Non-receptor Tyrosine Kinases in Cancer Development and Treatment

**11**

**Chapter 2**

**Abstract**

targeted therapies.

**1. Introduction**

threonine kinases [1].

forming a circuitry of regulation.

Non-receptor Tyrosine Kinases

This chapter presents a review about non-receptor tyrosine kinases, their structure, mechanisms of action and physiopathology, and how they are regulated and interact with other molecules and other signaling pathways, contributing to the regulation of fundamental cellular functions such as cell division and differentiation, stress responses, apoptosis, survival, and proliferation, gene expression, immune response, inter alia. Special emphasis will be assigned to the JAK family, the processes whereby it can be mutated/regulated and aberrantly activated, clinical significance and association with hematological disease progression and malignancy, mainly in myeloproliferative neoplasms. Consideration of these mechanisms may have important implications for selection of anti-cancer

**Keywords:** tyrosine kinase, non-receptor, JAK, mutation, driver mutations,

The existence and homeostasis of all living multicellular organisms depend on the existence of critical links established by several complex signaling pathways

The development of the Human Genome Project was crucial for the knowledge of the protein kinase, responsible for phosphorylation of other molecules, mostly proteins which can be grouped in two main classes, tyrosine kinases and serine-

Tyrosine kinases (TKs) are a family of more than 90 enzymes that act as fundamental mediators of all signal transduction processes, contributing to a variety of biological mechanisms in response to internal and external triggers, modulating cellular growth, differentiation, migration, metabolism, apoptosis, and survival [2, 3]. Though their activity is very well regulated in normal cells, recent studies have implicated TKs in human neoplastic disorder development and progression, including hematological malignancies [4], assuming a dominant oncoprotein status, either by acquiring transforming functions due to mutations by enhanced expression or by autocrine paracrine stimulation [2, 3]. These mechanisms of abnormal activation of TKs led to important efforts in the development of newly target-

directed molecules for cancer therapy as selective TK inhibitors [2–6].

myeloproliferative, malignancy, drug resistance

Role and Significance in

*Ana Azevedo, Susana Silva and José Rueff*

Hematological Malignancies

#### **Chapter 2**

## Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies

*Ana Azevedo, Susana Silva and José Rueff*

#### **Abstract**

This chapter presents a review about non-receptor tyrosine kinases, their structure, mechanisms of action and physiopathology, and how they are regulated and interact with other molecules and other signaling pathways, contributing to the regulation of fundamental cellular functions such as cell division and differentiation, stress responses, apoptosis, survival, and proliferation, gene expression, immune response, inter alia. Special emphasis will be assigned to the JAK family, the processes whereby it can be mutated/regulated and aberrantly activated, clinical significance and association with hematological disease progression and malignancy, mainly in myeloproliferative neoplasms. Consideration of these mechanisms may have important implications for selection of anti-cancer targeted therapies.

**Keywords:** tyrosine kinase, non-receptor, JAK, mutation, driver mutations, myeloproliferative, malignancy, drug resistance

#### **1. Introduction**

The existence and homeostasis of all living multicellular organisms depend on the existence of critical links established by several complex signaling pathways forming a circuitry of regulation.

The development of the Human Genome Project was crucial for the knowledge of the protein kinase, responsible for phosphorylation of other molecules, mostly proteins which can be grouped in two main classes, tyrosine kinases and serinethreonine kinases [1].

Tyrosine kinases (TKs) are a family of more than 90 enzymes that act as fundamental mediators of all signal transduction processes, contributing to a variety of biological mechanisms in response to internal and external triggers, modulating cellular growth, differentiation, migration, metabolism, apoptosis, and survival [2, 3]. Though their activity is very well regulated in normal cells, recent studies have implicated TKs in human neoplastic disorder development and progression, including hematological malignancies [4], assuming a dominant oncoprotein status, either by acquiring transforming functions due to mutations by enhanced expression or by autocrine paracrine stimulation [2, 3]. These mechanisms of abnormal activation of TKs led to important efforts in the development of newly targetdirected molecules for cancer therapy as selective TK inhibitors [2–6].

Tyrosine kinases are responsible for the selective phosphorylation of tyrosine residues in specific target protein substrates, using ATP, thus allowing transmission of signals from the cellular surface to cytoplasmic proteins and the nucleus, to regulate physiological circuits [2, 3, 5]. They can be further subdivided into two groups, receptor proteins and non-receptor proteins (which will be discussed below).

Briefly, receptor tyrosine kinases (RTKs) include several families, namely, epidermal growth factor receptor (EGFR), insulin receptor (IR), fibroblast growth factor receptor (FGFR), and platelet-derived growth factor receptors (PDGFR). They function as transducers of extracellular signals to cytoplasm and contain several domains, multiple extracellular ligand binding (e.g., EGF, PDGF, etc.) sites, a cytoplasmic portion with catalytic and regulation features, and a single transmembrane hydrophobic disulfide bond that links the two other regions [1, 5]. RTKs function as cell surface receptors, being activated by ligand binding to the extracellular domain, with subsequent dimerization of receptors and transphosphorylation in the cytoplasmic domain [5]. They constitute also enzymes with kinase activity, which are associated with altered gene expression, interfering with cellular division, migration, and survival functions [3].

Non-receptor tyrosine kinases (NRTKs) are organized into nine subfamilies based on sequence similarities, primarily within the kinase domains, and are able to regulate several cellular processes, such as cellular division, proliferation and survival, gene expression, and immune response, among others [3]. The role of their deregulation, genetic alterations, and abnormal activation in the development of hematological malignancies will be covered in this review.

Novel therapeutic compounds able to target kinases have been developed for the treatment of patients with this kind of disorders.

#### **2. Non-receptor tyrosine kinase families**

Non-receptor tyrosine kinases (NRTKs) are a subgroup of tyrosine kinases, intracellular cytoplasmic proteins, or anchored to the cell membrane, which can trigger intracellular signals derived from extracellular receptor [3]. They can be classified into nine subfamilies according to sequence similarities, primarily within the kinase domains. These include ABL, FES, JAK, ACK, SYK, TEC, FAK, SRC, and CSK family of kinases, which will be presented below in this section.

Unlike RTKs, NRTKs lack receptor-like features, such as an extracellular ligandbinding domain and a transmembrane-spanning domain, exhibiting considerable structural variability (**Figure 1**). They comprise a shared kinase domain, which spans approximately 300 residues and consists of an N-terminal portion (five stranded β-sheet and one α-helix), and a large cytoplasmic C-terminal domain (mainly α-helical). Moreover, they often possess several additional signaling or protein-protein interacting domains, such as SH2, SH3, and PH domains. The ATP molecule binds between the two domains, and the tyrosine sequence of the protein substrate links with the residues of the C terminal domain [5].

The activation of NRTKs involves several complex mechanisms of heterologous protein-protein interaction to enable cellular tyrosine kinase phosphorylation, highly regulated by antagonist effects of tyrosine kinase versus phosphatases, which results in the successive activation of specific signaling pathways and messenger proteins that regulate cellular functions, such as growth, division, and apoptosis [5].

In the last few years, it has been substantiated that NRTKs can suffer two types of oncogenic mutations, namely, intragenic point mutations, duplications, or deletions and insertions, or in addition chromosomal rearrangements may occur, resulting in the fusion of genes (e.g., most famously BCR-ABL), associated with

**13**

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies*

the development of hematological malignancies, either leukemia, lymphoma, or myeloma [3]. These mutations lead to aberrant kinase activation and signaling or a constitutive kinase activity, associated with the formation of oncogenes (or "driver mutations"), such as ABL, FES, SRC, and others, implicated in the process of hematopoiesis, contributing to cellular prolonged viability and survival [3]. Although some NRTK oncogenes exhibit structural, functional, and cellular localization differences, many of them share the same molecular pathways for cellular proliferation and viability regulation [3]. Later in this revision, we will focus the role of some NRTK families, mainly JAK, involved in the development of specific hematological malignancies, covering their associated genetic alterations and mutations, deregula-

*homology 2 domain; SH3, SRC homology 3 domain; SH4, SRC homology 4 domain.*

*Domain organization of the major non-receptor tyrosine kinase families (adapted from Siveen et al. [3]). Actin, actin-binding domain; Btk, Btk-type zinc finger motif; C, carboxy-terminus; CC, coiled coil motif; CRIB, Cdc42/Rac-interactive domain; DNA, DNA-binding domain; FAT, focal adhesion targeting domain; FCH, FES/Fer/Cdc-42 interactive protein homology domain; FERM, four-point-one, ezrin, radixin, moesin domain; JH2, Janus homology domain 2 (or pseudokinase domain); kinase, catalytic kinase domain (or SH1 domain); N, amino terminus; PH, pleckstrin homology domain; pr, proline-rich region; SH2, SRC* 

Recent advances have also been made in the development of specific kinase inhibitors and new therapies in order to target mutated kinases and inhibit their

While BCR-ABL occurs exclusively in leukemia, many of the subsequently discovered tyrosine kinase fusions occur in multiple tumor types, including both

The Abelson (ABL) kinase family includes ABL1 and ABL2 (ABL-related gene, ARG) proteins, which are ubiquitously expressed and necessary for normal cellular

NRTKs play a crucial role in several cellular mechanisms. Some examples are the involvement of JAK family in cell signaling, through activation of signal transducers and activators of transcription (STAT); the role in cellular growth of nuclear TKs (e.g., ABL), through activation of transcription factor Rb, and of ACKs via the induction of JAK and SRC; the regulation of cell adhesion and proliferation mediated by FAK; the association of Fyn and ACKs with signal transduction pathways and of TEC families with intracellular signaling processes; and the intervention of

activity, showing to be very effective and remarkably well tolerated [3].

*DOI: http://dx.doi.org/10.5772/intechopen.84873*

tion, and abnormal activation.

**Figure 1.**

SYK in immune response [3].

**2.1 ABL kinases**

liquid and solid malignancies [5].

function, encoded by ABL1 and ABL2 genes.

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies DOI: http://dx.doi.org/10.5772/intechopen.84873*

#### **Figure 1.**

*Tyrosine Kinases as Druggable Targets in Cancer*

sion, migration, and survival functions [3].

of hematological malignancies will be covered in this review.

treatment of patients with this kind of disorders.

**2. Non-receptor tyrosine kinase families**

Tyrosine kinases are responsible for the selective phosphorylation of tyrosine residues in specific target protein substrates, using ATP, thus allowing transmission of signals from the cellular surface to cytoplasmic proteins and the nucleus, to regulate physiological circuits [2, 3, 5]. They can be further subdivided into two groups, receptor proteins and non-receptor proteins (which will be discussed below). Briefly, receptor tyrosine kinases (RTKs) include several families, namely, epidermal growth factor receptor (EGFR), insulin receptor (IR), fibroblast growth factor receptor (FGFR), and platelet-derived growth factor receptors (PDGFR). They function as transducers of extracellular signals to cytoplasm and contain several domains, multiple extracellular ligand binding (e.g., EGF, PDGF, etc.) sites, a cytoplasmic portion with catalytic and regulation features, and a single transmembrane hydrophobic disulfide bond that links the two other regions [1, 5]. RTKs function as cell surface receptors, being activated by ligand binding to the extracellular domain, with subsequent dimerization of receptors and transphosphorylation in the cytoplasmic domain [5]. They constitute also enzymes with kinase activity, which are associated with altered gene expression, interfering with cellular divi-

Non-receptor tyrosine kinases (NRTKs) are organized into nine subfamilies based on sequence similarities, primarily within the kinase domains, and are able to regulate several cellular processes, such as cellular division, proliferation and survival, gene expression, and immune response, among others [3]. The role of their deregulation, genetic alterations, and abnormal activation in the development

Novel therapeutic compounds able to target kinases have been developed for the

Non-receptor tyrosine kinases (NRTKs) are a subgroup of tyrosine kinases, intracellular cytoplasmic proteins, or anchored to the cell membrane, which can trigger intracellular signals derived from extracellular receptor [3]. They can be classified into nine subfamilies according to sequence similarities, primarily within the kinase domains. These include ABL, FES, JAK, ACK, SYK, TEC, FAK, SRC, and

Unlike RTKs, NRTKs lack receptor-like features, such as an extracellular ligandbinding domain and a transmembrane-spanning domain, exhibiting considerable structural variability (**Figure 1**). They comprise a shared kinase domain, which spans approximately 300 residues and consists of an N-terminal portion (five stranded β-sheet and one α-helix), and a large cytoplasmic C-terminal domain (mainly α-helical). Moreover, they often possess several additional signaling or protein-protein interacting domains, such as SH2, SH3, and PH domains. The ATP molecule binds between the two domains, and the tyrosine sequence of the protein

The activation of NRTKs involves several complex mechanisms of heterologous

protein-protein interaction to enable cellular tyrosine kinase phosphorylation, highly regulated by antagonist effects of tyrosine kinase versus phosphatases, which results in the successive activation of specific signaling pathways and messenger proteins that regulate cellular functions, such as growth, division, and apoptosis [5]. In the last few years, it has been substantiated that NRTKs can suffer two types

of oncogenic mutations, namely, intragenic point mutations, duplications, or deletions and insertions, or in addition chromosomal rearrangements may occur, resulting in the fusion of genes (e.g., most famously BCR-ABL), associated with

CSK family of kinases, which will be presented below in this section.

substrate links with the residues of the C terminal domain [5].

**12**

*Domain organization of the major non-receptor tyrosine kinase families (adapted from Siveen et al. [3]). Actin, actin-binding domain; Btk, Btk-type zinc finger motif; C, carboxy-terminus; CC, coiled coil motif; CRIB, Cdc42/Rac-interactive domain; DNA, DNA-binding domain; FAT, focal adhesion targeting domain; FCH, FES/Fer/Cdc-42 interactive protein homology domain; FERM, four-point-one, ezrin, radixin, moesin domain; JH2, Janus homology domain 2 (or pseudokinase domain); kinase, catalytic kinase domain (or SH1 domain); N, amino terminus; PH, pleckstrin homology domain; pr, proline-rich region; SH2, SRC homology 2 domain; SH3, SRC homology 3 domain; SH4, SRC homology 4 domain.*

the development of hematological malignancies, either leukemia, lymphoma, or myeloma [3]. These mutations lead to aberrant kinase activation and signaling or a constitutive kinase activity, associated with the formation of oncogenes (or "driver mutations"), such as ABL, FES, SRC, and others, implicated in the process of hematopoiesis, contributing to cellular prolonged viability and survival [3]. Although some NRTK oncogenes exhibit structural, functional, and cellular localization differences, many of them share the same molecular pathways for cellular proliferation and viability regulation [3]. Later in this revision, we will focus the role of some NRTK families, mainly JAK, involved in the development of specific hematological malignancies, covering their associated genetic alterations and mutations, deregulation, and abnormal activation.

Recent advances have also been made in the development of specific kinase inhibitors and new therapies in order to target mutated kinases and inhibit their activity, showing to be very effective and remarkably well tolerated [3].

NRTKs play a crucial role in several cellular mechanisms. Some examples are the involvement of JAK family in cell signaling, through activation of signal transducers and activators of transcription (STAT); the role in cellular growth of nuclear TKs (e.g., ABL), through activation of transcription factor Rb, and of ACKs via the induction of JAK and SRC; the regulation of cell adhesion and proliferation mediated by FAK; the association of Fyn and ACKs with signal transduction pathways and of TEC families with intracellular signaling processes; and the intervention of SYK in immune response [3].

While BCR-ABL occurs exclusively in leukemia, many of the subsequently discovered tyrosine kinase fusions occur in multiple tumor types, including both liquid and solid malignancies [5].

#### **2.1 ABL kinases**

The Abelson (ABL) kinase family includes ABL1 and ABL2 (ABL-related gene, ARG) proteins, which are ubiquitously expressed and necessary for normal cellular function, encoded by ABL1 and ABL2 genes.

ABL family is involved in the regulation of several cellular mechanisms, namely, proliferation, migration, invasion and adhesion, reaction to DNA lesion and stress, and survival, through the interaction of distinct extracellular stimuli with specific signaling pathways [7]. Several growth factors, such as PDGF, EGFR, transforming growth factor β, and angiotensin subtype 1 receptors, are responsible for the activation of cytoplasmic c-ABL [8].

The identification of the fusion oncoprotein BCR-ABL1, which results from the translocation leading to the Philadelphia chromosome (Ph), by the American geneticist Janet Rowley (1925–2013) in 1972, formed by the reciprocal translocation between chromosomes 9 and 22 (t(9;22)(q34.1;q11.2)), and in 1985–1986, the knowledge of the *BCR-ABL1* transcript and its P210 fusion protein product, reinforced the role of ABL family in malignant disorders, especially hematological, such as acute myeloid leukemia (AML), chronic myeloid leukemia (CML), and acute lymphoblastic leukemia (ALL). The translocation of the breakpoint cluster region (BCR) sequences of chromosome 22 with the c-ABL tyrosine kinase of chromosome 9 gives origin to a fusion gene, responsible for the production of three oncoproteins. The BCR-ABL chimeric gene product has an enhanced tyrosine kinase activity, contributing to disease phenotype [2].

In 1996, in the era of the Human Genome Project development, these discoveries led Nicholas Lydon (b.1957), a British scientist, and Brian Druker (b. 1955), an American physician scientist, to the elaboration and therapeutic use of imatinib (a tyrosine kinase inhibitor) in CML [9].

The several products of malignant ABL fusion gene result in constitutively activated ABL kinases that can lead to cellular transformation and cancer. Activation of ABL kinases due to chromosome translocation is very rare in solid neoplasms, but usually there is overexpression, upstream oncogenic TKs or other chemokine receptors, inactivation of negative regulatory proteins, and/or oxidative stress [3].

There is a large number of signaling pathways that are activated by BCR-ABL, but those critical for BCR-ABL-dependent transformation include Gab2, Myc, CrkL, and STAT5 [3].

The first human malignancy to be associated to a specific genetic abnormality was chronic myelogenous leukemia, a clonal bone marrow stem cell malignancy, which accounts for 15–20% of adult leukemia's with a frequency of 1–2 cases per 100,000 individuals. It is more common in men and is rarely seen in children.

The formation of constitutively active chimeric BCR-ABL1 fusion oncoproteins leads to the creation of three distinct BCR-ABL variants, namely, p185, p210, and p230. The most common variant in CML is p210, in which the first exon of c-ABL has been replaced by BCR sequences, encoding either 927 or 902 amino acid, observed in hematopoietic cells of CML-stabilized patients, and in ALL and AML [3]. The p230 form is associated with acute leukemias, neutrophilic-CML, and rare cases of CML. The p185 form, containing BCR sequences from exon 1 fused to exons 2–11 of c-ABL, is found in about 20–30% of adults and about 3–5% of children with B-cell ALL [3].

BCR-ABL is the most common chromosomal translocation, but several other chromosomal abnormalities result in the expression of various fusion proteins, yet there are no activating point mutations identified in the ABL1/ABL2 genes [3].

BCR-ABL oncoprotein is the most frequent genetic defect found in adult ALL patients. Nearly 3–5% childhood and 25–40% adult cases of ALL have Philadelphia chromosome, associated with an aggressive phenotype and a worst prognosis [3].

The identification of BCR-ABL expression as the determinant leukemogenic event in CML and the use of BCR-ABL tyrosine kinase inhibitors (TKIs) since 2001 have changed the course of the disease and the management of patients, leading to a reduction in mortality rates and a consequent increase in the estimated prevalence of this disorder [10].

**15**

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies*

Imatinib mesylate, also known as STI571, was initially the standard of care for the first-line treatment of CML patients in chronic phase, due to its high long-term response rates and favorable tolerability profile compared with previous standard therapies [10]. The majority of kinase inhibitors are currently in clinical use to target BCR-ABL [11]. Imatinib is an ATP-competitive inhibitor that works by stabilizing the inactive ABL kinase domain conformation. Combining imatinib mesylate with standard chemotherapy also increases the overall long-term disease-

Approximately 15–30% (2–4% annually) of patients treated with imatinib discontinues treatment after 6 years due to resistance or intolerance, particularly in the accelerated and blast phase [10]. Nilotinib, dasatinib, bosutinib, and ponatinib

A literature review shows that pre-existing mutations at baseline confer a more aggressive disease phenotype and patients with advanced stages of the disease often

The role played by efflux ABC transporters in resistance to TKI in CML has deserved studies indicating its possible major role in drug resistance, besides the acquisition of mutations in the fusion leading to inefficacity of the TKI [12–14].

Feline sarcoma (FES) and FES-related (FER) proteins are proteins included in another group of NRTKs, called FES kinase family. These kinases are homologous to viral oncogenes responsible for cancerous transformation, namely, feline v-FES

Fer is ubiquitously expressed, while FES is a proto-oncogene expressed mostly in

myeloid hematopoietic, neuronal, epithelial, and vascular endothelial cells. There is recent evidence that both kinases are activated in AML blasts and regulate vital functions related with internal tandem duplication containing FLT3. FES is associated with phosphorylation/activation of STAT family, with signaling proteins such as phosphatidylinositol-4,5-bisphosphate 3-kinase, mitogen-activated protein kinases, and extracellular signal-regulated kinases and with signaling of the mutated oncogenic KIT receptor [15]. It is involved in several cellular mechanisms such as migration, survival and immune response, myeloid differentiation, and angiogenesis, through interaction with multiple cell surface growth factors and cytokine receptors (e.g., IL3, IL4, and GM-CSF receptors) [3]. Fer kinase partici-

FES kinases consist of a unique amino-terminal FCH (FES/FER/CDC-42-interacting protein homology) domain, three coiled coil motifs that promote oligomerization, a central SH2 domain for protein interactions, and a kinase domain in the carboxy-terminal region. FCH domain together with the first coiled coil motif corresponds to FCH-Bin-Amphiphysin-Rvs (F-BAR) domain (**Figure 1**) [16]. Although there is no negative regulatory SH3 domain, the catalytically repressed state of FES is strongly regulated through a tight interaction between SH2 and

Activation of FES kinase requires active phosphorylation of Tyr713 located inside the activation loop and of Tyr 811. Hyperactivation of FES kinase is necessary for deregulated proliferation in human lymphoid malignancies, but aberrant

Four somatic mutations within the kinase domain of FES were identified in colorectal cancers, and Fer mutations have been associated to small-cell lung

activation is not associated with human tumors [17].

are second-generation TKIs used for imatinib mesylate-resistant cases.

(Feline sarcoma) and avian v-fps (Fujinami poultry sarcoma).

*DOI: http://dx.doi.org/10.5772/intechopen.84873*

free survival in both adults and children [3].

do not respond to therapy or relapse [10].

**2.2 Feline sarcoma (FES) kinases**

pates in cell cycle progression.

kinase domain.

cancer [3].

#### *Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies DOI: http://dx.doi.org/10.5772/intechopen.84873*

Imatinib mesylate, also known as STI571, was initially the standard of care for the first-line treatment of CML patients in chronic phase, due to its high long-term response rates and favorable tolerability profile compared with previous standard therapies [10]. The majority of kinase inhibitors are currently in clinical use to target BCR-ABL [11]. Imatinib is an ATP-competitive inhibitor that works by stabilizing the inactive ABL kinase domain conformation. Combining imatinib mesylate with standard chemotherapy also increases the overall long-term diseasefree survival in both adults and children [3].

Approximately 15–30% (2–4% annually) of patients treated with imatinib discontinues treatment after 6 years due to resistance or intolerance, particularly in the accelerated and blast phase [10]. Nilotinib, dasatinib, bosutinib, and ponatinib are second-generation TKIs used for imatinib mesylate-resistant cases.

A literature review shows that pre-existing mutations at baseline confer a more aggressive disease phenotype and patients with advanced stages of the disease often do not respond to therapy or relapse [10].

The role played by efflux ABC transporters in resistance to TKI in CML has deserved studies indicating its possible major role in drug resistance, besides the acquisition of mutations in the fusion leading to inefficacity of the TKI [12–14].

#### **2.2 Feline sarcoma (FES) kinases**

*Tyrosine Kinases as Druggable Targets in Cancer*

(a tyrosine kinase inhibitor) in CML [9].

CrkL, and STAT5 [3].

tion of cytoplasmic c-ABL [8].

ABL family is involved in the regulation of several cellular mechanisms, namely, proliferation, migration, invasion and adhesion, reaction to DNA lesion and stress, and survival, through the interaction of distinct extracellular stimuli with specific signaling pathways [7]. Several growth factors, such as PDGF, EGFR, transforming growth factor β, and angiotensin subtype 1 receptors, are responsible for the activa-

The identification of the fusion oncoprotein BCR-ABL1, which results from the translocation leading to the Philadelphia chromosome (Ph), by the American geneticist Janet Rowley (1925–2013) in 1972, formed by the reciprocal translocation between chromosomes 9 and 22 (t(9;22)(q34.1;q11.2)), and in 1985–1986, the knowledge of the *BCR-ABL1* transcript and its P210 fusion protein product, reinforced the role of ABL family in malignant disorders, especially hematological, such as acute myeloid leukemia (AML), chronic myeloid leukemia (CML), and acute lymphoblastic leukemia (ALL). The translocation of the breakpoint cluster region (BCR) sequences of chromosome 22 with the c-ABL tyrosine kinase of chromosome 9 gives origin to a fusion gene, responsible for the production of three oncoproteins. The BCR-ABL chimeric gene product has an enhanced tyrosine kinase activity, contributing to disease phenotype [2].

In 1996, in the era of the Human Genome Project development, these discoveries led Nicholas Lydon (b.1957), a British scientist, and Brian Druker (b. 1955), an American physician scientist, to the elaboration and therapeutic use of imatinib

The several products of malignant ABL fusion gene result in constitutively activated ABL kinases that can lead to cellular transformation and cancer. Activation of ABL kinases due to chromosome translocation is very rare in solid neoplasms, but usually there is overexpression, upstream oncogenic TKs or other chemokine receptors, inactivation of negative regulatory proteins, and/or oxidative stress [3].

There is a large number of signaling pathways that are activated by BCR-ABL, but those critical for BCR-ABL-dependent transformation include Gab2, Myc,

The first human malignancy to be associated to a specific genetic abnormality was chronic myelogenous leukemia, a clonal bone marrow stem cell malignancy, which accounts for 15–20% of adult leukemia's with a frequency of 1–2 cases per 100,000 individuals. It is more common in men and is rarely seen in children. The formation of constitutively active chimeric BCR-ABL1 fusion oncoproteins leads to the creation of three distinct BCR-ABL variants, namely, p185, p210, and p230. The most common variant in CML is p210, in which the first exon of c-ABL has been replaced by BCR sequences, encoding either 927 or 902 amino acid, observed in hematopoietic cells of CML-stabilized patients, and in ALL and AML [3]. The p230 form is associated with acute leukemias, neutrophilic-CML, and rare cases of CML. The p185 form, containing BCR sequences from exon 1 fused to exons 2–11 of c-ABL, is found in

BCR-ABL is the most common chromosomal translocation, but several other chromosomal abnormalities result in the expression of various fusion proteins, yet there are no activating point mutations identified in the ABL1/ABL2 genes [3]. BCR-ABL oncoprotein is the most frequent genetic defect found in adult ALL patients. Nearly 3–5% childhood and 25–40% adult cases of ALL have Philadelphia chromosome, associated with an aggressive phenotype and a worst prognosis [3]. The identification of BCR-ABL expression as the determinant leukemogenic event in CML and the use of BCR-ABL tyrosine kinase inhibitors (TKIs) since 2001 have changed the course of the disease and the management of patients, leading to a reduction in mortality rates and a consequent increase in the estimated prevalence

about 20–30% of adults and about 3–5% of children with B-cell ALL [3].

**14**

of this disorder [10].

Feline sarcoma (FES) and FES-related (FER) proteins are proteins included in another group of NRTKs, called FES kinase family. These kinases are homologous to viral oncogenes responsible for cancerous transformation, namely, feline v-FES (Feline sarcoma) and avian v-fps (Fujinami poultry sarcoma).

Fer is ubiquitously expressed, while FES is a proto-oncogene expressed mostly in myeloid hematopoietic, neuronal, epithelial, and vascular endothelial cells.

There is recent evidence that both kinases are activated in AML blasts and regulate vital functions related with internal tandem duplication containing FLT3. FES is associated with phosphorylation/activation of STAT family, with signaling proteins such as phosphatidylinositol-4,5-bisphosphate 3-kinase, mitogen-activated protein kinases, and extracellular signal-regulated kinases and with signaling of the mutated oncogenic KIT receptor [15]. It is involved in several cellular mechanisms such as migration, survival and immune response, myeloid differentiation, and angiogenesis, through interaction with multiple cell surface growth factors and cytokine receptors (e.g., IL3, IL4, and GM-CSF receptors) [3]. Fer kinase participates in cell cycle progression.

FES kinases consist of a unique amino-terminal FCH (FES/FER/CDC-42-interacting protein homology) domain, three coiled coil motifs that promote oligomerization, a central SH2 domain for protein interactions, and a kinase domain in the carboxy-terminal region. FCH domain together with the first coiled coil motif corresponds to FCH-Bin-Amphiphysin-Rvs (F-BAR) domain (**Figure 1**) [16]. Although there is no negative regulatory SH3 domain, the catalytically repressed state of FES is strongly regulated through a tight interaction between SH2 and kinase domain.

Activation of FES kinase requires active phosphorylation of Tyr713 located inside the activation loop and of Tyr 811. Hyperactivation of FES kinase is necessary for deregulated proliferation in human lymphoid malignancies, but aberrant activation is not associated with human tumors [17].

Four somatic mutations within the kinase domain of FES were identified in colorectal cancers, and Fer mutations have been associated to small-cell lung cancer [3].

#### **2.3 JAK kinases**

This family comprises four members, JAK1, JAK2, JAK3, and TYK2, originally named "just another kinase." They owe their name due to the similarity of kinase (JH1) and pseudokinase (JH2) symmetrical domains with Janus, the Roman god of two faces [18, 19]. TYK2 was the first family member to be identified by Krolewski in 1990, through libraries of complementary DNA from human T lymphocytes, while JAK1, JAK2, and JAK3 were identified using conserved motif clonation of the catalytic domain [18]. They comprise seven homologous JH domains organized into four regions: kinase (JH1), pseudokinase (JH2), FERM (four-point-one, ezrin, radixin, moesin, including the N-terminal JH7, JH6, JH5, and part of JH4), and SH2 like (JH3 and part of JH4) (**Figure 1**) [20]. The carboxy-terminal portion of these molecules includes the distinctive kinase domain (JH1) which is catalytically active and the catalytically inactive pseudokinase domain (JH2) which is felt to regulate the activity of JH1. The other amino-terminal JH domains, JH3–JH7, mediate association with receptors. FERM domain regulates the binding to the membraneproximal part of the cytokine receptors [21].

In humans, JAK1 gene is located on chromosome 1p31.3, JAK2 gene on 9p24, JAK3 gene on 19p13.1, and TYK2 gene on 19p13.2 [9].

JAK proteins interact with different intracellular domains of cytokine receptors (discussed below) and are present in a variety of cellular subtypes. Expression is ubiquitous for JAK1, JAK2, and TYK2 but restricted to hematopoietic cells for JAK3 [9].

Many malignancies, including hematological neoplasms, are associated with deregulated activation of JAK family members, through aberrant cytokine production via autocrine/paracrine processes, point mutations within JAKs, or any other oncogene upstream of signaling cascade (discussed below).

Several studies reported various JAK mutations, mostly point mutations, occurring in all members [22–24]. *JAK2* V617F is one of the most studied mutations affecting JAK family, strongly associated with myeloproliferative neoplasms, which will be discussed in the next section of this chapter, and Hodgkin lymphoma and primary mediastinal B-cell lymphoma [3]. Other mutations have been described, such as 1) JAK1 A634D, localized in the pseudokinase domain, affecting signaling functions (STAT5), in AML, and T-cell and B-cell ALL; 2) JAK3 point mutations associated with various T-cell leukemia/lymphomas, poor prognosis and clinical outcome in juvenile myelomonocytic leukemia, and acute megakaryoblastic leukemia; 3) TYK2 kinase mutations have been reported in T-cell ALL and promote cell survival via activation of STAT1 as well BCL2 upregulation [3].

#### **2.4 ACK kinases**

ACKs also known as activated Cdc42 kinases are the fundamental components of signal transduction pathways linked to non-receptor tyrosine kinases. There are seven different types of ACKs, namely, ACK1/TNK2, ACK2, DACK, TNK1, ARK1, DPR2, and KOS1 [25].

The majority of these kinases include both N-terminal and C-terminal domains followed by a SH3 domain along with CRIB, which makes them unique NTRKs, and finally a kinase domain (**Figure 1**) [25].

ACK1 (ACK, TNK2, or activated Cdc42 kinase) is one of the most studied and well-known members of the ACKs. It is a ubiquitous 140-kDa protein located on the chromosome 3q, with the presence of multiple structural domains for its functional diversity, including cell survival, migration, growth, and proliferation, via acting as an integral cytosolic signal transducer for the array of receptor tyrosine

**17**

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies*

kinases (MERTK, EGFR, PDGFR, IR, etc.) to different intracellular effectors which includes both cytosolic and nuclear, and for epigenetic negative regulation on tumor suppressors [26]. It has been linked to several forms of human cancers, including gastric, breast, ovarian, pancreatic, colorectal, head, and neck squamous cell carcinomas, osteosarcoma, hepatocellular carcinoma, and prostate cancers [26]. Mutations in ACK1/TNK2 gene are the main oncogenic cause for AML, atypical CML, and chronic myelomonocytic leukemia. TNK1 has both tumor-suppressing and oncogenic potential as it can mitigate the growth of tumor cells by downregulating Ras-Raf1-MAPK pathway, induce apoptosis through NF-κB inhibition, and activate cellular transformation and growth of neoplastic cells. TNK1 has oncogenic potential implicated in hematological carcinogenesis such as in AML and Hodgkin's

Spleen tyrosine kinase (SYK) is one of the important classes of soluble cytosolic NRPKs and was first cloned in porcine spleen cells, with high expression hematopoietic cells [3]. It is a 72-kDa protein, encoded by SYK gene located on chromosome 9q22 and is highest homologous to ZAP-70, formed by two highly conserved SH2 domains with N-terminal and one tyrosine kinase domain at C-terminal (**Figure 1**) [3]. Activation of SYK occurs with the intervention of C-type lectins and integrins and the downstream signaling cascade, including VAV family members, phospholipase Cγ isoforms, the regulatory subunits of phosphoinositide 3-kinases, and the SH2 domain-containing leukocyte protein family members (SLP76 and

The SYK family is important in immune response between cell receptors and intracellular signaling mechanisms, through phosphorylation of cytosolic domain of the immunoreceptor tyrosine-based activation motifs (ITAMs), resulting in the conformational changes and further activation of SYK and signal transduction to other downstream target/effector proteins [27]. Its stimulatory effect on various survival pathways/signaling molecules supports the crucial role that SYK family has in many forms of hematological malignancies [28]. On the other hand, they also have a tumor-suppressive effect in the disorders of nonimmune origin [29]. Progress can be made in the development of targeted effective

TEC kinase family is the second largest subclass of the NRTKs. It includes five members, namely, Bruton's tyrosine kinase (BTK), interleukin 2-inducible T-cell kinase (ITK/EMT/TSK), tyrosine-protein kinase (RLK/TXK), bone marrow tyrosine kinase on chromosome (BMX/ETK), and tyrosine kinase expressed in hepatocellular carcinoma (TEC) [30]. Their structure is characterized by the presence of an amino-terminal (PH) that can bind phosphoinositides, enabling the interaction between phosphotyrosine-mediated and phospholipid-mediated signaling pathways, and Btk-type zinc finger (BTK) motif followed by two domains, SH3

TEC proteins are expressed in hematopoietic cells and involved in cellular signaling pathways of cytokine receptors, RTKs, lymphocyte surface antigens, G-proteincoupled receptors, and integrins, contributing to cellular growth and maturation of blood cells [3]. For example, it has been shown that BTK mutations are associated with B lymphocytes and other relevant cells contributing to the tumor microenvironment (e.g., dendritic cells, macrophages, myeloid-derived suppressor cells, and

and SH2, and a carboxy-terminal kinase domain (**Figure 1**).

*DOI: http://dx.doi.org/10.5772/intechopen.84873*

**2.5 SYK kinases**

SLP65) [27].

therapy.

**2.6 TEC kinases**

lymphoma, which may open new targets for therapy [3].

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies DOI: http://dx.doi.org/10.5772/intechopen.84873*

kinases (MERTK, EGFR, PDGFR, IR, etc.) to different intracellular effectors which includes both cytosolic and nuclear, and for epigenetic negative regulation on tumor suppressors [26]. It has been linked to several forms of human cancers, including gastric, breast, ovarian, pancreatic, colorectal, head, and neck squamous cell carcinomas, osteosarcoma, hepatocellular carcinoma, and prostate cancers [26].

Mutations in ACK1/TNK2 gene are the main oncogenic cause for AML, atypical CML, and chronic myelomonocytic leukemia. TNK1 has both tumor-suppressing and oncogenic potential as it can mitigate the growth of tumor cells by downregulating Ras-Raf1-MAPK pathway, induce apoptosis through NF-κB inhibition, and activate cellular transformation and growth of neoplastic cells. TNK1 has oncogenic potential implicated in hematological carcinogenesis such as in AML and Hodgkin's lymphoma, which may open new targets for therapy [3].

#### **2.5 SYK kinases**

*Tyrosine Kinases as Druggable Targets in Cancer*

proximal part of the cytokine receptors [21].

etic cells for JAK3 [9].

**2.4 ACK kinases**

DPR2, and KOS1 [25].

finally a kinase domain (**Figure 1**) [25].

JAK3 gene on 19p13.1, and TYK2 gene on 19p13.2 [9].

oncogene upstream of signaling cascade (discussed below).

cell survival via activation of STAT1 as well BCL2 upregulation [3].

This family comprises four members, JAK1, JAK2, JAK3, and TYK2, originally named "just another kinase." They owe their name due to the similarity of kinase (JH1) and pseudokinase (JH2) symmetrical domains with Janus, the Roman god of two faces [18, 19]. TYK2 was the first family member to be identified by Krolewski in 1990, through libraries of complementary DNA from human T lymphocytes, while JAK1, JAK2, and JAK3 were identified using conserved motif clonation of the catalytic domain [18]. They comprise seven homologous JH domains organized into four regions: kinase (JH1), pseudokinase (JH2), FERM (four-point-one, ezrin, radixin, moesin, including the N-terminal JH7, JH6, JH5, and part of JH4), and SH2 like (JH3 and part of JH4) (**Figure 1**) [20]. The carboxy-terminal portion of these molecules includes the distinctive kinase domain (JH1) which is catalytically active and the catalytically inactive pseudokinase domain (JH2) which is felt to regulate the activity of JH1. The other amino-terminal JH domains, JH3–JH7, mediate association with receptors. FERM domain regulates the binding to the membrane-

In humans, JAK1 gene is located on chromosome 1p31.3, JAK2 gene on 9p24,

Many malignancies, including hematological neoplasms, are associated with deregulated activation of JAK family members, through aberrant cytokine production via autocrine/paracrine processes, point mutations within JAKs, or any other

Several studies reported various JAK mutations, mostly point mutations, occurring in all members [22–24]. *JAK2* V617F is one of the most studied mutations affecting JAK family, strongly associated with myeloproliferative neoplasms, which will be discussed in the next section of this chapter, and Hodgkin lymphoma and primary mediastinal B-cell lymphoma [3]. Other mutations have been described, such as 1) JAK1 A634D, localized in the pseudokinase domain, affecting signaling functions (STAT5), in AML, and T-cell and B-cell ALL; 2) JAK3 point mutations associated with various T-cell leukemia/lymphomas, poor prognosis and clinical outcome in juvenile myelomonocytic leukemia, and acute megakaryoblastic leukemia; 3) TYK2 kinase mutations have been reported in T-cell ALL and promote

ACKs also known as activated Cdc42 kinases are the fundamental components of signal transduction pathways linked to non-receptor tyrosine kinases. There are seven different types of ACKs, namely, ACK1/TNK2, ACK2, DACK, TNK1, ARK1,

The majority of these kinases include both N-terminal and C-terminal domains followed by a SH3 domain along with CRIB, which makes them unique NTRKs, and

ACK1 (ACK, TNK2, or activated Cdc42 kinase) is one of the most studied and

well-known members of the ACKs. It is a ubiquitous 140-kDa protein located on the chromosome 3q, with the presence of multiple structural domains for its functional diversity, including cell survival, migration, growth, and proliferation, via acting as an integral cytosolic signal transducer for the array of receptor tyrosine

JAK proteins interact with different intracellular domains of cytokine receptors (discussed below) and are present in a variety of cellular subtypes. Expression is ubiquitous for JAK1, JAK2, and TYK2 but restricted to hematopoi-

**2.3 JAK kinases**

**16**

Spleen tyrosine kinase (SYK) is one of the important classes of soluble cytosolic NRPKs and was first cloned in porcine spleen cells, with high expression hematopoietic cells [3]. It is a 72-kDa protein, encoded by SYK gene located on chromosome 9q22 and is highest homologous to ZAP-70, formed by two highly conserved SH2 domains with N-terminal and one tyrosine kinase domain at C-terminal (**Figure 1**) [3]. Activation of SYK occurs with the intervention of C-type lectins and integrins and the downstream signaling cascade, including VAV family members, phospholipase Cγ isoforms, the regulatory subunits of phosphoinositide 3-kinases, and the SH2 domain-containing leukocyte protein family members (SLP76 and SLP65) [27].

The SYK family is important in immune response between cell receptors and intracellular signaling mechanisms, through phosphorylation of cytosolic domain of the immunoreceptor tyrosine-based activation motifs (ITAMs), resulting in the conformational changes and further activation of SYK and signal transduction to other downstream target/effector proteins [27]. Its stimulatory effect on various survival pathways/signaling molecules supports the crucial role that SYK family has in many forms of hematological malignancies [28]. On the other hand, they also have a tumor-suppressive effect in the disorders of nonimmune origin [29]. Progress can be made in the development of targeted effective therapy.

#### **2.6 TEC kinases**

TEC kinase family is the second largest subclass of the NRTKs. It includes five members, namely, Bruton's tyrosine kinase (BTK), interleukin 2-inducible T-cell kinase (ITK/EMT/TSK), tyrosine-protein kinase (RLK/TXK), bone marrow tyrosine kinase on chromosome (BMX/ETK), and tyrosine kinase expressed in hepatocellular carcinoma (TEC) [30]. Their structure is characterized by the presence of an amino-terminal (PH) that can bind phosphoinositides, enabling the interaction between phosphotyrosine-mediated and phospholipid-mediated signaling pathways, and Btk-type zinc finger (BTK) motif followed by two domains, SH3 and SH2, and a carboxy-terminal kinase domain (**Figure 1**).

TEC proteins are expressed in hematopoietic cells and involved in cellular signaling pathways of cytokine receptors, RTKs, lymphocyte surface antigens, G-proteincoupled receptors, and integrins, contributing to cellular growth and maturation of blood cells [3]. For example, it has been shown that BTK mutations are associated with B lymphocytes and other relevant cells contributing to the tumor microenvironment (e.g., dendritic cells, macrophages, myeloid-derived suppressor cells, and

endothelial cells) development impairment [31, 32], increasing the need of innovative immunochemotherapies, such as BTK inhibitors (e.g., ibrutinib), which have improved disease control rates but, unfortunately, not survival [33].

BTK, ITK, and TXK are predominately expressed in bone marrow cells, whereas BMX and TEC even extend to normal somatic cells (e.g., cardiac endothelium) [3, 30]. BMX is expressed in myeloid lineage hematopoietic cells (e.g., granulocytes and monocytes), endothelial cells, and numerous types of oncologic disorders, having a preponderant role in cellular survival, differentiation and motility, and playing a key role in inflammation and cancer [30]. Furthermore, TEC is expressed in hematopoietic cells, namely, myeloid and lymphoid, B and T, lineages; is involved in the stabilization, signaling, and activation of lymphocytes [34]; and acts as a regulator of pluripotent stem cells, through the regulation of fibroblast growth factor-2 secretion, associated with tumorigenesis and hepatocellular carcinoma progression [3].

#### **2.7 Focal adhesion kinases**

FAK family includes two members, namely, the ubiquitously expressed focal adhesion kinase and the associated adhesion focal tyrosine kinase (Pyk2), which is expressed in the central nervous system and in hematopoietic cells.

FAK and Pyk2 share a domain structure that includes an N-terminal FERM domain, followed by a residue linker region, a central kinase domain, a residue proline-rich low complexity region, and a C-terminal focal adhesion targeting domain (**Figure 1**) [35].

FAKs are involved in cell propagation and adhesion and in cell to microenvironment communications [36]. They are associated with B-lymphoblastic leukemia and lymphoma cells but are usually absent in leukemias/lymphomas of T-cell origin and in myeloma [3]. These kinases are involved in regulation of cellular proliferation and migration, via response to extracellular stimuli. Interaction with growth factor leads to phosphorylation/activation of SRC kinase, which in turn is associated with various signaling pathways, and modulates proliferation and survival of tumor cells in AML and MDS patients [37]. FAK overexpression has been associated with leukemic cell migration from the marrow to the circulating compartment, drug resistance, and poor survival outcome [3].

#### **2.8 SRC kinases**

The SRC family of tyrosine kinases (SFKs) is membrane-associated NRTKs, acting as key mediators of signal transduction pathways and modulators of RTK activation, promoting mitogenesis. This class includes 11 related kinases: BLK, FGR, FYN, HCK, LCK, LYN, c-SRC, c-YES, YRK, FRK (also known as RAK) and Srm [38].

Their structure includes in the amino-terminal region a membrane-targeting myristoylated or palmitoylated SH4 domain; a specific domain of 50–70 residues different for each member of the family, trailed by SH3, SH2, and kinase domains; and a short carboxy-terminal tail with an auto-inhibitory phosphorylation site (**Figure 1**) [39, 40].

BLK, FGR, HCK, LCK, and LYN expression predominates in hematopoietic cells, whereas c-SRC, c-YES, YRK, and FYN are highly expressed ubiquitously in platelets, neurons, and some epithelial tissues; Srm is found in keratinocytes; and Frk is present primarily in the bladder, breast, brain, colon, and lymphoid cells [38, 39].

SFKs are involved in a wealth of cellular mechanisms, such as cell survival regulation, DNA synthesis and division, actin cytoskeleton rearrangements, and

**19**

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies*

motility, through a major role in a variety of cellular signaling pathways activated by several RTKs (PDGF-R, EGF-R, FGF-R, IGF1-R, and CSF-R) and G-proteincoupled receptors [3]. Catalytic activity is exercised upon phosphorylation of a critical residue (Tyr419) within the activation loop and of the auto-inhibitory site Tyr530 within the carboxy-terminal tail, forming a closed auto-inhibited inactive conformation via the association of the SH2, SH3, and kinase domains by intramolecular interactions. However, these interactions could be broken by mutations or specific cellular triggers that are able to disrupt the inactive confor-

There is evidence that SFKs are involved in cancer development, by several different mechanisms. They are implicated in the regulation of cell-cell adhesion, involving different molecules, such as p120-catenin protein, a substrate of SRC; on the other hand, particularly SRC might be involved in the activation of STAT (STAT3 and STAT5) transcription factors which regulate cytokine signaling in hematopoietic cells and regulation of RAS/RAF/MEK/ERK MAPK and VEGF pathways and apoptosis molecules, having a role in the progression of CML, AML, CLL, and ALL. SFKs such as focal adhesion kinase, paxillin, and p130CAS have been implicated in monitoring of signaling pathways mediated by integrin, whose functional alterations are associated with several tumor types [3, 41]. SFKs are also associated with the development and signaling of T and B cells, particularly LCK,

Activation of SFKs due to mutation or binding to activating partners such as growth factor receptors (HER2/NWU, PDGF, EGFR, and c-kit), adaptor proteins, and other NRTKs (focal adhesion kinase and Bcr-ABL) can be detected in several cancers [45]. However, oncogenic mutations are rarely observed in the progression of hematopoietic malignancies such as leukemia and lymphomas (AML, ALL, CML, Burkitt's lymphoma, etc.), which are especially the result of constitutive activation of SFKs and amplification of anti-apoptotic and oncogenic downstream signaling pathways [41]. Moreover, there is evidence that SFKs promote cancer cell resistance to chemotherapy, radiation, and targeted RTK therapies. For example, Lyn and Hck have demonstrated upregulation and interaction with the oncogenic BCR-ABL fusion protein in specimens from patients with advanced CML and ALL

Due to the importance of SFKs in cancer development, it has been considered that inhibition of these proteins in combination with standard therapies may

C-terminal SRC kinases (CSK) and CSK-homologous kinase (CHK) are the two members included in this family of NRTKs. CSK is a 50-kDa protein ubiquitously expressed in all cells, primarily present in cytosol, with an amino-terminal SH3 domain followed by a SH2 domain and a carboxy-terminal kinase domain (**Figure 1**). CSK protein has no site for the activation loop for autophosphorylation nor a transmembrane domain or any fatty acyl modifications. However, the mobility of CSK to the membrane is a critical step in the regulation of its own activity, so that it is achieved by means of numerous scaffolding proteins (caveolin-1, paxillin, Dab2,

Chk is mainly expressed in the brain, hematopoietic cells, colon tissue, and

The binding of SH2-kinase and SH2–SH3 linkers to the amino-terminal lobe of the kinase domain stabilizes the active conformation. CSKs function as the major endogenous negative regulators of SFKs, as a result of CSK phosphorylation of

who showed relapse after imatinib mesylate treatment [46, 47].

represent a great clinical potential in disease control [48].

VE-cadherin, IGF-1R, IR, LIME, and SIT1) [49].

*DOI: http://dx.doi.org/10.5772/intechopen.84873*

mation of SFKs [3].

LYN, and FYN [39, 42–44].

**2.9 C-terminal SRC kinases**

smooth muscle cells [3].

#### *Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies DOI: http://dx.doi.org/10.5772/intechopen.84873*

motility, through a major role in a variety of cellular signaling pathways activated by several RTKs (PDGF-R, EGF-R, FGF-R, IGF1-R, and CSF-R) and G-proteincoupled receptors [3]. Catalytic activity is exercised upon phosphorylation of a critical residue (Tyr419) within the activation loop and of the auto-inhibitory site Tyr530 within the carboxy-terminal tail, forming a closed auto-inhibited inactive conformation via the association of the SH2, SH3, and kinase domains by intramolecular interactions. However, these interactions could be broken by mutations or specific cellular triggers that are able to disrupt the inactive conformation of SFKs [3].

There is evidence that SFKs are involved in cancer development, by several different mechanisms. They are implicated in the regulation of cell-cell adhesion, involving different molecules, such as p120-catenin protein, a substrate of SRC; on the other hand, particularly SRC might be involved in the activation of STAT (STAT3 and STAT5) transcription factors which regulate cytokine signaling in hematopoietic cells and regulation of RAS/RAF/MEK/ERK MAPK and VEGF pathways and apoptosis molecules, having a role in the progression of CML, AML, CLL, and ALL. SFKs such as focal adhesion kinase, paxillin, and p130CAS have been implicated in monitoring of signaling pathways mediated by integrin, whose functional alterations are associated with several tumor types [3, 41]. SFKs are also associated with the development and signaling of T and B cells, particularly LCK, LYN, and FYN [39, 42–44].

Activation of SFKs due to mutation or binding to activating partners such as growth factor receptors (HER2/NWU, PDGF, EGFR, and c-kit), adaptor proteins, and other NRTKs (focal adhesion kinase and Bcr-ABL) can be detected in several cancers [45]. However, oncogenic mutations are rarely observed in the progression of hematopoietic malignancies such as leukemia and lymphomas (AML, ALL, CML, Burkitt's lymphoma, etc.), which are especially the result of constitutive activation of SFKs and amplification of anti-apoptotic and oncogenic downstream signaling pathways [41]. Moreover, there is evidence that SFKs promote cancer cell resistance to chemotherapy, radiation, and targeted RTK therapies. For example, Lyn and Hck have demonstrated upregulation and interaction with the oncogenic BCR-ABL fusion protein in specimens from patients with advanced CML and ALL who showed relapse after imatinib mesylate treatment [46, 47].

Due to the importance of SFKs in cancer development, it has been considered that inhibition of these proteins in combination with standard therapies may represent a great clinical potential in disease control [48].

#### **2.9 C-terminal SRC kinases**

C-terminal SRC kinases (CSK) and CSK-homologous kinase (CHK) are the two members included in this family of NRTKs. CSK is a 50-kDa protein ubiquitously expressed in all cells, primarily present in cytosol, with an amino-terminal SH3 domain followed by a SH2 domain and a carboxy-terminal kinase domain (**Figure 1**). CSK protein has no site for the activation loop for autophosphorylation nor a transmembrane domain or any fatty acyl modifications. However, the mobility of CSK to the membrane is a critical step in the regulation of its own activity, so that it is achieved by means of numerous scaffolding proteins (caveolin-1, paxillin, Dab2, VE-cadherin, IGF-1R, IR, LIME, and SIT1) [49].

Chk is mainly expressed in the brain, hematopoietic cells, colon tissue, and smooth muscle cells [3].

The binding of SH2-kinase and SH2–SH3 linkers to the amino-terminal lobe of the kinase domain stabilizes the active conformation. CSKs function as the major endogenous negative regulators of SFKs, as a result of CSK phosphorylation of

*Tyrosine Kinases as Druggable Targets in Cancer*

progression [3].

**2.7 Focal adhesion kinases**

domain (**Figure 1**) [35].

**2.8 SRC kinases**

(**Figure 1**) [39, 40].

drug resistance, and poor survival outcome [3].

endothelial cells) development impairment [31, 32], increasing the need of innovative immunochemotherapies, such as BTK inhibitors (e.g., ibrutinib), which have

BMX and TEC even extend to normal somatic cells (e.g., cardiac endothelium) [3, 30]. BMX is expressed in myeloid lineage hematopoietic cells (e.g., granulocytes and monocytes), endothelial cells, and numerous types of oncologic disorders, having a preponderant role in cellular survival, differentiation and motility, and playing a key role in inflammation and cancer [30]. Furthermore, TEC is expressed in hematopoietic cells, namely, myeloid and lymphoid, B and T, lineages; is involved in the stabilization, signaling, and activation of lymphocytes [34]; and acts as a regulator of pluripotent stem cells, through the regulation of fibroblast growth factor-2 secretion, associated with tumorigenesis and hepatocellular carcinoma

FAK family includes two members, namely, the ubiquitously expressed focal adhesion kinase and the associated adhesion focal tyrosine kinase (Pyk2), which is

FAK and Pyk2 share a domain structure that includes an N-terminal FERM domain, followed by a residue linker region, a central kinase domain, a residue proline-rich low complexity region, and a C-terminal focal adhesion targeting

FAKs are involved in cell propagation and adhesion and in cell to microenvironment communications [36]. They are associated with B-lymphoblastic leukemia and lymphoma cells but are usually absent in leukemias/lymphomas of T-cell origin and in myeloma [3]. These kinases are involved in regulation of cellular proliferation and migration, via response to extracellular stimuli. Interaction with growth factor leads to phosphorylation/activation of SRC kinase, which in turn is associated with various signaling pathways, and modulates proliferation and survival of tumor cells in AML and MDS patients [37]. FAK overexpression has been associated with leukemic cell migration from the marrow to the circulating compartment,

The SRC family of tyrosine kinases (SFKs) is membrane-associated NRTKs, acting as key mediators of signal transduction pathways and modulators of RTK activation, promoting mitogenesis. This class includes 11 related kinases: BLK, FGR, FYN, HCK, LCK, LYN, c-SRC, c-YES, YRK, FRK (also known as RAK) and Srm [38]. Their structure includes in the amino-terminal region a membrane-targeting myristoylated or palmitoylated SH4 domain; a specific domain of 50–70 residues different for each member of the family, trailed by SH3, SH2, and kinase domains; and a short carboxy-terminal tail with an auto-inhibitory phosphorylation site

BLK, FGR, HCK, LCK, and LYN expression predominates in hematopoietic cells, whereas c-SRC, c-YES, YRK, and FYN are highly expressed ubiquitously in platelets, neurons, and some epithelial tissues; Srm is found in keratinocytes; and Frk is present primarily in the bladder, breast, brain, colon, and lymphoid cells [38,

SFKs are involved in a wealth of cellular mechanisms, such as cell survival regulation, DNA synthesis and division, actin cytoskeleton rearrangements, and

expressed in the central nervous system and in hematopoietic cells.

BTK, ITK, and TXK are predominately expressed in bone marrow cells, whereas

improved disease control rates but, unfortunately, not survival [33].

**18**

39].

the auto-inhibitory tyrosine residues in the SRC family kinase's C-terminal tail. Although its physiological importance is not known, several other signaling proteins such as paxillin, P2X3 receptor, c-Jun, and Lats can also serve as substrates of CSK [3].

These proteins have a critical role in the regulation of cell functions, such as growth, migration, differentiation, and immune response. Recent studies suggest that CSK can have a function as tumor suppressor through the inhibition of SFK oncogenic activity [3].

#### **3. Myeloproliferative neoplasms and their association with non-receptor tyrosine kinase families**

Myeloproliferative neoplasms (MPNs) are clonal hematopoietic malignancies resulting from the transformation of hematopoietic stem cells, leading to abnormal amplification of physiological signal transduction pathways and proliferation of one or more myeloid lineages. The *Word Health Organization (WHO) Classification of Tumours of Haematopoietic and Lymphoid Tissues* classified MPNs as chronic myeloid leukemia (CML), polycythemia vera (PV), essential thrombocythemia (ET), primary myelofibrosis (PMF) [50], chronic neutrophilic leukemia, and chronic eosinophilic leukemia not otherwise specified and MPNs unclassifiable [51]. In addition to primary (de novo), myelofibrosis can be secondary to PV (post-PV) or ET (post-ET) [52]. In the last revision of the WHO classification, in 2016, some changes were introduced, and mastocytosis ceased to be listed under the heading of MPNs [53].

Dameshek (1900–1969) was the first to conceptualize these groups of disorders, in 1951, highlighting the clinical and morphologic similarities between CML and Philadelphia-negative MPNs (PN-MPNs), namely, PV, ET, and PMF [54]. He realized that these disorders are caused by hyperproliferation in the bone marrow of more than one hematopoietic lineage, which proliferates "as a unit," and introduced the term "myeloproliferative disorders," indicating that these entities may correspond to a continuum of related syndromes. Moreover, he also postulated that the proliferative activity could be the result of a "hitherto undiscovered stimulus." However, the finding that bone marrow and peripheral blood cells from MPN patients can produce erythroid colonies in vitro without the stimulus of growth factor addition indicated the cell independent nature of these disorders [55].

But the "story" about MPNs had begun a few years before. Previously in 1845, John Hughes Bennett (1812–1875), an English pathologist working in Edinburgh, had described CML, and in 1879, a German surgeon, Gustav Heuck (1854–1940), underlined the morphological distinguishing features between PMF and CML, namely, the presence of bone marrow fibrosis, osteosclerosis, and extramedullary hematopoiesis in the former. Some years later in 1892, Louis Henri Vaquez (1860–1936), a French physician, was the first to describe PV, about a patient with marked erythrocytosis and hepatosplenomegaly, and in 1903 William Osler (1849–1919) took another step forward, distinguishing PV from both relative polycythemia and secondary polycythemia. The first description of ET is credited to Emil Epstein (1875–1951) and Alfred Goedel, two Austrian pathologists, who in 1934 published a case report of a "hemorrhagic thrombocythemia" in the absence of marked erythrocytosis.

In 1960, Peter Nowell (b. 1928) and David Hungerford (1927–1993), two American scientists working in Philadelphia, established the association between the Philadelphia (Ph) chromosome and CML [56], in contrast to PN-MPNs (PV, ET, and PMF).

**21**

(**Table 1**) [18].

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies*

X-linked glucose-6-phosphate dehydrogenase (G-6-PD) gene [9, 57–59].

poiesis, causing these organs to increase in size substantially.

Finally, the description of all four classic MPNs as clonal stem cell diseases was achieved by Philip Fialkow (1934–1996), an American physician scientist, through his studies developed between 1967 and 1981, on X chromosome inactivation patterns in women with PV, ET, PMF, and CML carrying a polymorphic variant of the

To better understand the pathophysiology of these disorders, the role of tyrosine kinases in all the process is crucial to elucidate some of the underlying mechanisms. Hematopoiesis is the process by which multipotent bone marrow-based stem cells (HSC) differentiate and mature into fully formed blood cells (namely, lymphoid, erythroid, megakaryocytes, and other myeloid cells), in response to external stimulus, such as erythropoietin (EPO), thrombopoietin (TPO), granulocytemacrophage colony-stimulating factor (GM-CSF), other stimulating growth factors, and several interleukins. Growth factors initiate signal transduction pathways (e.g., JAK-STAT pathway), which lead to the activation of transcription factors, and elicit different outcomes depending on the combination of factors and the cellular stage

In a healthy adult person, approximately 1011–1012 new blood cells are produced daily in order to maintain steady-state levels in the peripheral circulation. Besides bone marrow, in some cases and if necessary, the liver, thymus, and spleen may resume their hematopoietic function, in a process called extramedullary hemato-

Due to their essential roles as intracellular signaling effectors of hematopoietic cytokine receptor activation, the Janus kinase (JAK) family of tyrosine kinases have

JAK proteins (presented above) can link several intracellular domains of cyto-

Furthermore, a seven-member family of transcription factors named signal transducers and activators of transcription (STAT) are also involved in many cytokine signaling pathways. In 1994, Darnell and colleagues identified the first two members of the family, STAT1 and STAT2, by purification of factors linked to interferon (IFN)-stimulated genes, and the other family members were described subsequently [18]. These proteins act as transcriptional factors when they form homo- and heterodimers, among them, by phosphorylation at tyrosine residues in their SH2 domain, induced by upstream JAK proteins, activating different genes

The Janus kinase/signal transducers and activators for transcription (JAK/STAT) pathway regulate a large plethora of biological processes including cellular prolif-

All of these proteins are constitutively present in the cytoplasm without previous stimuli but can be quickly activated from the cellular membrane to the nucleus, by the binding of cytokines, growth factors, or hormones on cell surface receptors

Typically, Janus kinases function through their interaction with cytokine receptors that lack intrinsic kinase activity. Cytokines initiate signaling when ligand binding occurs (e.g., EPO, TPO) to the appropriate cytokine receptor (type 1 or type 2 cytokine receptors, e.g., EPO-R, MPL), which results in juxtaposition of JAKs, and bind to their specific cellular surface receptors, inducing several important conformational changes mainly oligomerization or multimerization of their receptors. JAK anchorage to the cytoplasmic domain of the cytokine receptor and phosphorylation of a tyrosine residue in the receptor follows, creating a docking site

aroused much interest since their discovery more than 20 years ago [60].

kine receptors and participate in a variety of cellular mechanisms [9].

and regulating downstream the JAK/STAT signaling pathway [18].

eration, differentiation, cell migration, and apoptosis [18].

*DOI: http://dx.doi.org/10.5772/intechopen.84873*

of differentiation.

**3.1 JAK-STAT signaling pathway**

#### *Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies DOI: http://dx.doi.org/10.5772/intechopen.84873*

Finally, the description of all four classic MPNs as clonal stem cell diseases was achieved by Philip Fialkow (1934–1996), an American physician scientist, through his studies developed between 1967 and 1981, on X chromosome inactivation patterns in women with PV, ET, PMF, and CML carrying a polymorphic variant of the X-linked glucose-6-phosphate dehydrogenase (G-6-PD) gene [9, 57–59].

To better understand the pathophysiology of these disorders, the role of tyrosine kinases in all the process is crucial to elucidate some of the underlying mechanisms.

Hematopoiesis is the process by which multipotent bone marrow-based stem cells (HSC) differentiate and mature into fully formed blood cells (namely, lymphoid, erythroid, megakaryocytes, and other myeloid cells), in response to external stimulus, such as erythropoietin (EPO), thrombopoietin (TPO), granulocytemacrophage colony-stimulating factor (GM-CSF), other stimulating growth factors, and several interleukins. Growth factors initiate signal transduction pathways (e.g., JAK-STAT pathway), which lead to the activation of transcription factors, and elicit different outcomes depending on the combination of factors and the cellular stage of differentiation.

In a healthy adult person, approximately 1011–1012 new blood cells are produced daily in order to maintain steady-state levels in the peripheral circulation. Besides bone marrow, in some cases and if necessary, the liver, thymus, and spleen may resume their hematopoietic function, in a process called extramedullary hematopoiesis, causing these organs to increase in size substantially.

#### **3.1 JAK-STAT signaling pathway**

*Tyrosine Kinases as Druggable Targets in Cancer*

CSK [3].

oncogenic activity [3].

these disorders [55].

of marked erythrocytosis.

**tyrosine kinase families**

the auto-inhibitory tyrosine residues in the SRC family kinase's C-terminal tail. Although its physiological importance is not known, several other signaling proteins such as paxillin, P2X3 receptor, c-Jun, and Lats can also serve as substrates of

These proteins have a critical role in the regulation of cell functions, such as growth, migration, differentiation, and immune response. Recent studies suggest that CSK can have a function as tumor suppressor through the inhibition of SFK

**3. Myeloproliferative neoplasms and their association with non-receptor** 

Myeloproliferative neoplasms (MPNs) are clonal hematopoietic malignancies resulting from the transformation of hematopoietic stem cells, leading to abnormal amplification of physiological signal transduction pathways and proliferation of one or more myeloid lineages. The *Word Health Organization (WHO) Classification of Tumours of Haematopoietic and Lymphoid Tissues* classified MPNs as chronic myeloid leukemia (CML), polycythemia vera (PV), essential thrombocythemia (ET), primary myelofibrosis (PMF) [50], chronic neutrophilic leukemia, and chronic eosinophilic leukemia not otherwise specified and MPNs unclassifiable [51]. In addition to primary (de novo), myelofibrosis can be secondary to PV (post-PV) or ET (post-ET) [52]. In the last revision of the WHO classification, in 2016, some changes were introduced, and mastocytosis ceased to be listed under the heading of MPNs [53]. Dameshek (1900–1969) was the first to conceptualize these groups of disorders, in 1951, highlighting the clinical and morphologic similarities between CML and Philadelphia-negative MPNs (PN-MPNs), namely, PV, ET, and PMF [54]. He realized that these disorders are caused by hyperproliferation in the bone marrow of more than one hematopoietic lineage, which proliferates "as a unit," and introduced the term "myeloproliferative disorders," indicating that these entities may correspond to a continuum of related syndromes. Moreover, he also postulated that the proliferative activity could be the result of a "hitherto undiscovered stimulus." However, the finding that bone marrow and peripheral blood cells from MPN patients can produce erythroid colonies in vitro without the stimulus of growth factor addition indicated the cell independent nature of

But the "story" about MPNs had begun a few years before. Previously in 1845, John Hughes Bennett (1812–1875), an English pathologist working in Edinburgh, had described CML, and in 1879, a German surgeon, Gustav Heuck (1854–1940), underlined the morphological distinguishing features between PMF and CML, namely, the presence of bone marrow fibrosis, osteosclerosis, and extramedullary hematopoiesis in the former. Some years later in 1892, Louis Henri Vaquez (1860–1936), a French physician, was the first to describe PV, about a patient with marked erythrocytosis and hepatosplenomegaly, and in 1903 William Osler (1849–1919) took another step forward, distinguishing PV from both relative polycythemia and secondary polycythemia. The first description of ET is credited to Emil Epstein (1875–1951) and Alfred Goedel, two Austrian pathologists, who in 1934 published a case report of a "hemorrhagic thrombocythemia" in the absence

In 1960, Peter Nowell (b. 1928) and David Hungerford (1927–1993), two American scientists working in Philadelphia, established the association between the Philadelphia (Ph) chromosome and CML [56], in contrast to PN-MPNs (PV, ET,

**20**

and PMF).

Due to their essential roles as intracellular signaling effectors of hematopoietic cytokine receptor activation, the Janus kinase (JAK) family of tyrosine kinases have aroused much interest since their discovery more than 20 years ago [60].

JAK proteins (presented above) can link several intracellular domains of cytokine receptors and participate in a variety of cellular mechanisms [9].

Furthermore, a seven-member family of transcription factors named signal transducers and activators of transcription (STAT) are also involved in many cytokine signaling pathways. In 1994, Darnell and colleagues identified the first two members of the family, STAT1 and STAT2, by purification of factors linked to interferon (IFN)-stimulated genes, and the other family members were described subsequently [18]. These proteins act as transcriptional factors when they form homo- and heterodimers, among them, by phosphorylation at tyrosine residues in their SH2 domain, induced by upstream JAK proteins, activating different genes and regulating downstream the JAK/STAT signaling pathway [18].

The Janus kinase/signal transducers and activators for transcription (JAK/STAT) pathway regulate a large plethora of biological processes including cellular proliferation, differentiation, cell migration, and apoptosis [18].

All of these proteins are constitutively present in the cytoplasm without previous stimuli but can be quickly activated from the cellular membrane to the nucleus, by the binding of cytokines, growth factors, or hormones on cell surface receptors (**Table 1**) [18].

Typically, Janus kinases function through their interaction with cytokine receptors that lack intrinsic kinase activity. Cytokines initiate signaling when ligand binding occurs (e.g., EPO, TPO) to the appropriate cytokine receptor (type 1 or type 2 cytokine receptors, e.g., EPO-R, MPL), which results in juxtaposition of JAKs, and bind to their specific cellular surface receptors, inducing several important conformational changes mainly oligomerization or multimerization of their receptors. JAK anchorage to the cytoplasmic domain of the cytokine receptor and phosphorylation of a tyrosine residue in the receptor follows, creating a docking site


#### **Table 1.**

*Cytokine and factor stimuli for JAK and STAT family activation.*

for the recruitment and activation of cytoplasmic signal transducers and activators of transcription (STATs: STAT3 and STAT5 in the case of JAK2, which is associated with PN-MPNs and will be taken as an example), through their SH2 domain. While STAT proteins are attached to the cytokine receptor, JAK proteins undergo autophosphorylation at a tyrosine residue, detaching the STAT protein from the cytokine receptor so that the STATs form homo- and heterodimers through their SH2 domain that will translocate to the nucleus. There, they bind to the promoter region of genes via specific DNA-binding domains to promote gene transcription.

The net result of STAT3 and STAT5 activation is apoptosis inhibition and a proliferative activity [61], playing an important role in growth factor-induced myeloid differentiation. STAT3 regulates cell growth through regulation of cyclins promoting cell cycle progression, as cyclin D1, and induces Bcl-2, resulting in an anti-apoptotic signal. Moreover, STAT3 may promote cellular differentiation by upregulating the expression and enhancing the transcriptional activity of CCAAT/enhancer-binding protein alpha (C/EBPα), a key transcription factor that drives myeloid differentiation [62]. STAT3 was also shown to play an important role in megakaryopoiesis, mainly through the expansion of megakaryocytic progenitor cells.

Normal differentiation of neutrophils, promoted by G-CSF, is disturbed by expression of a dominant negative form of STAT5. It has been suggested that STAT5 may induce the survival of myeloid progenitors via transcriptional upregulation of the anti-apoptotic protein BclxL and Pim kinase, inhibiting apoptosis of megakaryocytes, and mediates cell growth through induction of cyclin D1, thereby allowing myeloid differentiation to proceed [63].

EPO is secreted by interstitial kidney cells in response to reduction in blood oxygen concentration, transported to the bone marrow where it binds its receptor, EPO-R, and transmits an intercellular signal through a receptor conformational change, which stimulates an increased production of red blood cells [64–66]. The *JAK2* FERM domain constitutively binds to the EPO-R. EPO-induced EPO-R conformational change facilitates cross-phosphorylation and activation of the JAK2 proteins [67].

The amino-terminal extracellular TPO-R domain has a similar structure to EPO-R, which is critical in ligand binding, resulting in a significant overlap between

**23**

mas [60, 72].

**(PN-MPNs)**

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies*

JAK2 is involved in epigenetic modifications [18, 60, 71, 72].

(SOCS), and protein inhibitors of activated STATs [9]:

STAT recruitment to the binding sites [73, 74].

4.LNK sequesters JAK2 by direct binding [72].

cytokine receptors [75].

EPO- and TPO-stimulated pathways. As in EPO signaling, TPO stimulation causes the JAK2-dependent phosphorylation of STAT3 and STAT5, activation of the MAP kinase pathway, and activation of the PI3K/Akt survival pathway indirectly and can induce transcription of the pro-survival factor BclxL through STAT5- and PI3Kdependent pathways, promoting megakaryocyte differentiation. Overall, discovery of STAT, MAP kinase, and PI3K pathway stimulation downstream of the TPO-R gave a framework to understand the considerable overlap in phenotypic response to

JAK2 also serves as an endoplasmic reticulum chaperone for the EPO and TPO receptors, transporting them to the cell surface, and increases the total number of TPO receptors by stabilizing the mature form of the receptor, enhancing receptor recycling, and preventing receptor degradation [70]. On the other hand, nuclear

The JAK/STAT pathway is tightly regulated and inhibited at multiple levels by several protein families—tyrosine phosphatases, suppressors of cytokine signaling

1.SOCS, most notably SOCS1 and SOCS3, and CBL interact with activated JAKs and phosphorylated receptors or mark JAK for proteasomal degradation. CIS, SOCS1, SOCS2, and SOCS3 are members of the SOCS protein family. The synthesis of SOCS is induced by activated STATs resulting in a negative feedback loop, through interaction with activated JAKs and consequent inhibition of

2.Hematopoietic cells express SHP1. SHP1 belongs to the family of phosphotyrosine phosphatases (PTP); PTP dephosphorylates activated JAKs, STATs, and

3.Protein inhibitors of activated STATs (PIAS) interact with activated STATs, inhibit their dimerization, and prevent their binding to target DNA [72].

Mutations in all four JAKs have been associated with human diseases. Inherited

mutated JAK alleles lead to inactivated JAK3 and TYK2 in human immunodeficiency syndrome, while somatic mutations in JAK1, JAK2, and JAK3 result in constitutively active kinases in myeloproliferative diseases and leukemia/lympho-

A qualitative difference in the signaling state of STAT proteins has been described in PN-MPNs. ET progenitors have high phosphorylation levels of STAT1 and STAT5, whereas PV progenitors have only phosphorylated STAT5. The reasons behind this and other phenotypic differences are unclear but are potentially the result of a complex interplay between acquired and inherited variations, and pos-

sibly environmental exposure, all unique to each MPN patient [76].

**3.2 Philadelphia chromosome-negative myeloproliferative neoplasms** 

PN-MPNs (PV, ET, and PMF) are characterized by the clonal proliferation of one or more myeloid cell lineages (erythrocytic, granulocytic, or megakaryocytic), predominantly in the bone marrow, without altering the hematopoietic stem cell hierarchy, and involving JAK-STAT pathway. There is evidence of a normal and

*DOI: http://dx.doi.org/10.5772/intechopen.84873*

TPO and EPO [68, 69].

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies DOI: http://dx.doi.org/10.5772/intechopen.84873*

EPO- and TPO-stimulated pathways. As in EPO signaling, TPO stimulation causes the JAK2-dependent phosphorylation of STAT3 and STAT5, activation of the MAP kinase pathway, and activation of the PI3K/Akt survival pathway indirectly and can induce transcription of the pro-survival factor BclxL through STAT5- and PI3Kdependent pathways, promoting megakaryocyte differentiation. Overall, discovery of STAT, MAP kinase, and PI3K pathway stimulation downstream of the TPO-R gave a framework to understand the considerable overlap in phenotypic response to TPO and EPO [68, 69].

JAK2 also serves as an endoplasmic reticulum chaperone for the EPO and TPO receptors, transporting them to the cell surface, and increases the total number of TPO receptors by stabilizing the mature form of the receptor, enhancing receptor recycling, and preventing receptor degradation [70]. On the other hand, nuclear JAK2 is involved in epigenetic modifications [18, 60, 71, 72].

The JAK/STAT pathway is tightly regulated and inhibited at multiple levels by several protein families—tyrosine phosphatases, suppressors of cytokine signaling (SOCS), and protein inhibitors of activated STATs [9]:


4.LNK sequesters JAK2 by direct binding [72].

Mutations in all four JAKs have been associated with human diseases. Inherited mutated JAK alleles lead to inactivated JAK3 and TYK2 in human immunodeficiency syndrome, while somatic mutations in JAK1, JAK2, and JAK3 result in constitutively active kinases in myeloproliferative diseases and leukemia/lymphomas [60, 72].

A qualitative difference in the signaling state of STAT proteins has been described in PN-MPNs. ET progenitors have high phosphorylation levels of STAT1 and STAT5, whereas PV progenitors have only phosphorylated STAT5. The reasons behind this and other phenotypic differences are unclear but are potentially the result of a complex interplay between acquired and inherited variations, and possibly environmental exposure, all unique to each MPN patient [76].

#### **3.2 Philadelphia chromosome-negative myeloproliferative neoplasms (PN-MPNs)**

PN-MPNs (PV, ET, and PMF) are characterized by the clonal proliferation of one or more myeloid cell lineages (erythrocytic, granulocytic, or megakaryocytic), predominantly in the bone marrow, without altering the hematopoietic stem cell hierarchy, and involving JAK-STAT pathway. There is evidence of a normal and

*Tyrosine Kinases as Druggable Targets in Cancer*

JAK family

STAT family

**Table 1.**

**Cytokine or factor**

erythropoietin JAK3 IL-2, IL-7, IL-9, IL-15, IL-4

STAT2 IFN-α, IFN-β

STAT4 IL-12

STAT6 IL-4, IL-13

*Cytokine and factor stimuli for JAK and STAT family activation.*

STAT5 a/b

*Adapted from Becerra-Díaz et al. [18]*

TYK2 IL-6, I-11, IL-12, IL-13, CT-1, IFN-α, IFN-β, IL-10

STAT1 IL-2, IL-6, IL-10, IL-27, IFN-α, IFN-β, IFN-γ

STAT3 IL-6, IL-10, IL-27, LIF, growth hormone

for the recruitment and activation of cytoplasmic signal transducers and activators of transcription (STATs: STAT3 and STAT5 in the case of JAK2, which is associated with PN-MPNs and will be taken as an example), through their SH2 domain. While STAT proteins are attached to the cytokine receptor, JAK proteins undergo autophosphorylation at a tyrosine residue, detaching the STAT protein from the cytokine receptor so that the STATs form homo- and heterodimers through their SH2 domain that will translocate to the nucleus. There, they bind to the promoter region of genes via specific DNA-binding domains to promote gene transcription. The net result of STAT3 and STAT5 activation is apoptosis inhibition and a proliferative activity [61], playing an important role in growth factor-induced myeloid differentiation. STAT3 regulates cell growth through regulation of cyclins promoting cell cycle progression, as cyclin D1, and induces Bcl-2, resulting in an anti-apoptotic signal. Moreover, STAT3 may promote cellular differentiation by upregulating the expression and enhancing the transcriptional activity of CCAAT/enhancer-binding protein alpha (C/EBPα), a key transcription factor that drives myeloid differentiation [62]. STAT3 was also shown to play an important role in megakaryopoiesis,

Prolactin, growth hormone, thrombopoietin

JAK1 IL-2, IL-4, IL-6, IL-7, IL-9, IL-10, IL-11, IL-13, IL-15, IFN-α, IFN-β, IFN-γ, CT-1 JAK2 IL-3, IL-6, IL-11, IL-12, IL-13, IFN-γ, CT-1, growth hormone, prolactin,

mainly through the expansion of megakaryocytic progenitor cells.

myeloid differentiation to proceed [63].

Normal differentiation of neutrophils, promoted by G-CSF, is disturbed by expression of a dominant negative form of STAT5. It has been suggested that STAT5 may induce the survival of myeloid progenitors via transcriptional upregulation of the anti-apoptotic protein BclxL and Pim kinase, inhibiting apoptosis of megakaryocytes, and mediates cell growth through induction of cyclin D1, thereby allowing

EPO is secreted by interstitial kidney cells in response to reduction in blood oxygen concentration, transported to the bone marrow where it binds its receptor, EPO-R, and transmits an intercellular signal through a receptor conformational change, which stimulates an increased production of red blood cells [64–66]. The *JAK2* FERM domain constitutively binds to the EPO-R. EPO-induced EPO-R conformational change facilitates cross-phosphorylation and activation of the JAK2

The amino-terminal extracellular TPO-R domain has a similar structure to EPO-R, which is critical in ligand binding, resulting in a significant overlap between

**22**

proteins [67].

effective maturation, resulting in increased peripheral blood erythrocytes, granulocytes, and platelet counts [77].

Among the different PN-MPN entities, there is a frequent overlap of clinical, laboratory, and morphological data. Leukocytosis with neutrophilia, excessive megakaryocytic proliferation with thrombocytosis, myelofibrosis, and splenomegaly and hepatomegaly associated with the presence of extramedullary hematopoiesis can occur in any of these diseases.

PN-MPNs are considered as rare disorders, since their combined incidence is lower than 6 per 100,000 individuals per year [78]. Among the existent registries in the European Union, PN-MPNs have an annual incidence rate per 100,000 individuals per year ranging from 0.4 to 2.8 for PV (while the literature estimated 0.68–2.6), from 0.38 to 1.7 for ET (in the literature 0.6–2.5), and from 0.1 to 1.0 for PMF [79, 80]. There are few European studies reported on MPNs' prevalence [80]. However, according to the American data published in 2014, the prevalence per 100,000 individuals of PV (44–57) and ET (38–57) was much higher than that of MF (4–6) or subgroups with MF features (post-PV MF = 0.3–0.7; post-ET MF = 0.5–1.1) [81].

These groups of disorders occur in middle- or advanced-age adults, with a medium age of diagnosis of 65–67 years for PV, 65–70 years for ET, and 67–70 years for PMF [82]. However, it can be diagnosed in younger individuals, particularly if there is a familial predisposition [83]. Some reports indicate that ET is more common in women (particularly at younger ages) and PV in men, while in PMF both genders are nearly equally affected [51, 84, 85].

As demonstrated by European and international studies [86, 87], the distinction of MPNs in three nosological entities have a relevant prognostic significance. By and large, PN-MPN patients have a reduced life expectancy compared with general population, with PMF having the lowest overall survival (5.7 years), followed by PV with 15 years survival in 65% of cases and ET with an overall survival of more than 18–20 years [78, 88].

Despite insidious clinical onset, all PN-MPNs are at risk of clonal evolution and mortality. This is generally attributed to disease progression that may end in medullary failure (myelofibrosis or ineffective hematopoiesis) or transformation into other hematologic malignancies (the most common being acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS)) or the occurrence of bacterial infections and cardio- and cerebrovascular diseases, especially in younger patients [89, 90]. Fortunately, mortality due to these complications has been decreasing in the last few years [78].

#### *3.2.1 Driver genes and other mutations*

Until 2005 little was known about the etiology of PN-MPNs. The discovery of somatic mutations in Janus kinase 2 gene (*JAK2*), a member of the Janus kinase family located at chromosome 9 and first identified in 1993, was crucial. The identification of exon 14 V617F gain-of-function mutation, made by several independent groups of investigators [91–94], was one of the major genetic insights into the pathogenesis of the PN-MPNs and transformed the understanding of these disorders. It turned out to be the most important and most frequently recurring somatic mutation involved in PN-MPN pathogenesis, with the highest frequency (up to 95%) in PV, and 50–60% in ET and PMF patients (**Figure 2**) [9, 23, 55, 72, 95–99].

Although there is no gold standard and the choice of methodology is dependent on the application, quantitative real-time PCR is a useful method for detecting V617F mutation in *JAK2* gene [100].

**25**

**Figure 2.**

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies*

After *JAK2* V617F discovery in the majority of PN-MPN patients, there may have been an assumption of genetic uniformity, but the fact that approximately 50% of ET and PMF patients are *JAK2* V617F negative prompted the search for other putative genes in the JAK-STAT signaling pathway that could be mutated in these patients. In 2006, Pikman and colleagues [101] identified the mutations of thrombopoietin receptor (TPO-R) in myeloproliferative leukemia (MPL) virus oncogene. Moreover, a small proportion of patients with PV are *JAK2* V617F negative when tested by sensitive allele-specific assays [102], led only 1 year later, in 2007, to the identification by Scott and colleagues of a set of *JAK2* exon 12 mutations in *JAK2* V617F-negative patients with PV [103]. Although there is no gold standard and the choice of methodology is dependent on the application, quantitative real-time PCR and high-resolution melt-curve analysis are useful methods for detecting this type of mutation in *JAK2* gene [100]. One of the most recent discoveries was made by Kralovics in 2013, with the identification of calreticulin (CALR) mutation in 73% of MPN patients who do not bear the *JAK2* or *MPL* mutation (**Figure 2**) [106]. The identification of these other driver mutations (*JAK2* exon 12, *MPL*, and *CALR*) contributed to a better clarification of the pathophysiology of these disorders, their diagnostic tools, and therapeutic management [9, 91–94, 103, 107, 108]. In the majority of PN-MPN cases, *CALR, MPL*, and *JAK2* mutations are mutually exclusive, although rare exceptions can

It soon became clear that this group of diseases was far more genetically heterogeneous and complex than CML. Mutations other than in those driver genes and other genetic alterations have also been described in PN-MPNs and have shown to contribute to the establishment of the WHO diagnostic criteria, prognosis, and risk stratification in PN-MPNs [9, 90, 110, 111]. The majority of those mutations fall into one of the two categories—activation of the JAK-STAT pathway (*JAK2* V617F, *JAK2* exon 12, *MPL*, *LNK*, and probably *CALR*) [112] and aberrant epigenetic modification (*TET2*, *ASXL1*, and *EZH2*) [113]. A combination of mutations in these genes and environmental factors is likely the decisive factor of the development of

The receptors of bone marrow progenitor cells are highly sensitive to EPO (stimulates erythroblasts), TPO (induces proliferation and differentiation of

*Variation frequency of driver and other mutations in PN-MPNs [78, 104, 105].*

*DOI: http://dx.doi.org/10.5772/intechopen.84873*

occur [70, 109].

each one of these disorders.

*3.2.2 Molecular pathophysiology*

#### *Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies DOI: http://dx.doi.org/10.5772/intechopen.84873*

After *JAK2* V617F discovery in the majority of PN-MPN patients, there may have been an assumption of genetic uniformity, but the fact that approximately 50% of ET and PMF patients are *JAK2* V617F negative prompted the search for other putative genes in the JAK-STAT signaling pathway that could be mutated in these patients. In 2006, Pikman and colleagues [101] identified the mutations of thrombopoietin receptor (TPO-R) in myeloproliferative leukemia (MPL) virus oncogene. Moreover, a small proportion of patients with PV are *JAK2* V617F negative when tested by sensitive allele-specific assays [102], led only 1 year later, in 2007, to the identification by Scott and colleagues of a set of *JAK2* exon 12 mutations in *JAK2* V617F-negative patients with PV [103]. Although there is no gold standard and the choice of methodology is dependent on the application, quantitative real-time PCR and high-resolution melt-curve analysis are useful methods for detecting this type of mutation in *JAK2* gene [100].

One of the most recent discoveries was made by Kralovics in 2013, with the identification of calreticulin (CALR) mutation in 73% of MPN patients who do not bear the *JAK2* or *MPL* mutation (**Figure 2**) [106]. The identification of these other driver mutations (*JAK2* exon 12, *MPL*, and *CALR*) contributed to a better clarification of the pathophysiology of these disorders, their diagnostic tools, and therapeutic management [9, 91–94, 103, 107, 108]. In the majority of PN-MPN cases, *CALR, MPL*, and *JAK2* mutations are mutually exclusive, although rare exceptions can occur [70, 109].

It soon became clear that this group of diseases was far more genetically heterogeneous and complex than CML. Mutations other than in those driver genes and other genetic alterations have also been described in PN-MPNs and have shown to contribute to the establishment of the WHO diagnostic criteria, prognosis, and risk stratification in PN-MPNs [9, 90, 110, 111]. The majority of those mutations fall into one of the two categories—activation of the JAK-STAT pathway (*JAK2* V617F, *JAK2* exon 12, *MPL*, *LNK*, and probably *CALR*) [112] and aberrant epigenetic modification (*TET2*, *ASXL1*, and *EZH2*) [113]. A combination of mutations in these genes and environmental factors is likely the decisive factor of the development of each one of these disorders.

#### *3.2.2 Molecular pathophysiology*

The receptors of bone marrow progenitor cells are highly sensitive to EPO (stimulates erythroblasts), TPO (induces proliferation and differentiation of

*Variation frequency of driver and other mutations in PN-MPNs [78, 104, 105].*

*Tyrosine Kinases as Druggable Targets in Cancer*

poiesis can occur in any of these diseases.

genders are nearly equally affected [51, 84, 85].

cytes, and platelet counts [77].

MF = 0.5–1.1) [81].

18–20 years [78, 88].

the last few years [78].

*3.2.1 Driver genes and other mutations*

V617F mutation in *JAK2* gene [100].

effective maturation, resulting in increased peripheral blood erythrocytes, granulo-

Among the different PN-MPN entities, there is a frequent overlap of clinical, laboratory, and morphological data. Leukocytosis with neutrophilia, excessive megakaryocytic proliferation with thrombocytosis, myelofibrosis, and splenomegaly and hepatomegaly associated with the presence of extramedullary hemato-

PN-MPNs are considered as rare disorders, since their combined incidence is lower than 6 per 100,000 individuals per year [78]. Among the existent registries in the European Union, PN-MPNs have an annual incidence rate per 100,000 individuals per year ranging from 0.4 to 2.8 for PV (while the literature estimated 0.68–2.6), from 0.38 to 1.7 for ET (in the literature 0.6–2.5), and from 0.1 to 1.0 for PMF [79, 80]. There are few European studies reported on MPNs' prevalence [80]. However, according to the American data published in 2014, the prevalence per 100,000 individuals of PV (44–57) and ET (38–57) was much higher than that of MF (4–6) or subgroups with MF features (post-PV MF = 0.3–0.7; post-ET

These groups of disorders occur in middle- or advanced-age adults, with a medium age of diagnosis of 65–67 years for PV, 65–70 years for ET, and 67–70 years for PMF [82]. However, it can be diagnosed in younger individuals, particularly if there is a familial predisposition [83]. Some reports indicate that ET is more common in women (particularly at younger ages) and PV in men, while in PMF both

As demonstrated by European and international studies [86, 87], the distinction of MPNs in three nosological entities have a relevant prognostic significance. By and large, PN-MPN patients have a reduced life expectancy compared with general population, with PMF having the lowest overall survival (5.7 years), followed by PV with 15 years survival in 65% of cases and ET with an overall survival of more than

Despite insidious clinical onset, all PN-MPNs are at risk of clonal evolution and mortality. This is generally attributed to disease progression that may end in medullary failure (myelofibrosis or ineffective hematopoiesis) or transformation into other hematologic malignancies (the most common being acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS)) or the occurrence of bacterial infections and cardio- and cerebrovascular diseases, especially in younger patients [89, 90]. Fortunately, mortality due to these complications has been decreasing in

Until 2005 little was known about the etiology of PN-MPNs. The discovery of somatic mutations in Janus kinase 2 gene (*JAK2*), a member of the Janus kinase family located at chromosome 9 and first identified in 1993, was crucial. The identification of exon 14 V617F gain-of-function mutation, made by several independent groups of investigators [91–94], was one of the major genetic insights into the pathogenesis of the PN-MPNs and transformed the understanding of these disorders. It turned out to be the most important and most frequently recurring somatic mutation involved in PN-MPN pathogenesis, with the highest frequency (up to 95%) in PV, and 50–60% in ET and PMF patients (**Figure 2**) [9, 23, 55, 72, 95–99]. Although there is no gold standard and the choice of methodology is dependent

on the application, quantitative real-time PCR is a useful method for detecting

**24**

megakaryocytes), stem cell factor (SCF, promotes proliferation and self-renewal of multipotent hematopoietic primordial cells), granulocyte-stimulating factor (GSF, stimulates proliferation and differentiation of granulocytes), and interleukins. Cytokine hypersensitivity leads to monoclonal stimulation of the erythropoiesis, megakaryopoiesis, and granulopoiesis.

*JAK*2 serves as the cognate tyrosine kinase for the EPO and TPO receptors and can also be used by the G-CSF receptor, all of which lack an intrinsic kinase domain [9, 70]. Moreover, JAK2 is crucial for normal hematopoiesis, as demonstrated by abnormal erythropoiesis developed in JAK2-deficient mice [114]. It includes two main domains: one is an enzymatically active kinase domain (JAK homology 1 (JH1)), and the other corresponds to a catalytically inactive pseudokinase domain (JH2), which promotes an inhibitory affect that induces the inhibition of the kinase activity of JAK2 [114–116].

The most frequent mutation associated with PN-MPNs, *JAK2* V617F, is present in myeloblasts, granulocytes, erythroblasts, and all EPO-independent erythroid colonies. It consists of a gain-of-function missense mutation with a G to T (guanine to thymidine) substitution at nucleotide 1849, in exon 14 of the *JAK2* gene, resulting in the substitution of valine with phenylalanine at codon 617 in the inhibitory JH2 domain [102]. When V617F mutation occurs, the result is an increased activity in myeloid progenitor cells, which leads to proliferation and excessive production of mature cells [114, 116–119].

*JAK2* V617F activates signaling through the three main myeloid cytokine homodimeric receptors (EPO-R, MPL, and G-CSFR), which are involved in erythrocytosis, thrombocytosis, and neutrophilia, respectively. On the other hand, *CALR* or *MPL* mutants are restricted to MPL activation, explaining why *JAK2* V617F is associated with PV, ET, and PMF, whereas *CALR* and *MPL* mutants are found in ET and PMF [120].

In addition, expression of *JAK2* V617F results in constitutive activation of downstream signaling pathways including the JAK-STAT, MAPK/ERK, and phosphatidylinositol-3-kinase (PI3K/AKT) pathways [91–94] and later by interaction with p85, a regulatory subunit of PI3K, promoting proliferation and survival. Activated PI3K activates AKT, which in turn activates mammalian target of rapamycin (mTor) on Ser2448, which directly phosphorylates ribosomal p70S6 kinase (p70S6k). p70S6K and mTor are involved in angiogenesis by activation of vascular endothelial growth factor (VEGF) [61, 72]. It is known that this pathway is commonly activated in leukemia and lymphoma and is involved in inhibiting apoptosis in normal human erythroblasts. The PI3K/AKT pathway also induces the phosphorylation of BAD, a pro-apoptotic member of the Bcl2 family, via phosphorylated AKT (pAKT) and p70S6k, thus inhibiting BAD function and resulting in inhibition of apoptosis. BclxL is also activated by this pathway, resulting in inhibition of megakaryocyte apoptosis [61].

On the other hand, an increased activation of Ras-Erk signaling pathway was also demonstrated in PV patients. Ras is activated and activates Raf-1, which mediates the activation of MEK, which in turn activates extracellular signal-regulated kinase (ERK), one of members of the MAPK families. ERK phosphorylation also results in the inhibition of apoptosis, by blocking the function of BAD and activation of Bcl2. Therefore, due to the inactivation of the pro-apoptotic factor BAD and activation of BclxL and Bcl2, AKT and ERK together with *JAK2* V617F mutation suppress apoptosis and promote cellular survival, upregulating megakaryocytes and erythropoiesis [61].

In contrast to its effect on the EPO receptor, *JAK2* V617F appears to increase the quantity of immature MPL while increasing MPL degradation through ubiquitination and reducing its cell surface expression [70].

**27**

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies*

Several studies have shown that expression of *JAK2* V617F results in transformation of Ba/F3 cells, characterized by IL-3-independent growth, unlike wild-type JAK2 [91]. Due to *JAK2* V617F mutation and other mutations, hematopoietic progenitor cells can proliferate without the presence or induction by cytokines, resulting in factor-independent growth of the erythroid cell line and activation of signal transduction [102], mostly in PV homozygous cases. Yet, the presence of receptors is essential, leading to enhanced functional activity and increased sensitivity to cytokines and hematopoietic growth factors, such as interleukin 3 (IL-3), stem cell factor (SCF), granulocyte-macrophage CSF, and insulin-like growth factor-1

Recently, in 2017, Yao et al*.* demonstrated that activation of *JAK2* mutants can differentially link to selective cytokine receptors and change the signaling motifs, evidencing the molecular basis for phenotypic variants elicited by *JAK2* V617F or exon 12 mutations. On the basis of these findings, receptor-JAK2 interactions could evidence new targets of lineage-specific therapeutic tools against MPNs, which may

be considered in other cancers with aberrant JAK-STAT signaling [122].

Recent data also indicate that the *JAK2V617F* allele might escape negative

Unlike V617F where only a single codon is affected, exon 12 frameshift mutations comprise more than 40 different small deletions/duplications and substitutions of one or more amino acids between phenylalanines F533 and F547 (e.g., lysine for leucine at codon 539—K539 L), which are located in a linker between the JH2 pseudokinase and the SH2 domains [123]. However, just like *JAK2* V617F mutation, also exon 12 mutant alleles induce cytokine-independent/hypersensitive proliferation in EPO receptor (EPO-R) expressing cell lines and constitutive activation of JAK-STAT signaling [102]. The *JAK2* exon 12 mutations contribute primarily to erythroid myeloproliferation, associated with increasing levels of phosphorylated JAK2, STAT5, and Erk1/2 compared to patients with wild-type *JAK2*, and even higher activated JAK2 and ERK1/ERK2 levels than patients with the *JAK2* V617F

Although the complete cellular and molecular mechanisms involved in the pathophysiology of PN-MPNs have not yet been fully clarified [97, 107, 125–131], hyperactive JAK/STAT signaling pathway appears to be a constant, even in the presence of *CALR* mutations and the so-called "triple-negative" MPNs (nonmutated *JAK2*, *CALR*, and *MPL*), where the driver gene mutation is still unknown [55, 112].

In humans, *JAK2* V617F occurs at the stem cell level and is present in hematopoietic stem cell progenitors from affected individuals, but not usually in the germline, suggesting that this mutation is acquired as a somatic disease allele in the hematopoietic compartment [102]. It is believed to be myeloid lineage specific because it is present in erythroid and granulocyte-macrophage progenitors. *JAK2* V617F is not specific for an individual PN-MPN, nor does its absence exclude MPNs. Although the prevalence of *JAK2* V617F mutation differs among PN-MPNs, one of the most challenging aspects of the study of these disorders still is the explanation of pheno-

**3.3** *JAK2* **mutation's role in Philadelphia chromosome-negative myeloproliferative neoplasms and other disorders**

typic heterogeneity and mechanism of progression of the PN-MPNs [97].

About 25–30% of patients with PV and 2–4% with ET [102, 132] are homozygous for the *JAK2* V617F allele (loss of heterozygosity) as a result of mitotic recombination and duplication of the mutant allele, promoting uniparental disomy (UPD). Uniparental disomy of chromosomal locus 9p24, including *JAK2*, had previously been detected in PV, before identification of the *JAK2* V617F allele

*DOI: http://dx.doi.org/10.5772/intechopen.84873*

[23, 114, 121].

feedback by SOCS3 [72].

mutation [61, 103, 124].

#### *Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies DOI: http://dx.doi.org/10.5772/intechopen.84873*

Several studies have shown that expression of *JAK2* V617F results in transformation of Ba/F3 cells, characterized by IL-3-independent growth, unlike wild-type JAK2 [91]. Due to *JAK2* V617F mutation and other mutations, hematopoietic progenitor cells can proliferate without the presence or induction by cytokines, resulting in factor-independent growth of the erythroid cell line and activation of signal transduction [102], mostly in PV homozygous cases. Yet, the presence of receptors is essential, leading to enhanced functional activity and increased sensitivity to cytokines and hematopoietic growth factors, such as interleukin 3 (IL-3), stem cell factor (SCF), granulocyte-macrophage CSF, and insulin-like growth factor-1 [23, 114, 121].

Recently, in 2017, Yao et al*.* demonstrated that activation of *JAK2* mutants can differentially link to selective cytokine receptors and change the signaling motifs, evidencing the molecular basis for phenotypic variants elicited by *JAK2* V617F or exon 12 mutations. On the basis of these findings, receptor-JAK2 interactions could evidence new targets of lineage-specific therapeutic tools against MPNs, which may be considered in other cancers with aberrant JAK-STAT signaling [122].

Recent data also indicate that the *JAK2V617F* allele might escape negative feedback by SOCS3 [72].

Unlike V617F where only a single codon is affected, exon 12 frameshift mutations comprise more than 40 different small deletions/duplications and substitutions of one or more amino acids between phenylalanines F533 and F547 (e.g., lysine for leucine at codon 539—K539 L), which are located in a linker between the JH2 pseudokinase and the SH2 domains [123]. However, just like *JAK2* V617F mutation, also exon 12 mutant alleles induce cytokine-independent/hypersensitive proliferation in EPO receptor (EPO-R) expressing cell lines and constitutive activation of JAK-STAT signaling [102]. The *JAK2* exon 12 mutations contribute primarily to erythroid myeloproliferation, associated with increasing levels of phosphorylated JAK2, STAT5, and Erk1/2 compared to patients with wild-type *JAK2*, and even higher activated JAK2 and ERK1/ERK2 levels than patients with the *JAK2* V617F mutation [61, 103, 124].

Although the complete cellular and molecular mechanisms involved in the pathophysiology of PN-MPNs have not yet been fully clarified [97, 107, 125–131], hyperactive JAK/STAT signaling pathway appears to be a constant, even in the presence of *CALR* mutations and the so-called "triple-negative" MPNs (nonmutated *JAK2*, *CALR*, and *MPL*), where the driver gene mutation is still unknown [55, 112].

#### **3.3** *JAK2* **mutation's role in Philadelphia chromosome-negative myeloproliferative neoplasms and other disorders**

In humans, *JAK2* V617F occurs at the stem cell level and is present in hematopoietic stem cell progenitors from affected individuals, but not usually in the germline, suggesting that this mutation is acquired as a somatic disease allele in the hematopoietic compartment [102]. It is believed to be myeloid lineage specific because it is present in erythroid and granulocyte-macrophage progenitors. *JAK2* V617F is not specific for an individual PN-MPN, nor does its absence exclude MPNs. Although the prevalence of *JAK2* V617F mutation differs among PN-MPNs, one of the most challenging aspects of the study of these disorders still is the explanation of phenotypic heterogeneity and mechanism of progression of the PN-MPNs [97].

About 25–30% of patients with PV and 2–4% with ET [102, 132] are homozygous for the *JAK2* V617F allele (loss of heterozygosity) as a result of mitotic recombination and duplication of the mutant allele, promoting uniparental disomy (UPD). Uniparental disomy of chromosomal locus 9p24, including *JAK2*, had previously been detected in PV, before identification of the *JAK2* V617F allele

*Tyrosine Kinases as Druggable Targets in Cancer*

megakaryopoiesis, and granulopoiesis.

activity of JAK2 [114–116].

mature cells [114, 116–119].

inhibition of megakaryocyte apoptosis [61].

tion and reducing its cell surface expression [70].

and PMF [120].

megakaryocytes), stem cell factor (SCF, promotes proliferation and self-renewal of multipotent hematopoietic primordial cells), granulocyte-stimulating factor (GSF, stimulates proliferation and differentiation of granulocytes), and interleukins. Cytokine hypersensitivity leads to monoclonal stimulation of the erythropoiesis,

*JAK*2 serves as the cognate tyrosine kinase for the EPO and TPO receptors and can also be used by the G-CSF receptor, all of which lack an intrinsic kinase domain [9, 70]. Moreover, JAK2 is crucial for normal hematopoiesis, as demonstrated by abnormal erythropoiesis developed in JAK2-deficient mice [114]. It includes two main domains: one is an enzymatically active kinase domain (JAK homology 1 (JH1)), and the other corresponds to a catalytically inactive pseudokinase domain (JH2), which promotes an inhibitory affect that induces the inhibition of the kinase

The most frequent mutation associated with PN-MPNs, *JAK2* V617F, is present in myeloblasts, granulocytes, erythroblasts, and all EPO-independent erythroid colonies. It consists of a gain-of-function missense mutation with a G to T (guanine to thymidine) substitution at nucleotide 1849, in exon 14 of the *JAK2* gene, resulting in the substitution of valine with phenylalanine at codon 617 in the inhibitory JH2 domain [102]. When V617F mutation occurs, the result is an increased activity in myeloid progenitor cells, which leads to proliferation and excessive production of

*JAK2* V617F activates signaling through the three main myeloid cytokine homodimeric receptors (EPO-R, MPL, and G-CSFR), which are involved in erythrocytosis, thrombocytosis, and neutrophilia, respectively. On the other hand, *CALR* or *MPL* mutants are restricted to MPL activation, explaining why *JAK2* V617F is associated with PV, ET, and PMF, whereas *CALR* and *MPL* mutants are found in ET

In addition, expression of *JAK2* V617F results in constitutive activation of downstream signaling pathways including the JAK-STAT, MAPK/ERK, and phosphatidylinositol-3-kinase (PI3K/AKT) pathways [91–94] and later by interaction with p85, a regulatory subunit of PI3K, promoting proliferation and survival. Activated PI3K activates AKT, which in turn activates mammalian target of rapamycin (mTor) on Ser2448, which directly phosphorylates ribosomal p70S6 kinase (p70S6k). p70S6K and mTor are involved in angiogenesis by activation of vascular endothelial growth factor (VEGF) [61, 72]. It is known that this pathway is commonly activated in leukemia and lymphoma and is involved in inhibiting apoptosis in normal human erythroblasts. The PI3K/AKT pathway also induces the phosphorylation of BAD, a pro-apoptotic member of the Bcl2 family, via phosphorylated AKT (pAKT) and p70S6k, thus inhibiting BAD function and resulting in inhibition of apoptosis. BclxL is also activated by this pathway, resulting in

On the other hand, an increased activation of Ras-Erk signaling pathway was also demonstrated in PV patients. Ras is activated and activates Raf-1, which mediates the activation of MEK, which in turn activates extracellular signal-regulated kinase (ERK), one of members of the MAPK families. ERK phosphorylation also results in the inhibition of apoptosis, by blocking the function of BAD and activation of Bcl2. Therefore, due to the inactivation of the pro-apoptotic factor BAD and activation of BclxL and Bcl2, AKT and ERK together with *JAK2* V617F mutation suppress apoptosis and promote cellular survival, upregulating megakaryocytes

In contrast to its effect on the EPO receptor, *JAK2* V617F appears to increase the quantity of immature MPL while increasing MPL degradation through ubiquitina-

**26**

and erythropoiesis [61].

[102]. Mitotic recombination is more likely to occur in PV patients with mutation in exon 14 of the *JAK2* gene than in those with exon 12 mutations [133] and is an early genetic event in the development of PV, but not ET [102]. Although *JAK2* V617F homozygous subclones can be identified both in PV and ET patients, expression of a dominant homozygous subclone is almost exclusive in PV patients (~80% in PV and 50% in ET) [78, 119], originated by additional genetic or epigenetic events or, e.g., low levels of circulating erythropoietin in consequence of elevated hematocrit [119].

Although in the heterozygous state *JAK2* V617F-bearing receptors are still responsive to growth factors, in JAK2 V617F homozygosity, these receptors become autonomous with respect to growth factor [70], as referred earlier.

Almost all patients diagnosed with PV negative for *JAK2* V617F mutation are exon 12 positive (95% vs. 2–4%, respectively) [53, 103, 134–141]. Some studies have reported that Chinese PV patients have a relatively lower *JAK2* V617F mutation frequency (82%), in line with a Portuguese study [23], while the mutations in *JAK2* exon 12 are much more pervasive (13%), when compared to Westerns and other East Asians [139, 142].

Unlike *JAK2* V617F, which can be detected in any of the PN-MPNs, *JAK2* exon 12 mutations are almost exclusive of *JAK2* V617F-negative PV patients [24, 103]. PV patients who present *JAK2* exon 12 mutations, unlike those who are V617F positive, are not commonly homozygous [70, 103, 124, 138]. PV patients with the *JAK2* exon 12 mutations are usually younger than those with the *JAK2* V617F mutation and have a phenotype usually more benign than that of JAK2V617F, usually without panmyelosis [53], with normal leukocyte and platelet counts [61, 70]. Although *JAK2* V617F and exon 12 mutations express through the same C-terminal tyrosine kinase of JAK2, they originate very different phenotypic outcomes. These patients appear to be associated with a distinct syndrome, with higher hemoglobin concentrations, without concomitant leukocytosis or thrombocytosis (or minimal thrombocytosis), and isolated bone marrow erythroid hyperplasia [124], independently of the mutational variant [24, 124, 140]. The reasons for these various abnormal phenotypic readouts also remain unclear and are likely to be complex [124, 140]. The fact that exon 12 mutations are more frequently associated with erythrocytosis is consistent with their absence in ET but possible existence in PMF or AML secondary to PV [138]. However, there are exceptions as evidenced in some clinical reports [24]. Despite the phenotypical diversity, the clinical course and outcome seem overlapping between *JAK2* V617F and *JAK2* exon 12-positive patients, with convergent incidences of thrombosis, myelofibrosis, leukemia, and death [140]. There are also reports of the coexistence of *JAK2* V617F and *JAK2* exon 12 mutations as two separate clones [70, 140].

As published by Rumi and Cazzola [78], patients with the wild-type genotype for *JAK2* are extremely rare. However, a recent study [23] demonstrated a prevalence of 12.8% of patients with that genotype. This finding is consistent with the fact that the *JAK2* mutation expression alone may not be sufficient to induce the PV phenotype. However, larger studies are required to confirm this hypothesis.

Some reports have also suggested *JAK2* V617F clonal involvement of B [143, 144], T [143], and NK lymphocytes [83], also confirming the stem cell nature of *JAK2* V617F MPNs [102]. Lower frequencies of V617F mutation occur in PN-CML, chronic myelomonocytic leukemia, juvenile myelomonocytic leukemia, and rare cases of AML (megakaryocytic and in combination with other well-defined genetic abnormalities, such as BCR-ABL1) [145]. There is also evidence of association with certain solid tumors (generally non-hematological types) [51, 114, 117, 146–148]. Other mutations in the JAK2 pseudokinase domain (including point mutations involving R683) have been detected in about 20% of Down syndrome-associated

**29**

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies*

and other acute lymphoblastic leukemia and AML. A number of JAK2 fusion proteins, such as TEL-JAK2, PCM1-JAK2, and BCR-JAK2, lead to activation of JAK kinase activity and have also been associated with myeloid and lymphoid leukemia

Along with other driver mutations connected with clonal expansion of hematopoietic cells, *JAK2* V617F mutation might also represent a feature of the aging hematopoietic system in individuals without a malignant disease [149, 150]. There is increasing evidence that *JAK2* V617F is relatively frequent in the aging healthy population and is presently estimated to be 0.5% [120]. These individuals usually present higher erythrocyte, platelet, and leucocyte counts and are more likely to develop a hematological cancer. Aging is generally associated with a deregulation of hematopoietic stem cells, which lose their function and become myeloid-biased and less quiescent as a consequence of intrinsic and environmental changes, with *JAK2* V617F hematopoietic stem cells having higher competitive properties in this

Besides mutations and other molecular defects, various factors, such as gene burden and individual genetic background, may be responsible for predisposition

Several published data have shown the contribution and influence of *JAK2* V617F mutation allelic burden in the definition of phenotype and prognostic impact in PN-MPNs [151, 152]. *JAK2* V617F allelic burden corresponds to the ratio between mutant and wild-type *JAK2* in hematopoietic cells and is on the basis of a stronger activation of intracellular signaling pathways [153]. Between MPN patients there is a variability in the number of cells carrying the *JAK2* V617F mutation, and there is a

It is recognized that the allele burden tends to be higher in PV (due to the higher number of homozygous cases) and PMF, associated with the presence of acquired UPD, with defined hematological and clinical markers indicative of a more aggressive phenotype [153]. Indeed, a lower allele burden is generally observed in ET patients [97, 119, 152, 154, 155], but when it increases, some of them transform over time to PV or PMF. Importantly, ET patients positive for the *JAK2* V617F mutation have a "PV-like" phenotype compared to ET patients without this genetic abnormality. However, patients carrying *JAK2* V617F mutation do not have a higher risk of evolution to post-PV and post-ET myelofibrosis than patients

Another possible explanation concerns the concept of a "pre-JAK2" phase in which additional somatic mutations or inherited predisposing alleles present before the mutation are responsible for the clonal hematopoiesis, determine the phenotype, influence the risk of progression to AML, and might even be responsible for generating the mutation or act synergistically [55, 61]. In fact, although *JAK2* V617F mutation is crucial to the pathogenesis of PV, ET, and PMF, the existence of the same allele in three clinically distinct entities suggests that there might be additional inherited or acquired genetic predisposition. Indeed, a familial tendency has been identified in 72 families, which is consistent with an inherited genetic predisposi-

On the other hand, the role of the *JAK2* V617F mutation in the pathogenicity of the various MPNs may differ among different MPNs, involving the *JAK2* V617F mutation more often than others (e.g., ET vs. PV), which would indicate other oncogenic mutations or factors that may be determinant for certain cases other than

for developing an MPN, as well as influence their heterogeneity [78, 97].

*DOI: http://dx.doi.org/10.5772/intechopen.84873*

or atypical CML [60, 72].

context [120, 150].

*3.3.1 Prognosis and predictive factors*

without the mutation [61].

tion to MPNs [156].

*JAK2* V617F [97, 119, 157, 158].

variability in the alleles that carry the mutation.

#### *Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies DOI: http://dx.doi.org/10.5772/intechopen.84873*

and other acute lymphoblastic leukemia and AML. A number of JAK2 fusion proteins, such as TEL-JAK2, PCM1-JAK2, and BCR-JAK2, lead to activation of JAK kinase activity and have also been associated with myeloid and lymphoid leukemia or atypical CML [60, 72].

Along with other driver mutations connected with clonal expansion of hematopoietic cells, *JAK2* V617F mutation might also represent a feature of the aging hematopoietic system in individuals without a malignant disease [149, 150]. There is increasing evidence that *JAK2* V617F is relatively frequent in the aging healthy population and is presently estimated to be 0.5% [120]. These individuals usually present higher erythrocyte, platelet, and leucocyte counts and are more likely to develop a hematological cancer. Aging is generally associated with a deregulation of hematopoietic stem cells, which lose their function and become myeloid-biased and less quiescent as a consequence of intrinsic and environmental changes, with *JAK2* V617F hematopoietic stem cells having higher competitive properties in this context [120, 150].

#### *3.3.1 Prognosis and predictive factors*

*Tyrosine Kinases as Druggable Targets in Cancer*

of elevated hematocrit [119].

East Asians [139, 142].

separate clones [70, 140].

[102]. Mitotic recombination is more likely to occur in PV patients with mutation in exon 14 of the *JAK2* gene than in those with exon 12 mutations [133] and is an early genetic event in the development of PV, but not ET [102]. Although *JAK2* V617F homozygous subclones can be identified both in PV and ET patients, expression of a dominant homozygous subclone is almost exclusive in PV patients (~80% in PV and 50% in ET) [78, 119], originated by additional genetic or epigenetic events or, e.g., low levels of circulating erythropoietin in consequence

Although in the heterozygous state *JAK2* V617F-bearing receptors are still responsive to growth factors, in JAK2 V617F homozygosity, these receptors become

Almost all patients diagnosed with PV negative for *JAK2* V617F mutation are exon 12 positive (95% vs. 2–4%, respectively) [53, 103, 134–141]. Some studies have reported that Chinese PV patients have a relatively lower *JAK2* V617F mutation frequency (82%), in line with a Portuguese study [23], while the mutations in *JAK2* exon 12 are much more pervasive (13%), when compared to Westerns and other

Unlike *JAK2* V617F, which can be detected in any of the PN-MPNs, *JAK2* exon 12 mutations are almost exclusive of *JAK2* V617F-negative PV patients [24, 103]. PV patients who present *JAK2* exon 12 mutations, unlike those who are V617F positive, are not commonly homozygous [70, 103, 124, 138]. PV patients with the *JAK2* exon 12 mutations are usually younger than those with the *JAK2* V617F mutation and have a phenotype usually more benign than that of JAK2V617F, usually without panmyelosis [53], with normal leukocyte and platelet counts [61, 70]. Although *JAK2* V617F and exon 12 mutations express through the same C-terminal tyrosine kinase of JAK2, they originate very different phenotypic outcomes. These patients appear to be associated with a distinct syndrome, with higher hemoglobin concentrations, without concomitant leukocytosis or thrombocytosis (or minimal thrombocytosis), and isolated bone marrow erythroid hyperplasia [124], independently of the mutational variant [24, 124, 140]. The reasons for these various abnormal phenotypic readouts also remain unclear and are likely to be complex [124, 140]. The fact that exon 12 mutations are more frequently associated with erythrocytosis is consistent with their absence in ET but possible existence in PMF or AML secondary to PV [138]. However, there are exceptions as evidenced in some clinical reports [24]. Despite the phenotypical diversity, the clinical course and outcome seem overlapping between *JAK2* V617F and *JAK2* exon 12-positive patients, with convergent incidences of thrombosis, myelofibrosis, leukemia, and death [140]. There are also reports of the coexistence of *JAK2* V617F and *JAK2* exon 12 mutations as two

As published by Rumi and Cazzola [78], patients with the wild-type genotype for *JAK2* are extremely rare. However, a recent study [23] demonstrated a prevalence of 12.8% of patients with that genotype. This finding is consistent with the fact that the *JAK2* mutation expression alone may not be sufficient to induce the PV

Some reports have also suggested *JAK2* V617F clonal involvement of B [143, 144],

phenotype. However, larger studies are required to confirm this hypothesis.

T [143], and NK lymphocytes [83], also confirming the stem cell nature of *JAK2* V617F MPNs [102]. Lower frequencies of V617F mutation occur in PN-CML, chronic myelomonocytic leukemia, juvenile myelomonocytic leukemia, and rare cases of AML (megakaryocytic and in combination with other well-defined genetic abnormalities, such as BCR-ABL1) [145]. There is also evidence of association with certain solid tumors (generally non-hematological types) [51, 114, 117, 146–148]. Other mutations in the JAK2 pseudokinase domain (including point mutations involving R683) have been detected in about 20% of Down syndrome-associated

autonomous with respect to growth factor [70], as referred earlier.

**28**

Besides mutations and other molecular defects, various factors, such as gene burden and individual genetic background, may be responsible for predisposition for developing an MPN, as well as influence their heterogeneity [78, 97].

Several published data have shown the contribution and influence of *JAK2* V617F mutation allelic burden in the definition of phenotype and prognostic impact in PN-MPNs [151, 152]. *JAK2* V617F allelic burden corresponds to the ratio between mutant and wild-type *JAK2* in hematopoietic cells and is on the basis of a stronger activation of intracellular signaling pathways [153]. Between MPN patients there is a variability in the number of cells carrying the *JAK2* V617F mutation, and there is a variability in the alleles that carry the mutation.

It is recognized that the allele burden tends to be higher in PV (due to the higher number of homozygous cases) and PMF, associated with the presence of acquired UPD, with defined hematological and clinical markers indicative of a more aggressive phenotype [153]. Indeed, a lower allele burden is generally observed in ET patients [97, 119, 152, 154, 155], but when it increases, some of them transform over time to PV or PMF. Importantly, ET patients positive for the *JAK2* V617F mutation have a "PV-like" phenotype compared to ET patients without this genetic abnormality. However, patients carrying *JAK2* V617F mutation do not have a higher risk of evolution to post-PV and post-ET myelofibrosis than patients without the mutation [61].

Another possible explanation concerns the concept of a "pre-JAK2" phase in which additional somatic mutations or inherited predisposing alleles present before the mutation are responsible for the clonal hematopoiesis, determine the phenotype, influence the risk of progression to AML, and might even be responsible for generating the mutation or act synergistically [55, 61]. In fact, although *JAK2* V617F mutation is crucial to the pathogenesis of PV, ET, and PMF, the existence of the same allele in three clinically distinct entities suggests that there might be additional inherited or acquired genetic predisposition. Indeed, a familial tendency has been identified in 72 families, which is consistent with an inherited genetic predisposition to MPNs [156].

On the other hand, the role of the *JAK2* V617F mutation in the pathogenicity of the various MPNs may differ among different MPNs, involving the *JAK2* V617F mutation more often than others (e.g., ET vs. PV), which would indicate other oncogenic mutations or factors that may be determinant for certain cases other than *JAK2* V617F [97, 119, 157, 158].

Moreover, mutations in epigenetic regulators, transcription factors, and signaling components modify the course of the disease and can contribute to disease initiation and/ or progression [55]. Some studies performed in mice and humans led to the "host genetic factor" concept, acting as modifiers in combination with the mutation, for instance, single nucleotide polymorphisms (SNPs) [90, 110, 111, 159, 160]. Even gender could be an independent modifier, with women having a lower allele burden than men [61].

Also, the coexistence of autonomous *JAK2* mutant and *JAK2* wild-type clonal populations in the same patient can be an explanation. It is observed that *JAK2* positive AML patients are preceded by evolution to myelofibrosis during their disease course, in contrast to *JAK2* wild-type AML, which is preceded by chronicphase ET and PV patients [61].

On the other hand, the role of the JAK/STAT signaling pathway in the pathogenesis of MPNs and other cancers is questionable when taking into account the example of rare families hosting germline mutations leading to weak JAK expression. The mutations induce a hereditary thrombocytosis, but hematopoiesis is polyclonal, and there is no generation of hematological malignancies or solid tumors, indicating that JAK/STAT activation alone does not drive malignant disease [147].

In PV and ET, risk factors influencing survival include older age, leukocytosis, and thrombosis. In ET, the *JAK2* V617F mutation is associated with increased risk of thrombosis, leading to inclusion into the International Prognostic Score of Thrombosis for ET-thrombosis score [90, 94, 161]. Expansion of *JAK2*-mutated allele promotes the transformation of PV and ET to secondary myelofibrosis [153]. Furthermore, the presence of two or more mutations is associated with a worse survival and predicts shortened leukemia-free survival [162].

*JAK2* V617F has not been correlated to an increased risk of transformation to AML [90]; nevertheless, *JAK2* V617F-positive patients with MPN diagnosis can transform to *JAK2* V617F-negative AML [163].

The pathogenesis of thrombosis in PN-MPN patients is complex, involving clinical factors such as age, previous history of thrombotic events, obesity, hypertension, and hyperlipemia, as well increased blood cell counts (i.e., leukocytosis, erythrocytosis, and thrombocytosis), high hematocrit, and *JAK2* mutation [164]. The most important risk factor for future arterial and venous thrombosis in MPNs is the previous history of arterial and venous thrombosis, respectively [9]. The influence of the *JAK2* V617F mutational status and allele burden on the thrombotic risk has been evaluated and established in several studies among PN-MPNs [90]; however, regarding the presence of *MPL* mutation, the published results are discrepant [164]. Older (age > 60 years) patients are no longer considered "high risk," unless they have a history of thrombosis or are *JAK2*-mutated [9, 164].

In patients with ET, the frequency of thromboembolic events in different studies ranges from 10 to 30% at diagnosis and between 8 and 31% during follow-up [165], and the rate of fatal and nonfatal thrombotic events ranged from 2 to 4% patientyears, with a predominance of arterial events [164], whose risk is higher in patients with *JAK2* and *MPL* mutations [90, 166].

Risk factors for fibrotic transformation in PV include *JAK2* V617F allele burden of >50%; in ET they include advanced age and anemia, with the presence of *JAK2* V617F being associated with a lower risk of fibrotic transformation and *CALR* with a higher risk [9]. *JAK2* V617F mutational status may have prognostic significance in PV, ET, and PMF [102]. In PV, despite the phenotypic differences, the clinical course seems similar between *JAK2* V617F and *JAK2* exon 12-positive patients, with similar incidences of thrombosis, myelofibrosis, leukemia, and death [24, 140]. *JAK2/CALR* mutational status did not affect survival in ET [9]. In PMF and ET, triple-negative patients appear to have a less favorable prognosis than patients with

**31**

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies*

tend to have a better prognosis than patients with *JAK2* or *MPL* mutations.

a driver mutation (*JAK2*, *CALR*, or *MPL*), whereas patients with *CALR* mutations

Another important concern refers to the increased risk of generation of new non-hematological and nonmyeloid neoplasms in MPNs, with an incidence ratio of 1.2–1.4 and 3.4, respectively, compared to the general population [90, 167]. There is evidence that this risk is higher when *JAK2* V617F mutation is identified and other

The discovery of the *JAK2* mutations and their relation with the subsequent activation of the JAK-STAT pathway was crucial to the understanding of the pathogenesis of PV, ET, and PMF. This knowledge has led to the development of small-molecular *JAK* inhibitors to target autoimmune disease/immunosuppression (anti-*JAK1*, *JAK3*) and MPNs and leukemia/lymphoma (anti-*JAK2*, *JAK1*), which have been tested in several clinical trials, suggesting an overall reduction in JAK-STAT signaling and pro-inflammatory cytokines [141, 168, 169]. About 10 compounds were studied for MPNs, rheumatoid arthritis, psoriasis, and inflammatory bowel disease, all of them targeting the ATP-binding site of JAKs, but none is absolutely specific for any JAK [88]. Nevertheless, ruxolitinib (a *JAK1*, *JAK2* inhibitor, trade name Jakavi®) has been approved by the Food and Drug Administration (FDA) in November 2011, for use in myelofibrosis, and tofacitinib (a *JAK1*, J*AK3* inhibitor) has been approved for use in rheumatoid arthritis. The first two randomized controlled trials (Comfort I and II) on the effect of the JAK2 inhibitor ruxolitinib versus placebo and versus the best available therapy in intermediate-2 and high-risk PMF showed a decrease in spleen size and symptom burden in the experimental arm of both studies. In Comfort I, a survival benefit was also observed in the ruxolitinib arm compared to patients on placebo [170, 171]. Although ruxolitinib was recently approved for use in hydroxyurea-resistant PV, its role in routine clini-

The treatment options of PMF patients are currently limited, with stem cell transplant being the current treatment of choice for genetically or clinically high-risk disease. PMF patients may benefit from *JAK2* inhibition with immediate clinical value in the management of symptoms, through directly modulating the pro-growth signals of the JAK-STAT pathway, suppression of hematopoietic progenitor cell proliferation, and from downregulating specific pro-inflammatory

Ruxolitinib treatment substantially alleviates symptomatic splenomegaly and constitutional symptoms and improves quality of life in a significant proportion of patients with primary or post-PV/ET myelofibrosis [88]. Surprisingly, treatment with ruxolitinib is also effective in patients without mutated *JAK2*, suggesting that other, still unknown, underlying mechanisms are responsible for the increased JAK/ STAT pathway activity in PN-MPN patients. On the other hand, there is no convincing evidence of reduction in mutated allele burden, disease modification, nor

The identification of *JAK2* represented a milestone for the following studies and for today's knowledge, but the ongoing discovery of other mutations in MPNs will make possible the establishment of new drug targets and prognostic biomarkers that will for certain improve clinical practice and patients' outcome. All in all, it remains to be fully clarified whether *JAK2* mutations may be considered as "driver mutations" for MPNs or if they can act as "passenger mutations" which may alter-

*DOI: http://dx.doi.org/10.5772/intechopen.84873*

patient-related factors may be also present.

cal practice remains controversial [9, 52, 95, 172, 173].

cytokines produced by the affected clone [70, 113].

nate place with the former and have "driver" functions [129].

progression to AML [9, 174].

*3.3.2 Therapy management*

#### *Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies DOI: http://dx.doi.org/10.5772/intechopen.84873*

a driver mutation (*JAK2*, *CALR*, or *MPL*), whereas patients with *CALR* mutations tend to have a better prognosis than patients with *JAK2* or *MPL* mutations.

Another important concern refers to the increased risk of generation of new non-hematological and nonmyeloid neoplasms in MPNs, with an incidence ratio of 1.2–1.4 and 3.4, respectively, compared to the general population [90, 167]. There is evidence that this risk is higher when *JAK2* V617F mutation is identified and other patient-related factors may be also present.

#### *3.3.2 Therapy management*

*Tyrosine Kinases as Druggable Targets in Cancer*

phase ET and PV patients [61].

Moreover, mutations in epigenetic regulators, transcription factors, and signaling components modify the course of the disease and can contribute to disease initiation and/ or progression [55]. Some studies performed in mice and humans led to the "host genetic factor" concept, acting as modifiers in combination with the mutation, for instance, single nucleotide polymorphisms (SNPs) [90, 110, 111, 159, 160]. Even gender could be an independent modifier, with women having a lower allele burden than men [61]. Also, the coexistence of autonomous *JAK2* mutant and *JAK2* wild-type clonal populations in the same patient can be an explanation. It is observed that *JAK2* positive AML patients are preceded by evolution to myelofibrosis during their disease course, in contrast to *JAK2* wild-type AML, which is preceded by chronic-

On the other hand, the role of the JAK/STAT signaling pathway in the pathogenesis of MPNs and other cancers is questionable when taking into account the example of rare families hosting germline mutations leading to weak JAK expression. The mutations induce a hereditary thrombocytosis, but hematopoiesis is polyclonal, and there is no generation of hematological malignancies or solid tumors, indicating

In PV and ET, risk factors influencing survival include older age, leukocytosis, and thrombosis. In ET, the *JAK2* V617F mutation is associated with increased risk of thrombosis, leading to inclusion into the International Prognostic Score of Thrombosis for ET-thrombosis score [90, 94, 161]. Expansion of *JAK2*-mutated allele promotes the transformation of PV and ET to secondary myelofibrosis [153]. Furthermore, the presence of two or more mutations is associated with a worse

*JAK2* V617F has not been correlated to an increased risk of transformation to AML [90]; nevertheless, *JAK2* V617F-positive patients with MPN diagnosis can

The pathogenesis of thrombosis in PN-MPN patients is complex, involving clinical factors such as age, previous history of thrombotic events, obesity, hypertension, and hyperlipemia, as well increased blood cell counts (i.e., leukocytosis, erythrocytosis, and thrombocytosis), high hematocrit, and *JAK2* mutation [164]. The most important risk factor for future arterial and venous thrombosis in MPNs is the previous history of arterial and venous thrombosis, respectively [9]. The influence of the *JAK2* V617F mutational status and allele burden on the thrombotic risk has been evaluated and established in several studies among PN-MPNs [90]; however, regarding the presence of *MPL* mutation, the published results are discrepant [164]. Older (age > 60 years) patients are no longer considered "high risk,"

In patients with ET, the frequency of thromboembolic events in different studies ranges from 10 to 30% at diagnosis and between 8 and 31% during follow-up [165], and the rate of fatal and nonfatal thrombotic events ranged from 2 to 4% patientyears, with a predominance of arterial events [164], whose risk is higher in patients

Risk factors for fibrotic transformation in PV include *JAK2* V617F allele burden of >50%; in ET they include advanced age and anemia, with the presence of *JAK2* V617F being associated with a lower risk of fibrotic transformation and *CALR* with a higher risk [9]. *JAK2* V617F mutational status may have prognostic significance in PV, ET, and PMF [102]. In PV, despite the phenotypic differences, the clinical course seems similar between *JAK2* V617F and *JAK2* exon 12-positive patients, with similar incidences of thrombosis, myelofibrosis, leukemia, and death [24, 140]. *JAK2/CALR* mutational status did not affect survival in ET [9]. In PMF and ET, triple-negative patients appear to have a less favorable prognosis than patients with

unless they have a history of thrombosis or are *JAK2*-mutated [9, 164].

that JAK/STAT activation alone does not drive malignant disease [147].

survival and predicts shortened leukemia-free survival [162].

transform to *JAK2* V617F-negative AML [163].

with *JAK2* and *MPL* mutations [90, 166].

**30**

The discovery of the *JAK2* mutations and their relation with the subsequent activation of the JAK-STAT pathway was crucial to the understanding of the pathogenesis of PV, ET, and PMF. This knowledge has led to the development of small-molecular *JAK* inhibitors to target autoimmune disease/immunosuppression (anti-*JAK1*, *JAK3*) and MPNs and leukemia/lymphoma (anti-*JAK2*, *JAK1*), which have been tested in several clinical trials, suggesting an overall reduction in JAK-STAT signaling and pro-inflammatory cytokines [141, 168, 169]. About 10 compounds were studied for MPNs, rheumatoid arthritis, psoriasis, and inflammatory bowel disease, all of them targeting the ATP-binding site of JAKs, but none is absolutely specific for any JAK [88]. Nevertheless, ruxolitinib (a *JAK1*, *JAK2* inhibitor, trade name Jakavi®) has been approved by the Food and Drug Administration (FDA) in November 2011, for use in myelofibrosis, and tofacitinib (a *JAK1*, J*AK3* inhibitor) has been approved for use in rheumatoid arthritis. The first two randomized controlled trials (Comfort I and II) on the effect of the JAK2 inhibitor ruxolitinib versus placebo and versus the best available therapy in intermediate-2 and high-risk PMF showed a decrease in spleen size and symptom burden in the experimental arm of both studies. In Comfort I, a survival benefit was also observed in the ruxolitinib arm compared to patients on placebo [170, 171]. Although ruxolitinib was recently approved for use in hydroxyurea-resistant PV, its role in routine clinical practice remains controversial [9, 52, 95, 172, 173].

The treatment options of PMF patients are currently limited, with stem cell transplant being the current treatment of choice for genetically or clinically high-risk disease. PMF patients may benefit from *JAK2* inhibition with immediate clinical value in the management of symptoms, through directly modulating the pro-growth signals of the JAK-STAT pathway, suppression of hematopoietic progenitor cell proliferation, and from downregulating specific pro-inflammatory cytokines produced by the affected clone [70, 113].

Ruxolitinib treatment substantially alleviates symptomatic splenomegaly and constitutional symptoms and improves quality of life in a significant proportion of patients with primary or post-PV/ET myelofibrosis [88]. Surprisingly, treatment with ruxolitinib is also effective in patients without mutated *JAK2*, suggesting that other, still unknown, underlying mechanisms are responsible for the increased JAK/ STAT pathway activity in PN-MPN patients. On the other hand, there is no convincing evidence of reduction in mutated allele burden, disease modification, nor progression to AML [9, 174].

The identification of *JAK2* represented a milestone for the following studies and for today's knowledge, but the ongoing discovery of other mutations in MPNs will make possible the establishment of new drug targets and prognostic biomarkers that will for certain improve clinical practice and patients' outcome. All in all, it remains to be fully clarified whether *JAK2* mutations may be considered as "driver mutations" for MPNs or if they can act as "passenger mutations" which may alternate place with the former and have "driver" functions [129].

#### **4. Conclusions and future perspectives**

Non-receptor tyrosine kinases play an important role in the development of human malignancies, including hematological and others, and of inflammatory, and autoimmune diseases, through their profound involvement in the regulation of several vital cellular mechanisms, including cell proliferation, differentiation, maturation, apoptosis, and survival.

Targeting dysregulated NRTKs may prevent the process of tumorigenesis. The screening and clinical use of tyrosine kinase inhibitors, in combination with conventional treatments, have allowed the potential of targeted-based cancer therapy using specific cancer cell molecules, which are less toxic than traditional cytotoxic chemotherapy. The establishment of effective strategies in cancer research and patient care is mandatory.

#### **Acknowledgements**

This revision included data obtained from patients and controls who generously participate to whom authors gratefully acknowledge. Appreciation and thankfulness are extended to Luísa Manso Oliveira and Inês Sousa for their expert technical assistance.

The mentioned work was supported by funding through project UID/ BIM/00009/2016 (Centre for Toxicogenomics and Human Health (ToxOmics), from Fundação para a Ciência e Tecnologia (FCT), Portugal.

#### **Conflict of interest**

The authors claim no competing financial or intellectual conflicts of interest in the preparation and submission of this chapter.

#### **Author details**

Ana Azevedo1,2\*, Susana Silva1 and José Rueff<sup>1</sup>

1 Faculty of Medical Sciences, NOVA Medical School, Centre for Toxicogenomics and Human Health, Genetics, Oncology and Human Toxicology, NOVA University of Lisbon, Lisbon, Portugal

2 Department of Clinical Pathology, Hospital of São Francisco Xavier, West Lisbon Hospital Centre, Lisbon, Portugal

\*Address all correspondence to: ana.azevedo@nms.unl.pt

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

**33**

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies*

Clinical Medicine Insights: Oncology.

[11] Lamontanara AJ, Georgeon S, Tria G, Svergun DI, Hantschel O. The SH2 domain of ABL kinases regulates kinase autophosphorylation by controlling activation loop accessibility. Nature Communications. 2014;**5**:5470

[12] Gromicho M, Rueff J, Rodrigues AS. Dynamics of expression of drug transporters: Methods for appraisal. Methods in Molecular Biology.

[13] Gromicho M, Magalhães M, Torres F, Dinis J, Fernandes AR, Rendeiro P, et al. Instability of mRNA expression signatures of drug transporters in chronic myeloid leukemia patients resistant to imatinib. Oncology Reports.

[14] Gromicho M, Dinis J, Magalhães M, Fernandes AR, Tavares P, Laires A, et al. Development of imatinib and dasatinib resistance: Dynamics of expression of drug transporters ABCB1, ABCC1, ABCG2, MVP, and SLC22A1. Leukemia & Lymphoma.

[15] Voisset E, Lopez S, Dubreuil P, De Sepulveda P. The tyrosine kinase FES is an essential effector of KITD816V proliferation signal. Blood.

[16] Hellwig S, Miduturu CV, Kanda S, Zhang J, Filippakopoulos P, Salah E, et al. Small-molecule inhibitors of the c-FES protein-tyrosine kinase. Chemistry & Biology. 2012;**19**:529-540

[17] Condorelli F, Stec-Martyna E, Zaborowska J, Felli L, Gnemmi I, Ponassi M, et al. Role of the nonreceptor tyrosine kinase FES in cancer. Current Medicinal Chemistry.

2017;**11**:1179554917702870

2016;**1395**:75-85

2013;**29**:741-750

2011;**52**:1980-1990

2007;**110**:2593-2599

2011;**18**:2913-2920

*DOI: http://dx.doi.org/10.5772/intechopen.84873*

[1] Lahiry P, Torkamani A, Schork NJ, Hegele RA. Kinase mutations in human disease: Interpreting genotypephenotype relationships. Nature Reviews. Genetics. 2010;**11**:60-74

**References**

[2] Paul MK, Mukhopadhyay AK. Tyrosine kinase—Role and significance in cancer. International Journal of Medical Sciences. 2004;**1**:101-115

[3] Siveen KS, Prabhu KS, Achkar IW, Kuttikrishnan S, Shyam S, Khan AQ, et al. Role of non receptor tyrosine kinases in hematological malignances and its targeting by natural products.

Molecular Cancer. 2018;**17**:31

2011;**65**:819-828

[4] Kosior K, Lewandowska-Grygiel M, Giannopoulos K. Tyrosine kinase inhibitors in hematological malignancies. Postȩpy Higieny i Medycyny Doświadczalnej (Online).

[5] Du Z, Lovly CM. Mechanisms of receptor tyrosine kinase activation in cancer. Molecular Cancer. 2018;**17**:58

of inhibitors for protein tyrosine kinases. Oncogene. 2000;**19**:5690-5701

[7] Wang J, Pendergast AM. The emerging role of ABL kinases in solid tumors. Trends Cancer. 2015;**1**:110-123

[8] Sirvent A, Benistant C, Roche S. Cytoplasmic signalling by the c-ABL tyrosine kinase in normal and cancer cells. Biology of the Cell.

[9] Tefferi A. Myeloproliferative

of Hematology. 2016;**91**:50-58

neoplasms: A decade of discoveries and treatment advances. American Journal

[10] Azevedo AP, Reichert A, Afonso C, Alberca MD, Tavares P, Lima F. V280G mutation, potential role in imatinib resistance: First case report.

2008;**100**:617-631

[6] Al-Obeidi FA, Lam KS. Development

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies DOI: http://dx.doi.org/10.5772/intechopen.84873*

#### **References**

*Tyrosine Kinases as Druggable Targets in Cancer*

maturation, apoptosis, and survival.

patient care is mandatory.

**Acknowledgements**

**Conflict of interest**

**Author details**

Ana Azevedo1,2\*, Susana Silva1

of Lisbon, Lisbon, Portugal

Hospital Centre, Lisbon, Portugal

assistance.

**4. Conclusions and future perspectives**

Non-receptor tyrosine kinases play an important role in the development of human malignancies, including hematological and others, and of inflammatory, and autoimmune diseases, through their profound involvement in the regulation of several vital cellular mechanisms, including cell proliferation, differentiation,

Targeting dysregulated NRTKs may prevent the process of tumorigenesis. The screening and clinical use of tyrosine kinase inhibitors, in combination with conventional treatments, have allowed the potential of targeted-based cancer therapy using specific cancer cell molecules, which are less toxic than traditional cytotoxic chemotherapy. The establishment of effective strategies in cancer research and

This revision included data obtained from patients and controls who generously participate to whom authors gratefully acknowledge. Appreciation and thankfulness are extended to Luísa Manso Oliveira and Inês Sousa for their expert technical

The authors claim no competing financial or intellectual conflicts of interest in

The mentioned work was supported by funding through project UID/ BIM/00009/2016 (Centre for Toxicogenomics and Human Health (ToxOmics),

and José Rueff<sup>1</sup>

1 Faculty of Medical Sciences, NOVA Medical School, Centre for Toxicogenomics and Human Health, Genetics, Oncology and Human Toxicology, NOVA University

2 Department of Clinical Pathology, Hospital of São Francisco Xavier, West Lisbon

from Fundação para a Ciência e Tecnologia (FCT), Portugal.

the preparation and submission of this chapter.

**32**

provided the original work is properly cited.

\*Address all correspondence to: ana.azevedo@nms.unl.pt

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

[1] Lahiry P, Torkamani A, Schork NJ, Hegele RA. Kinase mutations in human disease: Interpreting genotypephenotype relationships. Nature Reviews. Genetics. 2010;**11**:60-74

[2] Paul MK, Mukhopadhyay AK. Tyrosine kinase—Role and significance in cancer. International Journal of Medical Sciences. 2004;**1**:101-115

[3] Siveen KS, Prabhu KS, Achkar IW, Kuttikrishnan S, Shyam S, Khan AQ, et al. Role of non receptor tyrosine kinases in hematological malignances and its targeting by natural products. Molecular Cancer. 2018;**17**:31

[4] Kosior K, Lewandowska-Grygiel M, Giannopoulos K. Tyrosine kinase inhibitors in hematological malignancies. Postȩpy Higieny i Medycyny Doświadczalnej (Online). 2011;**65**:819-828

[5] Du Z, Lovly CM. Mechanisms of receptor tyrosine kinase activation in cancer. Molecular Cancer. 2018;**17**:58

[6] Al-Obeidi FA, Lam KS. Development of inhibitors for protein tyrosine kinases. Oncogene. 2000;**19**:5690-5701

[7] Wang J, Pendergast AM. The emerging role of ABL kinases in solid tumors. Trends Cancer. 2015;**1**:110-123

[8] Sirvent A, Benistant C, Roche S. Cytoplasmic signalling by the c-ABL tyrosine kinase in normal and cancer cells. Biology of the Cell. 2008;**100**:617-631

[9] Tefferi A. Myeloproliferative neoplasms: A decade of discoveries and treatment advances. American Journal of Hematology. 2016;**91**:50-58

[10] Azevedo AP, Reichert A, Afonso C, Alberca MD, Tavares P, Lima F. V280G mutation, potential role in imatinib resistance: First case report. Clinical Medicine Insights: Oncology. 2017;**11**:1179554917702870

[11] Lamontanara AJ, Georgeon S, Tria G, Svergun DI, Hantschel O. The SH2 domain of ABL kinases regulates kinase autophosphorylation by controlling activation loop accessibility. Nature Communications. 2014;**5**:5470

[12] Gromicho M, Rueff J, Rodrigues AS. Dynamics of expression of drug transporters: Methods for appraisal. Methods in Molecular Biology. 2016;**1395**:75-85

[13] Gromicho M, Magalhães M, Torres F, Dinis J, Fernandes AR, Rendeiro P, et al. Instability of mRNA expression signatures of drug transporters in chronic myeloid leukemia patients resistant to imatinib. Oncology Reports. 2013;**29**:741-750

[14] Gromicho M, Dinis J, Magalhães M, Fernandes AR, Tavares P, Laires A, et al. Development of imatinib and dasatinib resistance: Dynamics of expression of drug transporters ABCB1, ABCC1, ABCG2, MVP, and SLC22A1. Leukemia & Lymphoma. 2011;**52**:1980-1990

[15] Voisset E, Lopez S, Dubreuil P, De Sepulveda P. The tyrosine kinase FES is an essential effector of KITD816V proliferation signal. Blood. 2007;**110**:2593-2599

[16] Hellwig S, Miduturu CV, Kanda S, Zhang J, Filippakopoulos P, Salah E, et al. Small-molecule inhibitors of the c-FES protein-tyrosine kinase. Chemistry & Biology. 2012;**19**:529-540

[17] Condorelli F, Stec-Martyna E, Zaborowska J, Felli L, Gnemmi I, Ponassi M, et al. Role of the nonreceptor tyrosine kinase FES in cancer. Current Medicinal Chemistry. 2011;**18**:2913-2920

[18] Becerra-Díaz M, Valderrama-Carvajal H, Terrazas LI. Signal transducers and activators of transcription (STAT) family members in helminth infections. International Journal of Biological Sciences. 2011;**7**:1371-1381

[19] McLornan D, Percy M, McMullin MF. JAK2 V617F: A single mutation in the myeloproliferative group of disorders. The Ulster Medical Journal. 2006;**75**:112-119

[20] Furqan M, Mukhi N, Lee B, Liu D. Dysregulation of JAK-STAT pathway in hematological malignancies and JAK inhibitors for clinical application. Biomarker Research. 2013;**1**:5

[21] Wilks AF. The JAK kinases: Not just another kinase drug discovery target. Seminars in Cell & Developmental Biology. 2008;**19**:319-328

[22] Constantinescu SN, Girardot M, Pecquet C. Mining for JAK-STAT mutations in cancer. Trends in Biochemical Sciences. 2008;**33**:122-131

[23] Azevedo AP, Silva SN, Reichert A, Lima F, Júnior E, Rueff J. Prevalence of the Janus kinase 2 Vs617F mutation in Philadelphia-negative myeloproliferative neoplasms in a Portuguese population. Biomedical Reports. 2017;**7**:370-376

[24] Pita ASA, Azevedo APDS, Reichert A, Silva CJPD, Henriques V, Mendes DS, et al. Atypical haematological presentation in a case of polycythaemia vera with a new variant mutation detected in exon 12: c.1605G>T (p.Met535Ile). Journal of Clinical Pathology. 2018;**71**:180-184

[25] Prieto-Echagüe V, Miller WT. Regulation of ACK-family nonreceptor tyrosine kinases. Journal of Signal Transduction. 2011;**2011**:742372

[26] Mahajan K, Mahajan NP. ACK1/ TNK2 tyrosine kinase: Molecular signaling and evolving role in cancers. Oncogene. 2015;**34**:4162-4167

[27] Mócsai A, Ruland J, Tybulewicz VL. The SYK tyrosine kinase: A crucial player in diverse biological functions. Nature Reviews. Immunology. 2010;**10**:387-402

[28] Liu D, Mamorska-Dyga A. SYK inhibitors in clinical development for hematological malignancies. Journal of Hematology & Oncology. 2017;**10**:145

[29] Krisenko MO, Geahlen RL. Calling in SYK: SYK's dual role as a tumor promoter and tumor suppressor in cancer. Biochimica et Biophysica Acta. 2015;**1853**:254-263

[30] Qiu L, Wang F, Liu S, Chen XL. Current understanding of tyrosine kinase BMX in inflammation and its inhibitors. Burns & Trauma. 2014;**2**:121-124

[31] Mohamed AJ, Yu L, Bäckesjö CM, Vargas L, Faryal R, Aints A, et al. Bruton's tyrosine kinase (Btk): Function, regulation, and transformation with special emphasis on the PH domain. Immunological Reviews. 2009;**228**:58-73

[32] Mohammad DK, Nore BF, Hussain A, Gustafsson MO, Mohamed AJ, Smith CI. Dual phosphorylation of Btk by Akt/protein kinase b provides docking for 14-3-3ζ, regulates shuttling, and attenuates both tonic and induced signaling in B cells. Molecular and Cellular Biology. 2013;**33**:3214-3226

[33] Tojo A. Kinase inhibitors against hematological malignancies. Nihon Rinsho. 2014;**72**:1118-1124

[34] Yu L, Simonson OE, Mohamed AJ, Smith CI. NF-kappaB regulates

**35**

2009;**228**:9-22

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies*

[44] Elias D, Ditzel HJ. Fyn is an important molecule in cancer pathogenesis and drug resistance. Pharmacological Research.

malignancies. Biochemical and

[45] Kim MS, Kim GM, Choi YJ, Kim HJ, Kim YJ, Jin W. c-SRC activation through a TrkA and c-SRC interaction is essential for cell proliferation and hematological

Biophysical Research Communications.

[46] Warmuth M, Damoiseaux R, Liu Y, Fabbro D, Gray N. SRC family kinases: Potential targets for the treatment of human cancer and leukemia. Current Pharmaceutical Design.

[47] Pene-Dumitrescu T, Smithgall TE. Expression of a SRC family kinase in chronic myelogenous leukemia cells induces resistance to imatinib in a kinase-dependent manner. The Journal of Biological Chemistry.

[48] Zhang S, Yu D. Targeting SRC family kinases in anti-cancer therapies:

[49] Okada M. Regulation of the SRC family kinases by CSK. International Journal of Biological Sciences.

Turning promise into triumph. Trends in Pharmacological Sciences.

[50] Vardiman JW, Harris NL, Brunning RD. The World Health Organization (WHO) classification of the myeloid neoplasms. Blood.

[51] Swerdlow SH, Campo E, Harris NL, Jaffe ES, Pileri SA, Stein H, et al. WHO Classification of Tumours of Haematopioetic and Lymphoid Tissues. Lyon: World Health

2015;**100**:250-254

2013;**441**:431-437

2003;**9**:2043-2059

2010;**285**:21446-21457

2012;**33**:122-128

2012;**8**:1385-1397

2002;**100**:2292-2302

Organization; 2008

*DOI: http://dx.doi.org/10.5772/intechopen.84873*

[35] Lietha D, Cai X, Ceccarelli DF, Li Y, Schaller MD, Eck MJ. Structural basis for the autoinhibition of focal adhesion

kinase. Cell. 2007;**129**:1177-1187

[36] Yin B. Focal adhesion kinase as a target in the treatment of hematological malignancies. Leukemia Research.

[37] Carter BZ, Mak PY, Wang X, Yang H, Garcia-Manero G, Mak DH, et al. Focal adhesion kinase as a potential

[38] Sen B, Johnson FM. Regulation of SRC family kinases in human cancers. Journal of Signal Transduction.

[39] Parsons SJ, Parsons JT. SRC family kinases, key regulators of signal transduction. Oncogene.

[40] Boggon TJ, Eck MJ. Structure and regulation of SRC family kinases.

[41] Ku M, Wall M, MacKinnon RN, Walkley CR, Purton LE, Tam C, et al. SRC family kinases and their role in hematological malignancies. Leukemia

[42] Palacios EH, Weiss A. Function of the SRC-family kinases, Lck and Fyn, in T-cell development and activation. Oncogene. 2004;**23**:7990-8000

[43] Salmond RJ, Filby A, Qureshi I, Caserta S, Zamoyska R. T-cell receptor proximal signaling via the SRC-family kinases, Lck and Fyn, influences T-cell activation, differentiation, and tolerance. Immunological Reviews.

Oncogene. 2004;**23**:7918-7927

& Lymphoma. 2015;**56**:577-586

the transcription of protein tyrosine kinase TEC. The FEBS Journal.

2009;**276**:6714-6724

2011;**35**:1416-1418

2017;**16**:1133-1144

2011;**2011**:865819

2004;**23**:7906-7909

target in AML and MDS. Molecular Cancer Therapeutics. *Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies DOI: http://dx.doi.org/10.5772/intechopen.84873*

the transcription of protein tyrosine kinase TEC. The FEBS Journal. 2009;**276**:6714-6724

*Tyrosine Kinases as Druggable Targets in Cancer*

[26] Mahajan K, Mahajan NP. ACK1/ TNK2 tyrosine kinase: Molecular signaling and evolving role in cancers.

[27] Mócsai A, Ruland J, Tybulewicz VL. The SYK tyrosine kinase: A crucial player in diverse biological functions. Nature Reviews. Immunology.

[28] Liu D, Mamorska-Dyga A. SYK inhibitors in clinical development for hematological malignancies. Journal of Hematology & Oncology.

[29] Krisenko MO, Geahlen RL. Calling in SYK: SYK's dual role as a tumor promoter and tumor suppressor in cancer. Biochimica et Biophysica Acta.

[30] Qiu L, Wang F, Liu S, Chen XL. Current understanding of tyrosine kinase BMX in inflammation and its inhibitors. Burns & Trauma.

[31] Mohamed AJ, Yu L, Bäckesjö CM, Vargas L, Faryal R, Aints A, et al. Bruton's tyrosine kinase (Btk): Function, regulation, and transformation with special emphasis on the PH domain. Immunological

[32] Mohammad DK, Nore BF, Hussain A, Gustafsson MO, Mohamed AJ, Smith CI. Dual phosphorylation of Btk by Akt/protein kinase b provides docking for 14-3-3ζ, regulates shuttling, and attenuates both tonic and induced signaling in B cells. Molecular and Cellular Biology.

[33] Tojo A. Kinase inhibitors against hematological malignancies. Nihon

[34] Yu L, Simonson OE, Mohamed AJ, Smith CI. NF-kappaB regulates

Reviews. 2009;**228**:58-73

2013;**33**:3214-3226

Rinsho. 2014;**72**:1118-1124

Oncogene. 2015;**34**:4162-4167

2010;**10**:387-402

2017;**10**:145

2015;**1853**:254-263

2014;**2**:121-124

transcription (STAT) family members in helminth infections. International Journal of Biological Sciences.

[19] McLornan D, Percy M, McMullin MF. JAK2 V617F: A single mutation in the myeloproliferative group of disorders. The Ulster Medical Journal.

[20] Furqan M, Mukhi N, Lee B, Liu D. Dysregulation of JAK-STAT pathway in hematological malignancies and JAK inhibitors for clinical application.

[21] Wilks AF. The JAK kinases: Not just another kinase drug discovery target. Seminars in Cell & Developmental

Biomarker Research. 2013;**1**:5

Biology. 2008;**19**:319-328

[22] Constantinescu SN, Girardot M, Pecquet C. Mining for JAK-STAT mutations in cancer. Trends in

Biochemical Sciences. 2008;**33**:122-131

[23] Azevedo AP, Silva SN, Reichert A, Lima F, Júnior E, Rueff J. Prevalence

[24] Pita ASA, Azevedo APDS, Reichert A, Silva CJPD, Henriques V, Mendes DS, et al. Atypical haematological presentation in a case of polycythaemia vera with a new variant mutation detected in exon 12: c.1605G>T (p.Met535Ile). Journal of Clinical Pathology. 2018;**71**:180-184

of the Janus kinase 2 Vs617F mutation in Philadelphia-negative myeloproliferative neoplasms in a Portuguese population. Biomedical

[25] Prieto-Echagüe V, Miller WT. Regulation of ACK-family nonreceptor tyrosine kinases. Journal of Signal Transduction.

2011;**2011**:742372

Reports. 2017;**7**:370-376

[18] Becerra-Díaz M, Valderrama-Carvajal H, Terrazas LI. Signal transducers and activators of

2011;**7**:1371-1381

2006;**75**:112-119

**34**

[35] Lietha D, Cai X, Ceccarelli DF, Li Y, Schaller MD, Eck MJ. Structural basis for the autoinhibition of focal adhesion kinase. Cell. 2007;**129**:1177-1187

[36] Yin B. Focal adhesion kinase as a target in the treatment of hematological malignancies. Leukemia Research. 2011;**35**:1416-1418

[37] Carter BZ, Mak PY, Wang X, Yang H, Garcia-Manero G, Mak DH, et al. Focal adhesion kinase as a potential target in AML and MDS. Molecular Cancer Therapeutics. 2017;**16**:1133-1144

[38] Sen B, Johnson FM. Regulation of SRC family kinases in human cancers. Journal of Signal Transduction. 2011;**2011**:865819

[39] Parsons SJ, Parsons JT. SRC family kinases, key regulators of signal transduction. Oncogene. 2004;**23**:7906-7909

[40] Boggon TJ, Eck MJ. Structure and regulation of SRC family kinases. Oncogene. 2004;**23**:7918-7927

[41] Ku M, Wall M, MacKinnon RN, Walkley CR, Purton LE, Tam C, et al. SRC family kinases and their role in hematological malignancies. Leukemia & Lymphoma. 2015;**56**:577-586

[42] Palacios EH, Weiss A. Function of the SRC-family kinases, Lck and Fyn, in T-cell development and activation. Oncogene. 2004;**23**:7990-8000

[43] Salmond RJ, Filby A, Qureshi I, Caserta S, Zamoyska R. T-cell receptor proximal signaling via the SRC-family kinases, Lck and Fyn, influences T-cell activation, differentiation, and tolerance. Immunological Reviews. 2009;**228**:9-22

[44] Elias D, Ditzel HJ. Fyn is an important molecule in cancer pathogenesis and drug resistance. Pharmacological Research. 2015;**100**:250-254

[45] Kim MS, Kim GM, Choi YJ, Kim HJ, Kim YJ, Jin W. c-SRC activation through a TrkA and c-SRC interaction is essential for cell proliferation and hematological malignancies. Biochemical and Biophysical Research Communications. 2013;**441**:431-437

[46] Warmuth M, Damoiseaux R, Liu Y, Fabbro D, Gray N. SRC family kinases: Potential targets for the treatment of human cancer and leukemia. Current Pharmaceutical Design. 2003;**9**:2043-2059

[47] Pene-Dumitrescu T, Smithgall TE. Expression of a SRC family kinase in chronic myelogenous leukemia cells induces resistance to imatinib in a kinase-dependent manner. The Journal of Biological Chemistry. 2010;**285**:21446-21457

[48] Zhang S, Yu D. Targeting SRC family kinases in anti-cancer therapies: Turning promise into triumph. Trends in Pharmacological Sciences. 2012;**33**:122-128

[49] Okada M. Regulation of the SRC family kinases by CSK. International Journal of Biological Sciences. 2012;**8**:1385-1397

[50] Vardiman JW, Harris NL, Brunning RD. The World Health Organization (WHO) classification of the myeloid neoplasms. Blood. 2002;**100**:2292-2302

[51] Swerdlow SH, Campo E, Harris NL, Jaffe ES, Pileri SA, Stein H, et al. WHO Classification of Tumours of Haematopioetic and Lymphoid Tissues. Lyon: World Health Organization; 2008

[52] Tefferi A, Barbui T. Polycythemia vera and essential thrombocythemia: 2017 update on diagnosis, riskstratification, and management. American Journal of Hematology. 2017;**92**:94-108

[53] Arber DA, Orazi A, Hasserjian R, Thiele J, Borowitz MJ, Le Beau MM, et al. The 2016 revision to the World Health Organization classification of myeloid neoplasms and acute leukemia. Blood. 2016;**127**:2391-2405

[54] DAMESHEK W. Some speculations on the myeloproliferative syndromes. Blood. 1951;**6**:372-375

[55] Skoda RC, Duek A, Grisouard J. Pathogenesis of myeloproliferative neoplasms. Experimental Hematology. 2015;**43**:599-608

[56] NOWELL PC, HUNGERFORD DA. Chromosome studies on normal and leukemic human leukocytes. Journal of the National Cancer Institute. 1960;**25**:85-109

[57] Adamson JW, Fialkow PJ, Murphy S, Prchal JF, Steinmann L. Polycythemia vera: Stem-cell and probable clonal origin of the disease. The New England Journal of Medicine. 1976;**295**:913-916

[58] Fialkow PJ. Glucose-6-phosphate dehydrogenase (G-6-PD) markers in Burkitt lymphoma and other malignancies. Haematology and Blood Transfusion. 1977;**20**:297-305

[59] Fialkow PJ, Faguet GB, Jacobson RJ, Vaidya K, Murphy S. Evidence that essential thrombocythemia is a clonal disorder with origin in a multipotent stem cell. Blood. 1981;**58**:916-919

[60] Babon JJ, Lucet IS, Murphy JM, Nicola NA, Varghese LN. The molecular regulation of Janus kinase (JAK) activation. Biochemical Journal. 2014;**462**:1-13

[61] Koopmans SM, Schouten HC, van Marion AM. BCR-ABL negative myeloproliferative neoplasia: A review of involved molecular mechanisms. Histology and Histopathology. 2015;**30**:151-161

[62] Numata A, Shimoda K, Kamezaki K, Haro T, Kakumitsu H, Shide K, et al. Signal transducers and activators of transcription 3 augments the transcriptional activity of CCAAT/ enhancer-binding protein alpha in granulocyte colony-stimulating factor signaling pathway. The Journal of Biological Chemistry. 2005;**280**:12621-12629

[63] Kieslinger M, Woldman I, Moriggl R, Hofmann J, Marine JC, Ihle JN, et al. Antiapoptotic activity of Stat5 required during terminal stages of myeloid differentiation. Genes & Development. 2000;**14**:232-244

[64] Jacobson LO, Goldwasser E, Fried W, Plzak L. Role of the kidney in erythropoiesis. Nature. 1957;**179**:633-634

[65] Maxwell AP, Lappin TR, Johnston CF, Bridges JM, McGeown MG. Erythropoietin production in kidney tubular cells. British Journal of Haematology. 1990;**74**:535-539

[66] Remy I, Wilson IA, Michnick SW. Erythropoietin receptor activation by a ligand-induced conformation change. Science. 1999;**283**:990-993

[67] Witthuhn BA, Quelle FW, Silvennoinen O, Yi T, Tang B, Miura O, et al. JAK2 associates with the erythropoietin receptor and is tyrosine phosphorylated and activated following stimulation with erythropoietin. Cell. 1993;**74**:227-236

[68] Kralovics R, Buser AS, Teo SS, Coers J, Tichelli A, van der Maas AP,

**37**

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies*

the diagnosis and treatment of myeloproliferative neoplasms. Current Opinion in Hematology.

[78] Rumi E, Cazzola M. Diagnosis, risk stratification, and response

[79] Titmarsh GJ, Duncombe AS, McMullin MF, O'Rorke M, Mesa R, De Vocht F, et al. How common are myeloproliferative neoplasms? A systematic review and meta-analysis. American Journal of Hematology.

[80] Moulard O, Mehta J, Fryzek J, Olivares R, Iqbal U, Mesa RA.

vera in the European Union. European Journal of Haematology.

[81] Mehta J, Wang H, Iqbal SU, Mesa R. Epidemiology of myeloproliferative neoplasms in the United States. Leukemia & Lymphoma.

[82] Srour SA, Devesa SS, Morton LM, Check DP, Curtis RE, Linet MS, et al. Incidence and patient survival of myeloproliferative neoplasms and myelodysplastic/myeloproliferative neoplasms in the United States, 2001-12.

British Journal of Haematology.

[83] Bellanné-Chantelot C, Chaumarel I, Labopin M, Bellanger F, Barbu V, De Toma C, et al. Genetic and clinical implications of the Val617Phe JAK2 mutation in 72 families with myeloproliferative disorders.

[84] Passamonti F, Rumi E, Arcaini L, Boveri E, Elena C, Pietra D, et al. Prognostic factors for thrombosis, myelofibrosis, and leukemia in essential thrombocythemia: A study

Epidemiology of myelofibrosis, essential thrombocythemia, and polycythemia

evaluation in classical myeloproliferative neoplasms. Blood. 2017;**129**:680-692

2016;**23**:137-143

2014;**89**:581-587

2014;**92**:289-297

2014;**55**:595-600

2016;**174**:382-396

Blood. 2006;**108**:346-352

*DOI: http://dx.doi.org/10.5772/intechopen.84873*

et al. Comparison of molecular markers in a cohort of patients with chronic myeloproliferative disorders. Blood.

[69] Li Y, Hetet G, Maurer AM, Chait Y, Dhermy D, Briere J. Spontaneous megakaryocyte colony formation in myeloproliferative disorders is not neutralizable by antibodies against IL3, IL6 and GM-CSF. British Journal of Haematology. 1994;**87**:471-476

[70] Spivak JL. Myeloproliferative Neoplasms. The New England Journal of

[71] Levine RL, Pardanani A, Tefferi A, Gilliland DG. Role of JAK2 in the pathogenesis and therapy of myeloproliferative disorders. Nature Reviews. Cancer. 2007;**7**:673-683

[72] Meyer SC, Levine RL. Molecular pathways: Molecular basis for

2014;**20**:2051-2059

2004;**117**:1281-1283

2005;**92**:845-857

sensitivity and resistance to JAK kinase inhibitors. Clinical Cancer Research.

[73] Rawlings JS, Rosler KM, Harrison

[74] Espert L, Dusanter-Fourt I, Chelbi-Alix MK. Negative regulation of the JAK/STAT: Pathway implication in tumorigenesis. Bulletin du Cancer.

[75] Valentino L, Pierre J. JAK/STAT signal transduction: Regulators and implication in hematological malignancies. Biochemical Pharmacology. 2006;**71**:713-721

[76] Jones AV, Cross NC. Inherited predisposition to myeloproliferative neoplasms. Therapeutic Advances in

[77] Passamonti F, Mora B, Maffioli M. New molecular genetics in

Hematology. 2013;**4**:237-253

DA. The JAK/STAT signaling pathway. Journal of Cell Science.

Medicine. 2017;**376**:2168-2181

2003;**102**:1869-1871

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies DOI: http://dx.doi.org/10.5772/intechopen.84873*

et al. Comparison of molecular markers in a cohort of patients with chronic myeloproliferative disorders. Blood. 2003;**102**:1869-1871

*Tyrosine Kinases as Druggable Targets in Cancer*

[52] Tefferi A, Barbui T. Polycythemia vera and essential thrombocythemia: 2017 update on diagnosis, riskstratification, and management. American Journal of Hematology.

[61] Koopmans SM, Schouten HC, van Marion AM. BCR-ABL negative myeloproliferative neoplasia: A review of involved molecular mechanisms. Histology and Histopathology.

[62] Numata A, Shimoda K, Kamezaki K, Haro T, Kakumitsu H, Shide K, et al. Signal transducers and activators of transcription 3 augments the transcriptional activity of CCAAT/ enhancer-binding protein alpha in granulocyte colony-stimulating factor signaling pathway. The Journal of Biological Chemistry.

[63] Kieslinger M, Woldman I, Moriggl R, Hofmann J, Marine JC, Ihle JN, et al. Antiapoptotic activity of Stat5 required during terminal stages of myeloid differentiation. Genes & Development.

2015;**30**:151-161

2005;**280**:12621-12629

2000;**14**:232-244

1957;**179**:633-634

1999;**283**:990-993

1993;**74**:227-236

[64] Jacobson LO, Goldwasser E, Fried W, Plzak L. Role of the kidney in erythropoiesis. Nature.

[65] Maxwell AP, Lappin TR, Johnston CF, Bridges JM, McGeown MG. Erythropoietin production in kidney tubular cells. British Journal of

Haematology. 1990;**74**:535-539

[67] Witthuhn BA, Quelle FW, Silvennoinen O, Yi T, Tang B, Miura O, et al. JAK2 associates with the erythropoietin receptor and is tyrosine phosphorylated and activated following stimulation with erythropoietin. Cell.

[68] Kralovics R, Buser AS, Teo SS, Coers J, Tichelli A, van der Maas AP,

[66] Remy I, Wilson IA, Michnick SW. Erythropoietin receptor activation by a ligand-induced conformation change. Science.

[53] Arber DA, Orazi A, Hasserjian R, Thiele J, Borowitz MJ, Le Beau MM, et al. The 2016 revision to the World Health Organization classification of myeloid neoplasms and acute leukemia.

[54] DAMESHEK W. Some speculations on the myeloproliferative syndromes.

[55] Skoda RC, Duek A, Grisouard J. Pathogenesis of myeloproliferative neoplasms. Experimental Hematology.

[56] NOWELL PC, HUNGERFORD DA. Chromosome studies on normal and leukemic human leukocytes. Journal of the National Cancer Institute.

[57] Adamson JW, Fialkow PJ, Murphy S, Prchal JF, Steinmann L. Polycythemia vera: Stem-cell and probable clonal origin of the disease. The New England Journal of Medicine.

[58] Fialkow PJ. Glucose-6-phosphate dehydrogenase (G-6-PD) markers in Burkitt lymphoma and other malignancies. Haematology and Blood

[59] Fialkow PJ, Faguet GB, Jacobson RJ, Vaidya K, Murphy S. Evidence that essential thrombocythemia is a clonal disorder with origin in a multipotent stem cell. Blood. 1981;**58**:916-919

Transfusion. 1977;**20**:297-305

[60] Babon JJ, Lucet IS, Murphy JM, Nicola NA, Varghese LN. The molecular regulation of Janus kinase (JAK) activation. Biochemical Journal.

Blood. 2016;**127**:2391-2405

Blood. 1951;**6**:372-375

2015;**43**:599-608

1960;**25**:85-109

1976;**295**:913-916

2017;**92**:94-108

**36**

2014;**462**:1-13

[69] Li Y, Hetet G, Maurer AM, Chait Y, Dhermy D, Briere J. Spontaneous megakaryocyte colony formation in myeloproliferative disorders is not neutralizable by antibodies against IL3, IL6 and GM-CSF. British Journal of Haematology. 1994;**87**:471-476

[70] Spivak JL. Myeloproliferative Neoplasms. The New England Journal of Medicine. 2017;**376**:2168-2181

[71] Levine RL, Pardanani A, Tefferi A, Gilliland DG. Role of JAK2 in the pathogenesis and therapy of myeloproliferative disorders. Nature Reviews. Cancer. 2007;**7**:673-683

[72] Meyer SC, Levine RL. Molecular pathways: Molecular basis for sensitivity and resistance to JAK kinase inhibitors. Clinical Cancer Research. 2014;**20**:2051-2059

[73] Rawlings JS, Rosler KM, Harrison DA. The JAK/STAT signaling pathway. Journal of Cell Science. 2004;**117**:1281-1283

[74] Espert L, Dusanter-Fourt I, Chelbi-Alix MK. Negative regulation of the JAK/STAT: Pathway implication in tumorigenesis. Bulletin du Cancer. 2005;**92**:845-857

[75] Valentino L, Pierre J. JAK/STAT signal transduction: Regulators and implication in hematological malignancies. Biochemical Pharmacology. 2006;**71**:713-721

[76] Jones AV, Cross NC. Inherited predisposition to myeloproliferative neoplasms. Therapeutic Advances in Hematology. 2013;**4**:237-253

[77] Passamonti F, Mora B, Maffioli M. New molecular genetics in

the diagnosis and treatment of myeloproliferative neoplasms. Current Opinion in Hematology. 2016;**23**:137-143

[78] Rumi E, Cazzola M. Diagnosis, risk stratification, and response evaluation in classical myeloproliferative neoplasms. Blood. 2017;**129**:680-692

[79] Titmarsh GJ, Duncombe AS, McMullin MF, O'Rorke M, Mesa R, De Vocht F, et al. How common are myeloproliferative neoplasms? A systematic review and meta-analysis. American Journal of Hematology. 2014;**89**:581-587

[80] Moulard O, Mehta J, Fryzek J, Olivares R, Iqbal U, Mesa RA. Epidemiology of myelofibrosis, essential thrombocythemia, and polycythemia vera in the European Union. European Journal of Haematology. 2014;**92**:289-297

[81] Mehta J, Wang H, Iqbal SU, Mesa R. Epidemiology of myeloproliferative neoplasms in the United States. Leukemia & Lymphoma. 2014;**55**:595-600

[82] Srour SA, Devesa SS, Morton LM, Check DP, Curtis RE, Linet MS, et al. Incidence and patient survival of myeloproliferative neoplasms and myelodysplastic/myeloproliferative neoplasms in the United States, 2001-12. British Journal of Haematology. 2016;**174**:382-396

[83] Bellanné-Chantelot C, Chaumarel I, Labopin M, Bellanger F, Barbu V, De Toma C, et al. Genetic and clinical implications of the Val617Phe JAK2 mutation in 72 families with myeloproliferative disorders. Blood. 2006;**108**:346-352

[84] Passamonti F, Rumi E, Arcaini L, Boveri E, Elena C, Pietra D, et al. Prognostic factors for thrombosis, myelofibrosis, and leukemia in essential thrombocythemia: A study of 605 patients. Haematologica. 2008;**93**:1645-1651

[85] Bai J, Xue Y, Ye L, Yao J, Zhou C, Shao Z, et al. Risk factors of long-term incidences of thrombosis, myelofibrosis and evolution into malignance in polycythemia vera: A single center experience from China. International Journal of Hematology. 2008;**88**:530-535

[86] Hultcrantz M, Kristinsson SY, Andersson TM, Landgren O, Eloranta S, Derolf AR, et al. Patterns of survival among patients with myeloproliferative neoplasms diagnosed in Sweden from 1973 to 2008: A population-based study. Journal of Clinical Oncology. 2012;**30**:2995-3001

[87] Tefferi A, Guglielmelli P, Larson DR, Finke C, Wassie EA, Pieri L, et al. Long-term survival and blast transformation in molecularly annotated essential thrombocythemia, polycythemia vera, and myelofibrosis. Blood. 2014;**124**:2507-2513 (quiz 2615)

[88] Tefferi A. Essential thrombocythemia, polycythemia vera, and myelofibrosis: Current management and the prospect of targeted therapy. American Journal of Hematology. 2008;**83**:491-497

[89] Hultcrantz M, Wilkes SR, Kristinsson SY, Andersson TM, Derolf Å, Eloranta S, et al. Risk and cause of death in patients diagnosed with myeloproliferative neoplasms in sweden between 1973 and 2005: A populationbased study. Journal of Clinical Oncology. 2015;**33**:2288-2295

[90] Azevedo AP, Silva SN, Reichert A, Lima F, Júnior E, Rueff J. Effects of polymorphic DNA genes involved in BER and caspase pathways on the clinical outcome of myeloproliferative neoplasms under treatment with hydroxyurea. Molecular Medicine Reports. 2018;**18**:5243-5255

[91] James C, Ugo V, Le Couédic JP, Staerk J, Delhommeau F, Lacout C, et al. A unique clonal JAK2 mutation leading to constitutive signalling causes polycythaemia vera. Nature. 2005;**434**:1144-1148

[92] Kralovics R, Passamonti F, Buser AS, Teo SS, Tiedt R, Passweg JR, et al. A gain-of-function mutation of JAK2 in myeloproliferative disorders. The New England Journal of Medicine. 2005;**352**:1779-1790

[93] Levine RL, Wadleigh M, Cools J, Ebert BL, Wernig G, Huntly BJ, et al. Activating mutation in the tyrosine kinase JAK2 in polycythemia vera, essential thrombocythemia, and myeloid metaplasia with myelofibrosis. Cancer Cell. 2005;**7**:387-397

[94] Baxter EJ, Scott LM, Campbell PJ, East C, Fourouclas N, Swanton S, et al. Acquired mutation of the tyrosine kinase JAK2 in human myeloproliferative disorders. Lancet. 2005;**365**:1054-1061

[95] Tefferi A, Pardanani A. Myeloproliferative neoplasms: A contemporary review. JAMA Osncology. 2015;**1**:97-105

[96] Levine RL. Mechanisms of mutations in myeloproliferative neoplasms. Best Practice & Research. Clinical Haematology. 2009;**22**:489-494

[97] Hinds DA, Barnholt KE, Mesa RA, Kiefer AK, Do CB, Eriksson N, et al. Germ line variants predispose to both JAK2 V617F clonal hematopoiesis and myeloproliferative neoplasms. Blood. 2016;**128**:1121-1128

[98] Cazzola M, Kralovics R. From Janus kinase 2 to calreticulin: The clinically relevant genomic landscape of myeloproliferative neoplasms. Blood. 2014;**123**:3714-3719

**39**

2010;**91**:165-173

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies*

[108] Mambet C, Matei L, Necula LG, Diaconu CC. A link between the driver mutations and dysregulated apoptosis in BCR-ABL1 negative myeloproliferative neoplasms. Journal of Immunoassay & Immunochemistry.

[109] Kang MG, Choi HW, Lee JH, Choi YJ, Choi HJ, Shin JH, et al. Coexistence of JAK2 and CALR mutations and their clinical implications in patients with essential thrombocythemia. Oncotarget.

[110] Azevedo AP, Silva SN, De Lima JP, Reichert A, Lima F, Júnior E, et al. DNA repair genes polymorphisms and genetic susceptibility to Philadelphianegative myeloproliferative neoplasms in a Portuguese population: The role of base excision repair genes polymorphisms. Oncology Letters.

[111] Azevedo AP, Silva S, Reichert A, Lima F, Júnior E, Rueff J. The role of caspase genes polymorphisms in genetic susceptibility to Philadelphia-negative myeloproliferative neoplasms in a Portuguese population. Pathology & Oncology Research. 2018; 1-9. DOI:

[112] Reuther GW. Myeloproliferative neoplasms: Molecular drivers and therapeutics. Progress in Molecular Biology and Translational Science.

[113] Tefferi A, Pardanani A. JAK inhibitors in myeloproliferative neoplasms: Rationale, current data and perspective. Blood Reviews.

patients. International Journal of Clinical and Experimental Pathology.

[114] Ebid GT, Ghareeb M, Salaheldin O, Kamel MM. Prevalence of the frequency of JAK2 (V617F) mutation in different myeloproliferative disorders in Egyptian

2016;**37**:331-345

2016;**7**:57036-57049

2017;**13**:4641-4650

10.1007/s12253-018-0411-y

2016;**144**:437-484

2011;**25**:229-237

2015;**8**:11555-11559

*DOI: http://dx.doi.org/10.5772/intechopen.84873*

[99] Oh ST, Gotlib J. JAK2 V617F and beyond: Role of genetics and aberrant signaling in the pathogenesis of myeloproliferative neoplasms. Expert Review of Hematology. 2010;**3**:323-337

[100] Bench AJ, Baxter EJ, Green AR. Methods for detecting mutations in the human JAK2 gene. Methods in Molecular Biology. 2013;**967**:115-131

[101] Pikman Y, Lee BH, Mercher T, McDowell E, Ebert BL, Gozo M, et al. MPLW515L is a novel somatic activating mutation in myelofibrosis with myeloid metaplasia. PLoS Medicine. 2006;**3**:e270

[102] Tefferi A. JAK and MPL mutations in myeloid malignancies. Leukemia &

Lymphoma. 2008;**49**:388-397

2007;**356**:459-468

[103] Scott LM, Tong W, Levine RL, Scott MA, Beer PA, Stratton MR, et al. JAK2 exon 12 mutations in polycythemia vera and idiopathic erythrocytosis. The New England Journal of Medicine.

[104] Nangalia J, Green TR. The evolving genomic landscape of myeloproliferative neoplasms. Hematology. American Society of Hematology. Education Program. 2014;**2014**:287-296

[105] Langabeer SE, Andrikovics H, Asp J, Bellosillo B, Carillo S, Haslam K, et al. MPN&MPNr-EuroNet, molecular diagnostics of myeloproliferative neoplasms. European Journal of Haematology. 2015;**95**:270-279

[106] Klampfl T, Gisslinger H,

Medicine. 2013;**369**:2379-2390

[107] Delhommeau F, Jeziorowska D, Marzac C, Casadevall N. Molecular aspects of myeloproliferative neoplasms. International Journal of Hematology.

Harutyunyan AS, Nivarthi H, Rumi E, Milosevic JD, et al. Somatic mutations of calreticulin in myeloproliferative neoplasms. The New England Journal of *Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies DOI: http://dx.doi.org/10.5772/intechopen.84873*

[99] Oh ST, Gotlib J. JAK2 V617F and beyond: Role of genetics and aberrant signaling in the pathogenesis of myeloproliferative neoplasms. Expert Review of Hematology. 2010;**3**:323-337

*Tyrosine Kinases as Druggable Targets in Cancer*

[91] James C, Ugo V, Le Couédic JP, Staerk J, Delhommeau F, Lacout C, et al. A unique clonal JAK2 mutation leading to constitutive signalling causes polycythaemia vera. Nature.

[92] Kralovics R, Passamonti F, Buser AS, Teo SS, Tiedt R, Passweg JR, et al. A gain-of-function mutation of JAK2 in myeloproliferative disorders. The New England Journal of Medicine.

[93] Levine RL, Wadleigh M, Cools J, Ebert BL, Wernig G, Huntly BJ, et al. Activating mutation in the tyrosine kinase JAK2 in polycythemia vera, essential thrombocythemia, and myeloid metaplasia with myelofibrosis. Cancer Cell.

[94] Baxter EJ, Scott LM, Campbell PJ, East C, Fourouclas N, Swanton S, et al. Acquired mutation of the tyrosine kinase JAK2 in human myeloproliferative disorders. Lancet.

2005;**434**:1144-1148

2005;**352**:1779-1790

2005;**7**:387-397

2005;**365**:1054-1061

2015;**1**:97-105

2009;**22**:489-494

2016;**128**:1121-1128

2014;**123**:3714-3719

[95] Tefferi A, Pardanani A. Myeloproliferative neoplasms: A

[96] Levine RL. Mechanisms of mutations in myeloproliferative neoplasms. Best Practice & Research. Clinical Haematology.

contemporary review. JAMA Osncology.

[97] Hinds DA, Barnholt KE, Mesa RA, Kiefer AK, Do CB, Eriksson N, et al. Germ line variants predispose to both JAK2 V617F clonal hematopoiesis and myeloproliferative neoplasms. Blood.

[98] Cazzola M, Kralovics R. From Janus kinase 2 to calreticulin: The clinically relevant genomic landscape of myeloproliferative neoplasms. Blood.

of 605 patients. Haematologica.

[85] Bai J, Xue Y, Ye L, Yao J, Zhou C, Shao Z, et al. Risk factors of long-term incidences of thrombosis, myelofibrosis and evolution into malignance in polycythemia vera: A single center experience from China. International Journal of Hematology.

[86] Hultcrantz M, Kristinsson SY, Andersson TM, Landgren O, Eloranta S, Derolf AR, et al. Patterns of survival among patients with myeloproliferative neoplasms diagnosed in Sweden from 1973 to 2008: A population-based study. Journal of Clinical Oncology.

[87] Tefferi A, Guglielmelli P, Larson DR, Finke C, Wassie EA, Pieri L, et al. Long-term survival and blast transformation in molecularly

annotated essential thrombocythemia, polycythemia vera, and myelofibrosis. Blood. 2014;**124**:2507-2513 (quiz 2615)

thrombocythemia, polycythemia vera, and myelofibrosis: Current management and the prospect of targeted therapy. American Journal of Hematology.

Kristinsson SY, Andersson TM, Derolf Å, Eloranta S, et al. Risk and cause of death in patients diagnosed with myeloproliferative neoplasms in sweden between 1973 and 2005: A populationbased study. Journal of Clinical Oncology. 2015;**33**:2288-2295

[90] Azevedo AP, Silva SN, Reichert A, Lima F, Júnior E, Rueff J. Effects of polymorphic DNA genes involved in BER and caspase pathways on the clinical outcome of myeloproliferative neoplasms under treatment with hydroxyurea. Molecular Medicine Reports. 2018;**18**:5243-5255

2008;**93**:1645-1651

2008;**88**:530-535

2012;**30**:2995-3001

[88] Tefferi A. Essential

[89] Hultcrantz M, Wilkes SR,

2008;**83**:491-497

**38**

[100] Bench AJ, Baxter EJ, Green AR. Methods for detecting mutations in the human JAK2 gene. Methods in Molecular Biology. 2013;**967**:115-131

[101] Pikman Y, Lee BH, Mercher T, McDowell E, Ebert BL, Gozo M, et al. MPLW515L is a novel somatic activating mutation in myelofibrosis with myeloid metaplasia. PLoS Medicine. 2006;**3**:e270

[102] Tefferi A. JAK and MPL mutations in myeloid malignancies. Leukemia & Lymphoma. 2008;**49**:388-397

[103] Scott LM, Tong W, Levine RL, Scott MA, Beer PA, Stratton MR, et al. JAK2 exon 12 mutations in polycythemia vera and idiopathic erythrocytosis. The New England Journal of Medicine. 2007;**356**:459-468

[104] Nangalia J, Green TR. The evolving genomic landscape of myeloproliferative neoplasms. Hematology. American Society of Hematology. Education Program. 2014;**2014**:287-296

[105] Langabeer SE, Andrikovics H, Asp J, Bellosillo B, Carillo S, Haslam K, et al. MPN&MPNr-EuroNet, molecular diagnostics of myeloproliferative neoplasms. European Journal of Haematology. 2015;**95**:270-279

[106] Klampfl T, Gisslinger H, Harutyunyan AS, Nivarthi H, Rumi E, Milosevic JD, et al. Somatic mutations of calreticulin in myeloproliferative neoplasms. The New England Journal of Medicine. 2013;**369**:2379-2390

[107] Delhommeau F, Jeziorowska D, Marzac C, Casadevall N. Molecular aspects of myeloproliferative neoplasms. International Journal of Hematology. 2010;**91**:165-173

[108] Mambet C, Matei L, Necula LG, Diaconu CC. A link between the driver mutations and dysregulated apoptosis in BCR-ABL1 negative myeloproliferative neoplasms. Journal of Immunoassay & Immunochemistry. 2016;**37**:331-345

[109] Kang MG, Choi HW, Lee JH, Choi YJ, Choi HJ, Shin JH, et al. Coexistence of JAK2 and CALR mutations and their clinical implications in patients with essential thrombocythemia. Oncotarget. 2016;**7**:57036-57049

[110] Azevedo AP, Silva SN, De Lima JP, Reichert A, Lima F, Júnior E, et al. DNA repair genes polymorphisms and genetic susceptibility to Philadelphianegative myeloproliferative neoplasms in a Portuguese population: The role of base excision repair genes polymorphisms. Oncology Letters. 2017;**13**:4641-4650

[111] Azevedo AP, Silva S, Reichert A, Lima F, Júnior E, Rueff J. The role of caspase genes polymorphisms in genetic susceptibility to Philadelphia-negative myeloproliferative neoplasms in a Portuguese population. Pathology & Oncology Research. 2018; 1-9. DOI: 10.1007/s12253-018-0411-y

[112] Reuther GW. Myeloproliferative neoplasms: Molecular drivers and therapeutics. Progress in Molecular Biology and Translational Science. 2016;**144**:437-484

[113] Tefferi A, Pardanani A. JAK inhibitors in myeloproliferative neoplasms: Rationale, current data and perspective. Blood Reviews. 2011;**25**:229-237

[114] Ebid GT, Ghareeb M, Salaheldin O, Kamel MM. Prevalence of the frequency of JAK2 (V617F) mutation in different myeloproliferative disorders in Egyptian patients. International Journal of Clinical and Experimental Pathology. 2015;**8**:11555-11559

[115] Jatiani SS, Baker SJ, Silverman LR, Reddy EP. JAK/STAT pathways in cytokine signaling and myeloproliferative disorders: Approaches for targeted therapies. Genes & Cancer. 2010;**1**:979-993

[116] Anand S, Stedham F, Beer P, Gudgin E, Ortmann CA, Bench A, et al. Effects of the JAK2 mutation on the hematopoietic stem and progenitor compartment in human myeloproliferative neoplasms. Blood. 2011;**118**:177-181

[117] Steensma DP, McClure RF, Karp JE, Tefferi A, Lasho TL, Powell HL, et al. JAK2 V617F is a rare finding in de novo acute myeloid leukemia, but STAT3 activation is common and remains unexplained. Leukemia. 2006;**20**:971-978

[118] Green DR, Llambi F. Cell death signaling. Cold Spring Harb Perspect Biol. Dec 2015;**7**(12):a006080. DOI: 10.1101/cshperspect.a006080

[119] Chen E, Mullally A. How does JAK2V617F contribute to the pathogenesis of myeloproliferative neoplasms? Hematology. American Society of Hematology. Education Program. 2014;**2014**:268-276

[120] Vainchenker W, Kralovics R. Genetic basis and molecular pathophysiology of classical myeloproliferative neoplasms. Blood. 2017;**129**:667-679

[121] Michiels JJ. Clinical, pathological and molecular features of the chronic myeloproliferative disorders: MPD 2005 and beyond. Hematology. 2005;**10**(Suppl 1):215-223

[122] Yao H, Ma Y, Hong Z, Zhao L, Monaghan SA, Hu MC, et al. Activating JAK2 mutants reveal cytokine receptor coupling differences that impact outcomes in myeloproliferative neoplasm. Leukemia. 2017

[123] Vorechovsky I, Jones AV, Cross NC. Why do we see JAK2 exon 12 mutations in myeloproliferative neoplasms? Leukemia. 2013;**27**:1930-1932

[124] Godfrey AL, Chen E, Massie CE, Silber Y, Pagano F, Bellosillo B, et al. STAT1 activation in association with JAK2 exon 12 mutations. Haematologica. 2016;**101**:e15-e19

[125] Bolufer P, Barragan E, Collado M, Cervera J, López JA, Sanz MA. Influence of genetic polymorphisms on the risk of developing leukemia and on disease progression. Leukemia Research. 2006;**30**:1471-1491

[126] Beer PA, Delhommeau F, LeCouédic JP, Dawson MA, Chen E, Bareford D, et al. Two routes to leukemic transformation after a JAK2 mutation-positive myeloproliferative neoplasm. Blood. 2010;**115**:2891-2900

[127] Kilpivaara O, Levine RL. JAK2 and MPL mutations in myeloproliferative neoplasms: Discovery and science. Leukemia. 2008;**22**:1813-1817

[128] Björkholm M, Hultcrantz M, Derolf Å. Leukemic transformation in myeloproliferative neoplasms: Therapy-related or unrelated? Best Practice & Research. Clinical Haematology. 2014;**27**:141-153

[129] Rueff J, Rodrigues AS. Cancer drug resistance: A brief overview from a genetic viewpoint. Methods in Molecular Biology. 2016;**1395**:1-18

[130] Rice KL, Lin X, Wolniak K, Ebert BL, Berkofsky-Fessler W, Buzzai M, et al. Analysis of genomic aberrations and gene expression profiling identifies novel lesions and pathways in myeloproliferative neoplasms. Blood Cancer Journal. 2011;**1**:e40

**41**

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies*

[138] Scott LM. The JAK2 exon 12 mutations: A comprehensive review. American Journal of Hematology.

[139] Park CH, Lee KO, Jang JH, Jung CW, Kim JW, Kim SH, et al. High frequency of JAK2 exon 12 mutations in Korean patients with polycythaemia vera: Novel mutations and clinical significance. Journal of Clinical Pathology. 2016;**69**:737-741

[140] Passamonti F, Elena C, Schnittger S, Skoda RC, Green AR, Girodon F, et al. Molecular and clinical features of the myeloproliferative neoplasm associated with JAK2 exon 12 mutations. Blood.

[141] Pardanani A, Lasho TL, Finke C, Hanson CA, Tefferi A. Prevalence and clinicopathologic correlates of JAK2 exon 12 mutations in JAK2V617Fnegative polycythemia vera. Leukemia.

[142] Wu Z, Zhang X, Xu X, Chen Y, Hu T, Kang Z, et al. The mutation profile of JAK2 and CALR in Chinese Han patients

negative myeloproliferative neoplasms. Journal of Hematology & Oncology.

with Philadelphia chromosome-

[143] Ishii T, Bruno E, Hoffman R, Xu M. Involvement of various hematopoietic-cell lineages by the JAK2V617F mutation in polycythemia vera. Blood. 2006;**108**:3128-3134

[144] Mousinho F, Santos PSE, Azevedo AP, Pereira JM, Lemos R, Matos S, et al. Concomitant

lymphoproliferative neoplasms, distinct progenitors: A case report and review of the literature. Molecular and Clinical Oncology. 2018;**9**:347-349

[145] Mousinho F, Azevedo AP, Mendes T, Santos PSE, Cerqueira R, Matos S,et al. Concomitant

myeloproliferative and

2011;**86**:668-676

2011;**117**:2813-2816

2007;**21**:1960-1963

2014;**7**:48

*DOI: http://dx.doi.org/10.5772/intechopen.84873*

[131] Campregher PV, Santos FP, Perini GF, Hamerschlak N. Molecular biology of Philadelphia-negative myeloproliferative neoplasms. Revista Brasileira de Hematologia e Hemoterapia. 2012;**34**:150-155

[132] Vannucchi AM, Antonioli E, Guglielmelli P, Rambaldi A, Barosi G, Marchioli R, et al. Clinical profile of homozygous JAK2 617V>F mutation in patients with polycythemia vera or essential thrombocythemia. Blood.

[133] Pietra D, Li S, Brisci A, Passamonti F, Rumi E, Theocharides A, et al. Somatic mutations of JAK2 exon 12 in patients with JAK2 (V617F)-negative myeloproliferative disorders. Blood.

[134] Passamonti F, Rumi E, Pietra D, Elena C, Boveri E, Arcaini L, et al. A prospective study of 338 patients with polycythemia vera: The impact of JAK2 (V617F) allele burden and leukocytosis on fibrotic or leukemic disease transformation and vascular complications. Leukemia.

[135] Butcher CM, Hahn U, To LB, Gecz J, Wilkins EJ, Scott HS, et al. Two

in JAK2V617F-negative polycythaemia

novel JAK2 exon 12 mutations

[136] Rumi E, Pietra D, Ferretti V, Klampfl T, Harutyunyan AS, Milosevic JD, et al. JAK2 or CALR mutation status defines subtypes of essential thrombocythemia with substantially different clinical course and outcomes. Blood.

[137] Scott LM, Beer PA, Bench AJ, Erber WN, Green AR. Prevalence of JAK2 V617F and exon 12 mutations in polycythaemia vera. British Journal of Haematology. 2007;**139**:511-512

vera patients. Leukemia.

2008;**22**:870-873

2014;**123**:1544-1551

2007;**110**:840-846

2008;**111**:1686-1689

2010;**24**:1574-1579

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies DOI: http://dx.doi.org/10.5772/intechopen.84873*

[131] Campregher PV, Santos FP, Perini GF, Hamerschlak N. Molecular biology of Philadelphia-negative myeloproliferative neoplasms. Revista Brasileira de Hematologia e Hemoterapia. 2012;**34**:150-155

*Tyrosine Kinases as Druggable Targets in Cancer*

[123] Vorechovsky I, Jones AV, Cross NC. Why do we see JAK2 exon 12 mutations in myeloproliferative

[124] Godfrey AL, Chen E, Massie CE, Silber Y, Pagano F, Bellosillo B, et al. STAT1 activation in association with JAK2 exon 12 mutations. Haematologica. 2016;**101**:e15-e19

[125] Bolufer P, Barragan E, Collado M, Cervera J, López JA, Sanz MA. Influence of genetic polymorphisms on the risk of developing leukemia and on disease progression. Leukemia Research.

[126] Beer PA, Delhommeau F, LeCouédic JP, Dawson MA, Chen E, Bareford D, et al. Two routes to leukemic transformation after a

myeloproliferative neoplasm. Blood.

[127] Kilpivaara O, Levine RL. JAK2 and MPL mutations in myeloproliferative neoplasms: Discovery and science. Leukemia. 2008;**22**:1813-1817

[128] Björkholm M, Hultcrantz M, Derolf Å. Leukemic transformation in

[129] Rueff J, Rodrigues AS. Cancer drug resistance: A brief overview from a genetic viewpoint. Methods in Molecular Biology. 2016;**1395**:1-18

[130] Rice KL, Lin X, Wolniak K, Ebert BL, Berkofsky-Fessler W, Buzzai M, et al. Analysis of genomic aberrations

and gene expression profiling identifies novel lesions and pathways in myeloproliferative neoplasms. Blood

Cancer Journal. 2011;**1**:e40

myeloproliferative neoplasms: Therapy-related or unrelated? Best Practice & Research. Clinical Haematology. 2014;**27**:141-153

JAK2 mutation-positive

2010;**115**:2891-2900

2006;**30**:1471-1491

neoplasms? Leukemia. 2013;**27**:1930-1932

[115] Jatiani SS, Baker SJ, Silverman

[116] Anand S, Stedham F, Beer P, Gudgin E, Ortmann CA, Bench A, et al. Effects of the JAK2 mutation on the hematopoietic stem and progenitor compartment in human myeloproliferative neoplasms. Blood.

[117] Steensma DP, McClure RF, Karp JE, Tefferi A, Lasho TL, Powell HL, et al. JAK2 V617F is a rare finding in de novo acute myeloid leukemia, but STAT3 activation is common and remains unexplained. Leukemia.

[118] Green DR, Llambi F. Cell death signaling. Cold Spring Harb Perspect Biol. Dec 2015;**7**(12):a006080. DOI: 10.1101/cshperspect.a006080

[119] Chen E, Mullally A. How does JAK2V617F contribute to the pathogenesis of myeloproliferative neoplasms? Hematology. American Society of Hematology. Education Program. 2014;**2014**:268-276

[120] Vainchenker W, Kralovics R. Genetic basis and molecular pathophysiology of classical

2017;**129**:667-679

myeloproliferative neoplasms. Blood.

[121] Michiels JJ. Clinical, pathological and molecular features of the chronic myeloproliferative disorders: MPD 2005 and beyond. Hematology. 2005;**10**(Suppl 1):215-223

[122] Yao H, Ma Y, Hong Z, Zhao L, Monaghan SA, Hu MC, et al. Activating JAK2 mutants reveal cytokine receptor coupling differences that impact outcomes in myeloproliferative neoplasm. Leukemia. 2017

LR, Reddy EP. JAK/STAT pathways in cytokine signaling and myeloproliferative disorders: Approaches for targeted therapies. Genes & Cancer. 2010;**1**:979-993

2011;**118**:177-181

2006;**20**:971-978

**40**

[132] Vannucchi AM, Antonioli E, Guglielmelli P, Rambaldi A, Barosi G, Marchioli R, et al. Clinical profile of homozygous JAK2 617V>F mutation in patients with polycythemia vera or essential thrombocythemia. Blood. 2007;**110**:840-846

[133] Pietra D, Li S, Brisci A, Passamonti F, Rumi E, Theocharides A, et al. Somatic mutations of JAK2 exon 12 in patients with JAK2 (V617F)-negative myeloproliferative disorders. Blood. 2008;**111**:1686-1689

[134] Passamonti F, Rumi E, Pietra D, Elena C, Boveri E, Arcaini L, et al. A prospective study of 338 patients with polycythemia vera: The impact of JAK2 (V617F) allele burden and leukocytosis on fibrotic or leukemic disease transformation and vascular complications. Leukemia. 2010;**24**:1574-1579

[135] Butcher CM, Hahn U, To LB, Gecz J, Wilkins EJ, Scott HS, et al. Two novel JAK2 exon 12 mutations in JAK2V617F-negative polycythaemia vera patients. Leukemia. 2008;**22**:870-873

[136] Rumi E, Pietra D, Ferretti V, Klampfl T, Harutyunyan AS, Milosevic JD, et al. JAK2 or CALR mutation status defines subtypes of essential thrombocythemia with substantially different clinical course and outcomes. Blood. 2014;**123**:1544-1551

[137] Scott LM, Beer PA, Bench AJ, Erber WN, Green AR. Prevalence of JAK2 V617F and exon 12 mutations in polycythaemia vera. British Journal of Haematology. 2007;**139**:511-512

[138] Scott LM. The JAK2 exon 12 mutations: A comprehensive review. American Journal of Hematology. 2011;**86**:668-676

[139] Park CH, Lee KO, Jang JH, Jung CW, Kim JW, Kim SH, et al. High frequency of JAK2 exon 12 mutations in Korean patients with polycythaemia vera: Novel mutations and clinical significance. Journal of Clinical Pathology. 2016;**69**:737-741

[140] Passamonti F, Elena C, Schnittger S, Skoda RC, Green AR, Girodon F, et al. Molecular and clinical features of the myeloproliferative neoplasm associated with JAK2 exon 12 mutations. Blood. 2011;**117**:2813-2816

[141] Pardanani A, Lasho TL, Finke C, Hanson CA, Tefferi A. Prevalence and clinicopathologic correlates of JAK2 exon 12 mutations in JAK2V617Fnegative polycythemia vera. Leukemia. 2007;**21**:1960-1963

[142] Wu Z, Zhang X, Xu X, Chen Y, Hu T, Kang Z, et al. The mutation profile of JAK2 and CALR in Chinese Han patients with Philadelphia chromosomenegative myeloproliferative neoplasms. Journal of Hematology & Oncology. 2014;**7**:48

[143] Ishii T, Bruno E, Hoffman R, Xu M. Involvement of various hematopoietic-cell lineages by the JAK2V617F mutation in polycythemia vera. Blood. 2006;**108**:3128-3134

[144] Mousinho F, Santos PSE, Azevedo AP, Pereira JM, Lemos R, Matos S, et al. Concomitant myeloproliferative and lymphoproliferative neoplasms, distinct progenitors: A case report and review of the literature. Molecular and Clinical Oncology. 2018;**9**:347-349

[145] Mousinho F, Azevedo AP, Mendes T, Santos PSE, Cerqueira R, Matos S,et al. Concomitant

presence of JAK2V617F mutation and BCR-ABL translocation in two patients: A new entity or a variant of myeloproliferative neoplasms (Case report). Molecular Medicine Reports. 2018;**18**:1001-1006

[146] Vainchenker W, Constantinescu SN. JAK/STAT signaling in hematological malignancies. Oncogene. 2013;**32**:2601-2613

[147] Thomas SJ, Snowden JA, Zeidler MP, Danson SJ. The role of JAK/STAT signalling in the pathogenesis, prognosis and treatment of solid tumours. British Journal of Cancer. 2015;**113**:365-371

[148] Nielsen C, Birgens HS, Nordestgaard BG, Kjaer L, Bojesen SE. The JAK2 V617F somatic mutation, mortality and cancer risk in the general population. Haematologica. 2011;**96**:450-453

[149] Jaiswal S, Fontanillas P, Flannick J, Manning A, Grauman PV, Mar BG, et al. Age-related clonal hematopoiesis associated with adverse outcomes. The New England Journal of Medicine. 2014;**371**:2488-2498

[150] Nielsen C, Birgens HS, Nordestgaard BG, Bojesen SE. Diagnostic value of JAK2 V617F somatic mutation for myeloproliferative cancer in 49 488 individuals from the general population. British Journal of Haematology. 2013;**160**:70-79

[151] Nielsen C, Bojesen SE, Nordestgaard BG, Kofoed KF, Birgens HS. JAK2V617F somatic mutation in the general population: Myeloproliferative neoplasm development and progression rate. Haematologica. 2014;**99**:1448-1455

[152] Larsen TS, Pallisgaard N, Møller MB, Hasselbalch HC. The JAK2 V617F allele burden in essential thrombocythemia, polycythemia vera and primary myelofibrosis--impact on disease phenotype. European Journal of Haematology. 2007;**79**:508-515

[153] Vannucchi AM, Pieri L, Guglielmelli P. JAK2 allele burden in the myeloproliferative neoplasms: Effects on phenotype, prognosis and change with treatment. Therapeutic Advances in Hematology. 2011;**2**:21-32

[154] Duletić AN, Dekanić A, Hadzisejdić I, Kusen I, Matusan-Ilijas K, Grohovac D, et al. JAK2-v617F mutation is associated with clinical and laboratory features of myeloproliferative neoplasms. Collegium Antropologicum. 2012;**36**:859-865

[155] Ha JS, Kim YK, Jung SI, Jung HR, Chung IS. Correlations between Janus kinase 2 V617F allele burdens and clinicohematologic parameters in myeloproliferative neoplasms. Annals of Laboratory Medicine. 2012;**32**:385-391

[156] Pardanani A, Lasho T, McClure R, Lacy M, Tefferi A. Discordant distribution of JAK2V617F mutation in siblings with familial myeloproliferative disorders. Blood. 2006;**107**:4572-4573

[157] Jones AV, Kreil S, Zoi K, Waghorn K, Curtis C, Zhang L, et al. Widespread occurrence of the JAK2 V617F mutation in chronic myeloproliferative disorders. Blood. 2005;**106**:2162-2168

[158] Lundberg P, Takizawa H, Kubovcakova L, Guo G, Hao-Shen H, Dirnhofer S, et al. Myeloproliferative neoplasms can be initiated from a single hematopoietic stem cell expressing JAK2-V617F. The Journal of Experimental Medicine. 2014;**211**:2213-2230

[159] Bastos HN, Antão MR, Silva SN, Azevedo AP, Manita I, Teixeira V, et al. Association of polymorphisms in genes of the homologous recombination DNA repair pathway and thyroid cancer risk. Thyroid. 2009;**19**:1067-1075

**43**

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies*

transformation and new nonmyeloid

thrombocythemia and polycythemia vera. Blood. 2012;**119**:5221-5228

[168] Pardanani A, Fridley BL, Lasho TL, Gilliland DG, Tefferi A. Host genetic variation contributes to phenotypic diversity in myeloproliferative disorders.

malignancies in essential

Blood. 2008;**111**:2785-2789

2008;**22**:1790-1792

2012;**366**:787-798

2012;**366**:799-807

2017;**129**:693-703

2017;**10**:459-477

[169] Lasho TL, Tefferi A, Hood JD, Verstovsek S, Gilliland DG, Pardanani A. TG101348, a JAK2 selective antagonist, inhibits primary hematopoietic cells derived from myeloproliferative disorder patients with JAK2V617F, MPLW515K or JAK2 exon 12 mutations as well as mutation negative patients. Leukemia.

[170] Harrison C, Kiladjian JJ, Al-Ali HK, Gisslinger H, Waltzman R, Stalbovskaya V, et al. JAK inhibition with ruxolitinib versus best available therapy for myelofibrosis. The New England Journal of Medicine.

[171] Verstovsek S, Mesa RA, Gotlib J, Levy RS, Gupta V, DiPersio JF, et al. A double-blind, placebo-controlled trial of ruxolitinib for myelofibrosis. The New England Journal of Medicine.

[172] Hobbs GS, Rozelle S, Mullally A. The development and use of Janus kinase 2 inhibitors for the treatment of myeloproliferative neoplasms. Hematology/Oncology Clinics of North

[173] Vannucchi AM, Harrison CN. Emerging treatments for classical myeloproliferative neoplasms. Blood.

[174] Stahl M, Zeidan AM. Management of myelofibrosis: JAK inhibition and beyond. Expert Review of Hematology.

America. 2017;**31**:613-626

*DOI: http://dx.doi.org/10.5772/intechopen.84873*

[160] Conde J, Silva SN, Azevedo AP, Teixeira V, Pina JE, Rueff J, et al. Association of common variants in mismatch repair genes and breast cancer susceptibility: A multigene study. BMC

Cancer. 2009;**9**:344

[161] Barbui T, Vannucchi AM,

Buxhofer-Ausch V, De Stefano V, Betti S, Rambaldi A, et al. Practice-relevant revision of IPSET-thrombosis based on 1019 patients with WHO-defined essential thrombocythemia. Blood Cancer Journal. 2015;**5**:e369

[162] Guglielmelli P, Lasho TL, Rotunno G, Score J, Mannarelli C, Pancrazzi A, et al. The number of prognostically detrimental mutations and prognosis in primary myelofibrosis: An international

study of 797 patients. Leukemia.

[163] Theocharides A, Boissinot M, Girodon F, Garand R, Teo SS, Lippert E, et al. Leukemic blasts in transformed JAK2-V617F-positive myeloproliferative disorders are frequently negative for the JAK2-V617F mutation. Blood.

[164] Barbui T, Finazzi G, Falanga A. Myeloproliferative neoplasms and thrombosis. Blood. 2013;**122**:2176-2184

[165] Carobbio A, Finazzi G, Guerini V, Spinelli O, Delaini F, Marchioli R, et al.

thrombosis in essential thrombocythemia: Interaction with treatment, standard risk factors, and JAK2 mutation status. Blood.

[166] Tefferi A, Vannucchi AM. Genetic risk assessment in myeloproliferative neoplasms. Mayo Clinic Proceedings.

Leukocytosis is a risk factor for

2014;**28**:1804-1810

2007;**110**:375-379

2007;**109**:2310-2313

2017;**92**:1283-1290

[167] Hernández-Boluda JC, Pereira A, Cervantes F, Alvarez-Larrán A, Collado M, Such E, et al. A polymorphism in the XPD gene predisposes to leukemic

*Non-receptor Tyrosine Kinases Role and Significance in Hematological Malignancies DOI: http://dx.doi.org/10.5772/intechopen.84873*

[160] Conde J, Silva SN, Azevedo AP, Teixeira V, Pina JE, Rueff J, et al. Association of common variants in mismatch repair genes and breast cancer susceptibility: A multigene study. BMC Cancer. 2009;**9**:344

*Tyrosine Kinases as Druggable Targets in Cancer*

disease phenotype. European Journal of

Guglielmelli P. JAK2 allele burden in the myeloproliferative neoplasms: Effects on phenotype, prognosis and change with treatment. Therapeutic Advances

Haematology. 2007;**79**:508-515

[153] Vannucchi AM, Pieri L,

in Hematology. 2011;**2**:21-32

[154] Duletić AN, Dekanić A,

2012;**36**:859-865

2012;**32**:385-391

Hadzisejdić I, Kusen I, Matusan-Ilijas K, Grohovac D, et al. JAK2-v617F mutation is associated with clinical and laboratory features of myeloproliferative neoplasms. Collegium Antropologicum.

[155] Ha JS, Kim YK, Jung SI, Jung HR, Chung IS. Correlations between Janus kinase 2 V617F allele burdens and clinicohematologic parameters in myeloproliferative neoplasms. Annals of Laboratory Medicine.

[156] Pardanani A, Lasho T, McClure R, Lacy M, Tefferi A. Discordant distribution of JAK2V617F mutation in siblings with familial myeloproliferative disorders. Blood. 2006;**107**:4572-4573

[157] Jones AV, Kreil S, Zoi K, Waghorn K, Curtis C, Zhang L, et al. Widespread occurrence of the JAK2 V617F mutation in chronic myeloproliferative disorders.

Blood. 2005;**106**:2162-2168

2014;**211**:2213-2230

[158] Lundberg P, Takizawa H, Kubovcakova L, Guo G, Hao-Shen H, Dirnhofer S, et al. Myeloproliferative neoplasms can be initiated from a single hematopoietic stem cell expressing JAK2-V617F. The Journal of Experimental Medicine.

[159] Bastos HN, Antão MR, Silva SN, Azevedo AP, Manita I, Teixeira V, et al. Association of polymorphisms in genes of the homologous recombination DNA repair pathway and thyroid cancer risk.

Thyroid. 2009;**19**:1067-1075

presence of JAK2V617F mutation and BCR-ABL translocation in two patients: A new entity or a variant of myeloproliferative neoplasms (Case report). Molecular Medicine Reports.

[146] Vainchenker W, Constantinescu

[147] Thomas SJ, Snowden JA, Zeidler MP, Danson SJ. The role of JAK/STAT signalling in the pathogenesis, prognosis and treatment of solid tumours. British Journal of Cancer. 2015;**113**:365-371

Nordestgaard BG, Kjaer L, Bojesen SE. The JAK2 V617F somatic mutation, mortality and cancer risk in the general population. Haematologica.

[149] Jaiswal S, Fontanillas P, Flannick J, Manning A, Grauman PV, Mar BG, et al. Age-related clonal hematopoiesis associated with adverse outcomes. The New England Journal of Medicine.

Diagnostic value of JAK2 V617F somatic

Nordestgaard BG, Kofoed KF, Birgens HS. JAK2V617F somatic mutation in the general population: Myeloproliferative

[152] Larsen TS, Pallisgaard N, Møller MB, Hasselbalch HC. The JAK2 V617F allele burden in essential thrombocythemia, polycythemia vera and primary myelofibrosis--impact on

hematological malignancies. Oncogene.

2018;**18**:1001-1006

2013;**32**:2601-2613

2011;**96**:450-453

2014;**371**:2488-2498

[150] Nielsen C, Birgens HS, Nordestgaard BG, Bojesen SE.

mutation for myeloproliferative cancer in 49 488 individuals from the general population. British Journal of

Haematology. 2013;**160**:70-79

[151] Nielsen C, Bojesen SE,

neoplasm development and progression rate. Haematologica.

2014;**99**:1448-1455

SN. JAK/STAT signaling in

[148] Nielsen C, Birgens HS,

**42**

[161] Barbui T, Vannucchi AM, Buxhofer-Ausch V, De Stefano V, Betti S, Rambaldi A, et al. Practice-relevant revision of IPSET-thrombosis based on 1019 patients with WHO-defined essential thrombocythemia. Blood Cancer Journal. 2015;**5**:e369

[162] Guglielmelli P, Lasho TL, Rotunno G, Score J, Mannarelli C, Pancrazzi A, et al. The number of prognostically detrimental mutations and prognosis in primary myelofibrosis: An international study of 797 patients. Leukemia. 2014;**28**:1804-1810

[163] Theocharides A, Boissinot M, Girodon F, Garand R, Teo SS, Lippert E, et al. Leukemic blasts in transformed JAK2-V617F-positive myeloproliferative disorders are frequently negative for the JAK2-V617F mutation. Blood. 2007;**110**:375-379

[164] Barbui T, Finazzi G, Falanga A. Myeloproliferative neoplasms and thrombosis. Blood. 2013;**122**:2176-2184

[165] Carobbio A, Finazzi G, Guerini V, Spinelli O, Delaini F, Marchioli R, et al. Leukocytosis is a risk factor for thrombosis in essential thrombocythemia: Interaction with treatment, standard risk factors, and JAK2 mutation status. Blood. 2007;**109**:2310-2313

[166] Tefferi A, Vannucchi AM. Genetic risk assessment in myeloproliferative neoplasms. Mayo Clinic Proceedings. 2017;**92**:1283-1290

[167] Hernández-Boluda JC, Pereira A, Cervantes F, Alvarez-Larrán A, Collado M, Such E, et al. A polymorphism in the XPD gene predisposes to leukemic

transformation and new nonmyeloid malignancies in essential thrombocythemia and polycythemia vera. Blood. 2012;**119**:5221-5228

[168] Pardanani A, Fridley BL, Lasho TL, Gilliland DG, Tefferi A. Host genetic variation contributes to phenotypic diversity in myeloproliferative disorders. Blood. 2008;**111**:2785-2789

[169] Lasho TL, Tefferi A, Hood JD, Verstovsek S, Gilliland DG, Pardanani A. TG101348, a JAK2 selective antagonist, inhibits primary hematopoietic cells derived from myeloproliferative disorder patients with JAK2V617F, MPLW515K or JAK2 exon 12 mutations as well as mutation negative patients. Leukemia. 2008;**22**:1790-1792

[170] Harrison C, Kiladjian JJ, Al-Ali HK, Gisslinger H, Waltzman R, Stalbovskaya V, et al. JAK inhibition with ruxolitinib versus best available therapy for myelofibrosis. The New England Journal of Medicine. 2012;**366**:787-798

[171] Verstovsek S, Mesa RA, Gotlib J, Levy RS, Gupta V, DiPersio JF, et al. A double-blind, placebo-controlled trial of ruxolitinib for myelofibrosis. The New England Journal of Medicine. 2012;**366**:799-807

[172] Hobbs GS, Rozelle S, Mullally A. The development and use of Janus kinase 2 inhibitors for the treatment of myeloproliferative neoplasms. Hematology/Oncology Clinics of North America. 2017;**31**:613-626

[173] Vannucchi AM, Harrison CN. Emerging treatments for classical myeloproliferative neoplasms. Blood. 2017;**129**:693-703

[174] Stahl M, Zeidan AM. Management of myelofibrosis: JAK inhibition and beyond. Expert Review of Hematology. 2017;**10**:459-477

**45**

**Chapter 3**

**Abstract**

JAK, an Oncokinase in

Hematological Cancer

*Borja Guerra and Leandro Fernández-Pérez*

*Mercedes de Mirecki-Garrido, Patricia Martín-Rodríguez,* 

Janus kinases (JAKs) play an essential role in the regulation of cytokine signaling. They control cell survival, proliferation, differentiation, immune response, and hematopoiesis. Deregulation of JAK signaling has been associated to the pathogenesis of numerous immune-inflammatory diseases, hematological malignancies, and solid tumors. Thus, JAK proteins have emerged as attractive therapeutic targets in the last decade. The discovery of the gain-of-function JAK2 mutation (JAK2 V617F) as the main cause of polycythemia vera—a chronic myeloproliferative syndrome led to the development of the JAK inhibitor ruxolitinib. This key finding opened the door to the search for new therapeutic agents able to suppress the constitutive activation of JAK signaling in hematological cancers and other tumors. However, given the conserved nature of the kinase domain among JAK family members, and the interrelated roles of JAK kinases in many physiological processes, including hematopoiesis and immunity, the broad usage of JAK inhibitors in hematology is challenged by their narrow therapeutic window. Novel therapies are, therefore, needed. This chapter focuses on the understanding of the complex signaling of JAK proteins in cancerous cells, the various JAK aberrations implicated in myeloproliferative neoplasms, leukemia, and lymphoma, and the clinically available JAK

**Keywords:** blood cancer, hematological tumor, JAK, STAT, mutation, JAK2 V617F

The Janus kinase (JAK) signal transducer and activators of transcription (STAT) intracellular pathway connects the signaling from extracellular cytokines, hormones, and growth factors, with the nuclear transcriptional machinery [1]. It is expressed in animals from flies to humans, being highly evolutionarily conserved [2]. The cascade consists of the tyrosine kinase JAK, the transcription factor STAT, and different regulatory proteins. In mammals, four JAKs and seven STATs have been identified [3]. JAK/STAT signaling controls numerous essential cellular responses, including cell proliferation, differentiation, migration, immune response, apoptosis, and cell survival, according to the signal, cell context, and tissue [4, 5]. These cellular events are crucial to a wide range of biological functions

*Carlota Recio, Haidée Aranda-Tavío,* 

*Miguel Guerra-Rodríguez,* 

inhibitors in cancer therapy.

**1. Introduction**

#### **Chapter 3**

## JAK, an Oncokinase in Hematological Cancer

*Carlota Recio, Haidée Aranda-Tavío, Miguel Guerra-Rodríguez, Mercedes de Mirecki-Garrido, Patricia Martín-Rodríguez, Borja Guerra and Leandro Fernández-Pérez*

#### **Abstract**

Janus kinases (JAKs) play an essential role in the regulation of cytokine signaling. They control cell survival, proliferation, differentiation, immune response, and hematopoiesis. Deregulation of JAK signaling has been associated to the pathogenesis of numerous immune-inflammatory diseases, hematological malignancies, and solid tumors. Thus, JAK proteins have emerged as attractive therapeutic targets in the last decade. The discovery of the gain-of-function JAK2 mutation (JAK2 V617F) as the main cause of polycythemia vera—a chronic myeloproliferative syndrome led to the development of the JAK inhibitor ruxolitinib. This key finding opened the door to the search for new therapeutic agents able to suppress the constitutive activation of JAK signaling in hematological cancers and other tumors. However, given the conserved nature of the kinase domain among JAK family members, and the interrelated roles of JAK kinases in many physiological processes, including hematopoiesis and immunity, the broad usage of JAK inhibitors in hematology is challenged by their narrow therapeutic window. Novel therapies are, therefore, needed. This chapter focuses on the understanding of the complex signaling of JAK proteins in cancerous cells, the various JAK aberrations implicated in myeloproliferative neoplasms, leukemia, and lymphoma, and the clinically available JAK inhibitors in cancer therapy.

**Keywords:** blood cancer, hematological tumor, JAK, STAT, mutation, JAK2 V617F

#### **1. Introduction**

The Janus kinase (JAK) signal transducer and activators of transcription (STAT) intracellular pathway connects the signaling from extracellular cytokines, hormones, and growth factors, with the nuclear transcriptional machinery [1]. It is expressed in animals from flies to humans, being highly evolutionarily conserved [2]. The cascade consists of the tyrosine kinase JAK, the transcription factor STAT, and different regulatory proteins. In mammals, four JAKs and seven STATs have been identified [3]. JAK/STAT signaling controls numerous essential cellular responses, including cell proliferation, differentiation, migration, immune response, apoptosis, and cell survival, according to the signal, cell context, and tissue [4, 5]. These cellular events are crucial to a wide range of biological functions like hematopoiesis, immune development, inflammatory response, adipogenesis, and angiogenesis, among others [6]. Under normal physiological conditions, JAK/ STAT pathway signaling is strictly regulated. However, in different pathological conditions such as cancer, atherosclerosis, rheumatoid arthritis, or diabetes, an "aberrant" regulation of JAK/STAT signaling has been described [6]. Mutations on JAK proteins have been reported in certain cancers, highlighting hematological cancers (HCs). Generally, these are JAK gain-of-function mutations that promote constitutive STAT activation, which triggers tumorigenesis, high-grade inflammation, or hypergrowing, among other pathological consequences [7]. As consequence, JAK inhibitors are gaining prominence in clinical use, mainly in the treatment of HCs driven by JAK mutations, or in those tumors in which JAK/STAT pathway is determinant for the pathogenesis [8, 9]. Interestingly, not only in HCs therapy, but also in the treatment of advanced solid tumors such as pancreatic cancer and triplenegative breast cancer, and certain autoimmune and inflammatory diseases such as rheumatoid arthritis, JAK inhibitors are under clinical trial [10, 11].

### **2. The JAK/STAT pathway**

#### **2.1 JAKs**

JAK proteins are nonreceptor tyrosine kinases that are essential for the activation of signaling mediated by receptors for cytokines, hormones, or several growth factors. The family includes four 120–130 kDa proteins, named JAK1, JAK2, JAK3, and TYK2, with seven defined regions of homology, called JAK homology (JH) domains (JH1–JH7) [5] (**Figure 1**). The C-terminal region includes the kinase (JH1) and the pseudokinase (JH2) domains. JH1 domain contains tyrosine residues in the activation loop, essential for JAK activation. The pseudokinase domain JH2 is structurally analogous to JH1 and participates on its activity regulation but lacks characteristic residues of tyrosine kinases, which makes it catalytically inactive [12]. Next, the SH2-related domain is constituted by JH3 and part of JH4; this region mediates JAK docking to phosphorylated tyrosine residues [13]. The other half of JH4 to JH7 domains compose the N-terminal region, called FERM (four-point-one, ezrin, radixin, and moesin), which are involved in the association between JAK and cytokine receptors [12].

#### **2.2 STATs**

The STAT family consists of seven members, named STAT1 to STAT4, STAT5A, STAT5B, and STAT6, of 80–100 kDa, which share highly conserved homology

**47**

**2.4 Regulation of JAK/STAT pathway**

*JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

**2.3 Pathway signaling**

regions. These include (a) an N-terminal domain, (b) a spiral domain, (c) a DNA-binding domain, (d) a SH2 domain, and (e) a transactivation domain at the C-terminal end [7] (**Figure 1**). The N-terminal region is the less conserved one among the STATs, and it is implicated in some STAT dimer-dimer and other protein interactions. The spiral coiled-coil domain is responsible for many other proteinprotein interactions [6]. The STAT binding to DNA is mediated by the DNA-binding domain, which defines that STAT dimers recognize an 8- to 10-base pair inverted repeat DNA element with a consensus sequence of 5′-TTCN2–4GAA-3′. Differential binding affinity of an activated STAT dimer for a single target DNA sequence is determined by the number of nucleotides between TTC and GAA [14]. The SH2 domain is responsible to target STATs to specific tyrosine-phosphorylated peptide sequences within their binding molecules, thus controlling a broad range of intracellular signaling functions [7]. The transactivation domain holds two aminoacidic residues (tyrosine and serine) essential for STAT activity; so that JAK-promoted tyrosine phosphorylation leads to STAT dimerization, whereas STAT serine phosphorylation mediated by mitogen-activated protein kinases (MAPKs) enhances its transcriptional activity [7, 15]. All these domains are essential for STAT biological functions in response to extracellular stimuli such as cytokines or growth factors.

External stimuli (i.e., cytokines, growth factors) bind their receptors in the cellular membrane activating receptor-associated JAK autophosphorylation and subsequent activation. This event triggers a conformational change in JAK structure, which gets it ready for binding substrate and exerting its kinase activity. JAK binding sites are then exposed to the cytoplasm, where STAT monomers are found themselves in latency. STATs are recruited to the recognition areas at JAK-binding sites being phosphorylated by JAKs, which triggers their dimerization in homodimers (STAT1, STAT3, STAT4, STAT5A, and STAT5B) or heterodimers (STAT1-STAT2 and STAT1- STAT3). Consequently, active STAT dimers translocate into the nucleus where they bind to DNA, activating or repressing the transcription of their target genes [3, 6] (**Figure 2**). According to the cellular context, the external stimuli implicated, and the receptors engaged, different JAKs and STATs can be activated [16, 17] (**Table 1**). Interestingly, through a noncanonical signaling, other tyrosine kinases different from JAKs can activate STAT factors, including membrane-bound growth factor receptor tyrosine kinases (e.g., epidermal growth factor receptor—EGFR, platelet-derived growth factor receptor—PDGFR) and nonreceptor tyrosine kinases (e.g., the proto-oncogene tyrosine kinases Src and Bcr-Abl) [2, 18]. Furthermore, STAT has been shown to be able to form dimers and exert biological activity in absence of canonical JAK tyrosine phosphorylation [19]. In fact, activated JAK2 has been reported that it can enter the nucleus where it mediates epigenetic modifications of histones [20]. Furthermore, a fraction of inactive STAT5 has been found to be localized in the nucleus (instead of in the cytoplasm as the canonical signaling describes), where it is not susceptible of being phosphorylated by tyrosine kinases, mediating chromatin stabilization [21, 22].

Owing to the implication of JAK/STAT pathway in many relevant biological processes, its endogenous regulation is tight and precise. Besides, since deregulated JAKs and STATs have been associated with several pathological disorders, most of JAK/STAT modulators have been largely assessed as interesting therapeutic approaches. One of the conventional JAK/STAT modulators is protein tyrosine

**Figure 1.** *JAKs and STATs structural domains.*

#### *JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

*Tyrosine Kinases as Druggable Targets in Cancer*

**2. The JAK/STAT pathway**

cytokine receptors [12].

**2.2 STATs**

**2.1 JAKs**

like hematopoiesis, immune development, inflammatory response, adipogenesis, and angiogenesis, among others [6]. Under normal physiological conditions, JAK/ STAT pathway signaling is strictly regulated. However, in different pathological conditions such as cancer, atherosclerosis, rheumatoid arthritis, or diabetes, an "aberrant" regulation of JAK/STAT signaling has been described [6]. Mutations on JAK proteins have been reported in certain cancers, highlighting hematological cancers (HCs). Generally, these are JAK gain-of-function mutations that promote constitutive STAT activation, which triggers tumorigenesis, high-grade inflammation, or hypergrowing, among other pathological consequences [7]. As consequence, JAK inhibitors are gaining prominence in clinical use, mainly in the treatment of HCs driven by JAK mutations, or in those tumors in which JAK/STAT pathway is determinant for the pathogenesis [8, 9]. Interestingly, not only in HCs therapy, but also in the treatment of advanced solid tumors such as pancreatic cancer and triplenegative breast cancer, and certain autoimmune and inflammatory diseases such as

rheumatoid arthritis, JAK inhibitors are under clinical trial [10, 11].

JAK proteins are nonreceptor tyrosine kinases that are essential for the activation of signaling mediated by receptors for cytokines, hormones, or several growth factors. The family includes four 120–130 kDa proteins, named JAK1, JAK2, JAK3, and TYK2, with seven defined regions of homology, called JAK homology (JH) domains (JH1–JH7) [5] (**Figure 1**). The C-terminal region includes the kinase (JH1) and the pseudokinase (JH2) domains. JH1 domain contains tyrosine residues in the activation loop, essential for JAK activation. The pseudokinase domain JH2 is structurally analogous to JH1 and participates on its activity regulation but lacks characteristic residues of tyrosine kinases, which makes it catalytically inactive [12]. Next, the SH2-related domain is constituted by JH3 and part of JH4; this region mediates JAK docking to phosphorylated tyrosine residues [13]. The other half of JH4 to JH7 domains compose the N-terminal region, called FERM (four-point-one, ezrin, radixin, and moesin), which are involved in the association between JAK and

The STAT family consists of seven members, named STAT1 to STAT4, STAT5A,

STAT5B, and STAT6, of 80–100 kDa, which share highly conserved homology

**46**

**Figure 1.**

*JAKs and STATs structural domains.*

regions. These include (a) an N-terminal domain, (b) a spiral domain, (c) a DNA-binding domain, (d) a SH2 domain, and (e) a transactivation domain at the C-terminal end [7] (**Figure 1**). The N-terminal region is the less conserved one among the STATs, and it is implicated in some STAT dimer-dimer and other protein interactions. The spiral coiled-coil domain is responsible for many other proteinprotein interactions [6]. The STAT binding to DNA is mediated by the DNA-binding domain, which defines that STAT dimers recognize an 8- to 10-base pair inverted repeat DNA element with a consensus sequence of 5′-TTCN2–4GAA-3′. Differential binding affinity of an activated STAT dimer for a single target DNA sequence is determined by the number of nucleotides between TTC and GAA [14]. The SH2 domain is responsible to target STATs to specific tyrosine-phosphorylated peptide sequences within their binding molecules, thus controlling a broad range of intracellular signaling functions [7]. The transactivation domain holds two aminoacidic residues (tyrosine and serine) essential for STAT activity; so that JAK-promoted tyrosine phosphorylation leads to STAT dimerization, whereas STAT serine phosphorylation mediated by mitogen-activated protein kinases (MAPKs) enhances its transcriptional activity [7, 15]. All these domains are essential for STAT biological functions in response to extracellular stimuli such as cytokines or growth factors.

#### **2.3 Pathway signaling**

External stimuli (i.e., cytokines, growth factors) bind their receptors in the cellular membrane activating receptor-associated JAK autophosphorylation and subsequent activation. This event triggers a conformational change in JAK structure, which gets it ready for binding substrate and exerting its kinase activity. JAK binding sites are then exposed to the cytoplasm, where STAT monomers are found themselves in latency. STATs are recruited to the recognition areas at JAK-binding sites being phosphorylated by JAKs, which triggers their dimerization in homodimers (STAT1, STAT3, STAT4, STAT5A, and STAT5B) or heterodimers (STAT1-STAT2 and STAT1- STAT3). Consequently, active STAT dimers translocate into the nucleus where they bind to DNA, activating or repressing the transcription of their target genes [3, 6] (**Figure 2**). According to the cellular context, the external stimuli implicated, and the receptors engaged, different JAKs and STATs can be activated [16, 17] (**Table 1**).

Interestingly, through a noncanonical signaling, other tyrosine kinases different from JAKs can activate STAT factors, including membrane-bound growth factor receptor tyrosine kinases (e.g., epidermal growth factor receptor—EGFR, platelet-derived growth factor receptor—PDGFR) and nonreceptor tyrosine kinases (e.g., the proto-oncogene tyrosine kinases Src and Bcr-Abl) [2, 18]. Furthermore, STAT has been shown to be able to form dimers and exert biological activity in absence of canonical JAK tyrosine phosphorylation [19]. In fact, activated JAK2 has been reported that it can enter the nucleus where it mediates epigenetic modifications of histones [20]. Furthermore, a fraction of inactive STAT5 has been found to be localized in the nucleus (instead of in the cytoplasm as the canonical signaling describes), where it is not susceptible of being phosphorylated by tyrosine kinases, mediating chromatin stabilization [21, 22].

#### **2.4 Regulation of JAK/STAT pathway**

Owing to the implication of JAK/STAT pathway in many relevant biological processes, its endogenous regulation is tight and precise. Besides, since deregulated JAKs and STATs have been associated with several pathological disorders, most of JAK/STAT modulators have been largely assessed as interesting therapeutic approaches. One of the conventional JAK/STAT modulators is protein tyrosine

**Figure 2.** *JAK/STAT signaling pathway.*


#### **Table 1.**

*Differential activation of JAK/STAT pathway upon ligand binding.*

phosphatases (PTPs), which negatively regulate the signaling of the pathway by dephosphorylating the JAK-associated receptor and/or JAK itself. Furthermore, the protein inhibitors of activated STATs (PIAS) constitute another classical group of JAK/STAT negative regulators. This family of proteins can inhibit STAT signaling

**49**

*JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

and (3) proteasomal degradation [23].

B cell numbers and aberrant myelopoiesis [27].

**4. Aberrant JAK signaling and hematological cancer development**

The multifactorial process of tumorigenesis is characterized by cellular fail in sensing and repairing DNA damage, loss of regulation of cell cycle progression and

**3. JAK in hematopoiesis**

and function by directly preventing STAT from binding DNA or indirectly inhibiting STAT dimerization [23, 24]. But doubtlessly, the most broadly studied group of negative modulators of JAK/STAT signaling is the family of Suppressor of Cytokine Signaling (SOCS) proteins [25]. The family comprises eight members (SOCS1–7 and CIS) of 20–30 kDa, which show different structural domains including a N-terminal domain of variable length, little conserved; a central Src homology region that contains an extended SH2 sequence that leads to SOCS binding to tyrosine-phosphorylated residues either on the associated receptor or at JAK protein; and a highly conserved C-terminal domain, called SOCS box [25, 26]. Furthermore, SOCS1 and SOCS3 share a small kinase inhibitory region (KIR) located at their N-terminal region, which is implicated in the inhibition of JAK-catalytic activity. SOCS proteins exert a negative feedback loop mechanism, so that activated STATs induce the expression of SOCS, which then control STAT transduction signaling (**Figure 2**). The mechanisms by which SOCS proteins suppress JAK/STAT signaling include (1) binding to JAK catalytic site and subsequent inhibition of its kinase activity; (2) competition with STAT for the binding sites on the associated receptor;

Hematopoiesis is a multistep process by which blood cells, which have a limited life span, are continuously renewed. It is initiated in the bone marrow with the proliferation and differentiation of pluripotent hematopoietic stem cells, which undergo asymmetric divisions and differentiate into lineage-committed progenitors that eventually give rise to specialized blood cells [9]. Deregulation in hematopoiesis leads to the accumulation of intermediate progenitors or mature cells in the bone marrow, blood, or lymphoid tissues driving hematological malignancies [9]. Hematopoietic cytokines including erythropoietin (EPO), thrombopoietin (TPO), granulocyte colony-stimulating factor (GM-CSF), among others, tightly regulate hematopoiesis. They maintain regular levels of blood cells or induce their production according to physiological needs. These cytokines bind to their cognate receptors at the cell membrane, which generally (except some tyrosine kinases such as c-KIT, FLT-3, or GM-CSF receptor) lack intrinsic enzymatic activity at their intracellular part. Nevertheless, these receptor chains are constitutively associated with a JAK kinase, which mediates cytokine-induced signaling [9]. During myelopoiesis, JAK2 has been found to respond upon EPO, TPO, G-CSF, GM-CSF, IL-3, and IL-5 binding, mediating myeloid cell proliferation and differentiation [9], whereas in lymphopoiesis are mainly JAK1 and JAK3, which cooperate by binding to specific cytokine receptors (IL-2R, IL-4R, IL-7R and IL-15R). It has been suggested that JAK1 functions as the primary signaling effector since JAK3 is a JAK1 scaffold [9]. Gene disruption studies have confirmed the essential role of JAK proteins in hematopoiesis. JAK1-deficient mice showed perinatal lethality and defective lymphoid development [27]. Lack of JAK2 expression resulted in an embryonic lethality due to a block in erythropoiesis but with intact lymphoid development [27]. JAK3 deficiency revealed severe combined immunodeficiency with low functional T and

#### *JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

*Tyrosine Kinases as Druggable Targets in Cancer*

**48**

**Table 1.**

**Figure 2.**

**factor**

*JAK/STAT signaling pathway.*

**Cytokine/growth** 

IL-12, G-CSF, EPO, TPO, PRL, GH

CSF-1/M-CSF, EGF, PDGF, insulin

IFN (α,β,γ) JAK1, JAK2,

IL-4, IL-13 JAK1, JAK2,

IL-6, IL-11, leptin, CT-1 JAK2, JAK1,

**Activated JAK**

IL-10 JAK1, TYK2 STAT1, STAT3 Immune regulation

IL-2, IL-7, IL-9, IL-15 JAK1, JAK3 STAT5, STAT3 Immune regulation, asthma

IL-3, IL-5, GM-CSF JAK2 STAT5 Myeloid reconstitution, asthma

JAK1 STAT1, STAT3, STAT5

STAT5, STAT3, STAT4, STAT1

STAT1, STAT3, STAT5

TYK2

JAK3

TYK2

*Differential activation of JAK/STAT pathway upon ligand binding.*

JAK2, JAK1, TYK2

**Activated STAT Therapeutic area**

STAT6 Allergy, asthma

STAT1, STAT2 Immune regulation, cancer

Immune regulation, myeloid reconstitution, anemia, platelet production, growth, aging

Inflammation, platelet production, obesity, cardiovascular disease

Cancer, diabetes

phosphatases (PTPs), which negatively regulate the signaling of the pathway by dephosphorylating the JAK-associated receptor and/or JAK itself. Furthermore, the protein inhibitors of activated STATs (PIAS) constitute another classical group of JAK/STAT negative regulators. This family of proteins can inhibit STAT signaling

and function by directly preventing STAT from binding DNA or indirectly inhibiting STAT dimerization [23, 24]. But doubtlessly, the most broadly studied group of negative modulators of JAK/STAT signaling is the family of Suppressor of Cytokine Signaling (SOCS) proteins [25]. The family comprises eight members (SOCS1–7 and CIS) of 20–30 kDa, which show different structural domains including a N-terminal domain of variable length, little conserved; a central Src homology region that contains an extended SH2 sequence that leads to SOCS binding to tyrosine-phosphorylated residues either on the associated receptor or at JAK protein; and a highly conserved C-terminal domain, called SOCS box [25, 26]. Furthermore, SOCS1 and SOCS3 share a small kinase inhibitory region (KIR) located at their N-terminal region, which is implicated in the inhibition of JAK-catalytic activity. SOCS proteins exert a negative feedback loop mechanism, so that activated STATs induce the expression of SOCS, which then control STAT transduction signaling (**Figure 2**). The mechanisms by which SOCS proteins suppress JAK/STAT signaling include (1) binding to JAK catalytic site and subsequent inhibition of its kinase activity; (2) competition with STAT for the binding sites on the associated receptor; and (3) proteasomal degradation [23].

#### **3. JAK in hematopoiesis**

Hematopoiesis is a multistep process by which blood cells, which have a limited life span, are continuously renewed. It is initiated in the bone marrow with the proliferation and differentiation of pluripotent hematopoietic stem cells, which undergo asymmetric divisions and differentiate into lineage-committed progenitors that eventually give rise to specialized blood cells [9]. Deregulation in hematopoiesis leads to the accumulation of intermediate progenitors or mature cells in the bone marrow, blood, or lymphoid tissues driving hematological malignancies [9]. Hematopoietic cytokines including erythropoietin (EPO), thrombopoietin (TPO), granulocyte colony-stimulating factor (GM-CSF), among others, tightly regulate hematopoiesis. They maintain regular levels of blood cells or induce their production according to physiological needs. These cytokines bind to their cognate receptors at the cell membrane, which generally (except some tyrosine kinases such as c-KIT, FLT-3, or GM-CSF receptor) lack intrinsic enzymatic activity at their intracellular part. Nevertheless, these receptor chains are constitutively associated with a JAK kinase, which mediates cytokine-induced signaling [9]. During myelopoiesis, JAK2 has been found to respond upon EPO, TPO, G-CSF, GM-CSF, IL-3, and IL-5 binding, mediating myeloid cell proliferation and differentiation [9], whereas in lymphopoiesis are mainly JAK1 and JAK3, which cooperate by binding to specific cytokine receptors (IL-2R, IL-4R, IL-7R and IL-15R). It has been suggested that JAK1 functions as the primary signaling effector since JAK3 is a JAK1 scaffold [9]. Gene disruption studies have confirmed the essential role of JAK proteins in hematopoiesis. JAK1-deficient mice showed perinatal lethality and defective lymphoid development [27]. Lack of JAK2 expression resulted in an embryonic lethality due to a block in erythropoiesis but with intact lymphoid development [27]. JAK3 deficiency revealed severe combined immunodeficiency with low functional T and B cell numbers and aberrant myelopoiesis [27].

#### **4. Aberrant JAK signaling and hematological cancer development**

The multifactorial process of tumorigenesis is characterized by cellular fail in sensing and repairing DNA damage, loss of regulation of cell cycle progression and apoptosis, and expression of aberrant patterns of growth signaling and angiogenesis [28, 29]. Numerous studies have provided strong evidence for the key role that JAK kinases play in hematologic cancer genesis and progression. This is not surprising considering the close relation between JAKs and cytokine and growth factor signaling, hematopoiesis, proliferation, apoptosis, and immune response, processes that, when deregulated, contribute to tumor development [29, 30]. Either gain-offunction mutations in JAKs, cognate JAK tyrosine kinases, or JAK associate receptors, the generation of fusion proteins, or the loss of negative feedback regulation of JAK signaling can contribute to constitutive and aberrant STAT signaling and therefore to oncogenesis [18]. The first evidence of the strong implication of JAK kinases in HCs was the identification of oncogenic fusion proteins involving JAK kinase domain (e.g., TEL/ETV6-JAK2) [31]. Subsequently, other JAK2 fusion proteins and JAK2 gene amplifications have been identified. However, although they were more recently discovered, JAK point somatic mutations are the most common JAK deregulations found in hematological tumors, being the mutation JAK2 V617F found in more than half of all classical myeloproliferative disorders (MPDs) [32]. Besides, other JAK mutations are associated to hematological malignancies, such as JAK1 mutations, found in 10–20% of T-ALL, and other JAK2 mutations associated to ∼20% of Down syndrome (DS)-associated B-ALL [32] (**Table 2**). Interestingly, the discovery of all these mutations has highlighted JAK proteins as potent drug targets and biomarkers for HCs.

#### **4.1 JAK2 mutations**

#### *4.1.1 JAK2V617F mutation in myeloproliferative disorders*

Myeloproliferative disorders (MPDs) are a group of chronic clonal malignancies arising from the expansion of mature hematopoietic progenitor cells [33]. The World Health Organization (WHO) distinguishes two MPDs subtypes: (a) chronic myelogenous leukemia (CML) involving the Philadelphia (Ph) chromosome, frequently associated to BCR-ABL fusion oncoprotein and (b) a set of Ph-negative MPDs syndromes mainly referred to polycythemia vera (PV), essential thrombocythemia (ET), and idiopathic myelofibrosis (IMF) [34]. Two key features of this second group are the ability of cytokine-independent blood colony formation [33, 35] and hypersensitivity to numerous cytokines [36, 37]. However, each subtype is characterized by the clonal production of different hematologic lineages. PV and ET present, for example, an increased production of platelets and red cells. Accumulating evidences over the last decade establish that Ph-negative MPDs frequently carry a JAK2 single point somatic mutation at chromosome 9p24, exon 14


**51**

**Figure 3.**

*JAK2 point mutations.*

*JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

ence on platelet number [44, 47].

(JAK2V617F—Val617Phe) [38, 39]. This genetic abnormality has stem cell nature, affecting all cells of the myeloid lineages [36, 40], whereas clonal involvement of the lymphoid lineage is controversial, and its effects are less understood. Larsen and colleagues detected the JAK2V617F mutation in both B lymphocytes and T lymphocytes in a subgroup of patients with Ph-negative MPDs. Their results suggested an early stem cell origin with both lymphoid and myeloid differentiation possibilities [41]. JAK2V617F mutation is present in 50–60% of patients with ET and IMF and in most of cases of PV [35, 36, 40]. Some reports have also related JAK2V617F mutation to other myeloid malignancies like chronic myelomonocytic leukemia (CMML), myelodysplasia (MD), and, in rare cases, acute myelogenous leukemia (AML) [42]. Additionally, in other less frequent leukemias like mediastinal B cell lymphoma and Hodgkin lymphoma, both with amplification of the JAK2V617F mutation, researchers were conscious about an epigenetic role of aberrant JAK2 kinase, leading to histone H3 phosphorylation, thereby promoting gene expression [43]. The origin of JAK2V617F mutation is localized within the pseudokinase domain, JH2 of JAK2 gene [36]. JAK2 activation requires Y1007 phosphorylation [33] and its activation is crucial for cytokine-mediated signaling from the EPO receptor and other type I cytokine receptors [44]. In this sense, JAK2V617F somatic mutation is phosphorylated at Y1007, conferring constitutive activation of JAK2 tyrosine kinase by decreasing the autoinhibitory effect of JH2, thereby recapitulating cytokine receptor downstream signaling pathways, among these STAT5 and ERK (extracellular signal-regulated kinase) [33, 35, 45] (**Figure 3**). The discovery could be performed by tyrosine kinase gene sequencing in MPD patients [35, 36] and by assessing the role of JAK2V617F mutation in different in vitro studies. Cellular transformation of cytokine-dependent cell lines like Ba/F3, Ba/F3-EpoR, and FDCP-EpoR with JAK2 mutant variant led to cytokine-independent signaling triggered by JAK2 constitutive phosphorylation and induced erythrocytosis; whereas concomitant wild-type JAK2 overexpression restored or alternatively decreased the effects of the mutation in vitro [35, 40]. Lower levels of JAK2V617F required coexpression of dimeric type 1 cytokine receptor as a scaffold for the independence of hormone signaling status in Ba/F3 cells [46]. Retroviral transplant mouse models have evidenced that JAK2V617F presence is enough for reproducing PV and IMF diseases in vivo [33, 35, 46]. However, its related effects on ET remained insufficiently understood [45], exposing no sufficient JAK2V617F influ-

Three hypotheses have been suggested for explaining the causes of phenotype variability exhibited by JAK2V617F: gene dosage background, unidentified mutations, and receptor interaction with JAK2 during myeloid and erythroid differentiation [35, 42]. In the first case, mice genotyping of the JAK2V617F gene showed increased expression of this protein in homozygote samples, leading to PV or IMF like diseases. Homozygous form of this single-point mutation is found in at least 30% of PV patients, probably due to mitotic recombination [36, 40]. On the other

#### **Table 2.**

*Mutation in human JAKs and disease association.*

#### *JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

*Tyrosine Kinases as Druggable Targets in Cancer*

targets and biomarkers for HCs.

*4.1.1 JAK2V617F mutation in myeloproliferative disorders*

**JAK Mutation Associated disease**

K539L PV

Deletion of IREED ALL

T478S, V623A AML

JAK3 A572V, V722I, P132T Acute megakaryoblastic myeloid leukemia

T875N Acute megakaryoblastic myeloid leukemia

JAK2 V617F MPDs

JAK1 A634D T-ALL

*Mutation in human JAKs and disease association.*

Myeloproliferative disorders (MPDs) are a group of chronic clonal malignancies arising from the expansion of mature hematopoietic progenitor cells [33]. The World Health Organization (WHO) distinguishes two MPDs subtypes: (a) chronic myelogenous leukemia (CML) involving the Philadelphia (Ph) chromosome, frequently associated to BCR-ABL fusion oncoprotein and (b) a set of Ph-negative MPDs syndromes mainly referred to polycythemia vera (PV), essential thrombocythemia (ET), and idiopathic myelofibrosis (IMF) [34]. Two key features of this second group are the ability of cytokine-independent blood colony formation [33, 35] and hypersensitivity to numerous cytokines [36, 37]. However, each subtype is characterized by the clonal production of different hematologic lineages. PV and ET present, for example, an increased production of platelets and red cells. Accumulating evidences over the last decade establish that Ph-negative MPDs frequently carry a JAK2 single point somatic mutation at chromosome 9p24, exon 14

**4.1 JAK2 mutations**

apoptosis, and expression of aberrant patterns of growth signaling and angiogenesis [28, 29]. Numerous studies have provided strong evidence for the key role that JAK kinases play in hematologic cancer genesis and progression. This is not surprising considering the close relation between JAKs and cytokine and growth factor signaling, hematopoiesis, proliferation, apoptosis, and immune response, processes that, when deregulated, contribute to tumor development [29, 30]. Either gain-offunction mutations in JAKs, cognate JAK tyrosine kinases, or JAK associate receptors, the generation of fusion proteins, or the loss of negative feedback regulation of JAK signaling can contribute to constitutive and aberrant STAT signaling and therefore to oncogenesis [18]. The first evidence of the strong implication of JAK kinases in HCs was the identification of oncogenic fusion proteins involving JAK kinase domain (e.g., TEL/ETV6-JAK2) [31]. Subsequently, other JAK2 fusion proteins and JAK2 gene amplifications have been identified. However, although they were more recently discovered, JAK point somatic mutations are the most common JAK deregulations found in hematological tumors, being the mutation JAK2 V617F found in more than half of all classical myeloproliferative disorders (MPDs) [32]. Besides, other JAK mutations are associated to hematological malignancies, such as JAK1 mutations, found in 10–20% of T-ALL, and other JAK2 mutations associated to ∼20% of Down syndrome (DS)-associated B-ALL [32] (**Table 2**). Interestingly, the discovery of all these mutations has highlighted JAK proteins as potent drug

**50**

**Table 2.**

(JAK2V617F—Val617Phe) [38, 39]. This genetic abnormality has stem cell nature, affecting all cells of the myeloid lineages [36, 40], whereas clonal involvement of the lymphoid lineage is controversial, and its effects are less understood. Larsen and colleagues detected the JAK2V617F mutation in both B lymphocytes and T lymphocytes in a subgroup of patients with Ph-negative MPDs. Their results suggested an early stem cell origin with both lymphoid and myeloid differentiation possibilities [41]. JAK2V617F mutation is present in 50–60% of patients with ET and IMF and in most of cases of PV [35, 36, 40]. Some reports have also related JAK2V617F mutation to other myeloid malignancies like chronic myelomonocytic leukemia (CMML), myelodysplasia (MD), and, in rare cases, acute myelogenous leukemia (AML) [42]. Additionally, in other less frequent leukemias like mediastinal B cell lymphoma and Hodgkin lymphoma, both with amplification of the JAK2V617F mutation, researchers were conscious about an epigenetic role of aberrant JAK2 kinase, leading to histone H3 phosphorylation, thereby promoting gene expression [43]. The origin of JAK2V617F mutation is localized within the pseudokinase domain, JH2 of JAK2 gene [36]. JAK2 activation requires Y1007 phosphorylation [33] and its activation is crucial for cytokine-mediated signaling from the EPO receptor and other type I cytokine receptors [44]. In this sense, JAK2V617F somatic mutation is phosphorylated at Y1007, conferring constitutive activation of JAK2 tyrosine kinase by decreasing the autoinhibitory effect of JH2, thereby recapitulating cytokine receptor downstream signaling pathways, among these STAT5 and ERK (extracellular signal-regulated kinase) [33, 35, 45] (**Figure 3**). The discovery could be performed by tyrosine kinase gene sequencing in MPD patients [35, 36] and by assessing the role of JAK2V617F mutation in different in vitro studies. Cellular transformation of cytokine-dependent cell lines like Ba/F3, Ba/F3-EpoR, and FDCP-EpoR with JAK2 mutant variant led to cytokine-independent signaling triggered by JAK2 constitutive phosphorylation and induced erythrocytosis; whereas concomitant wild-type JAK2 overexpression restored or alternatively decreased the effects of the mutation in vitro [35, 40]. Lower levels of JAK2V617F required coexpression of dimeric type 1 cytokine receptor as a scaffold for the independence of hormone signaling status in Ba/F3 cells [46]. Retroviral transplant mouse models have evidenced that JAK2V617F presence is enough for reproducing PV and IMF diseases in vivo [33, 35, 46]. However, its related effects on ET remained insufficiently understood [45], exposing no sufficient JAK2V617F influence on platelet number [44, 47].

Three hypotheses have been suggested for explaining the causes of phenotype variability exhibited by JAK2V617F: gene dosage background, unidentified mutations, and receptor interaction with JAK2 during myeloid and erythroid differentiation [35, 42]. In the first case, mice genotyping of the JAK2V617F gene showed increased expression of this protein in homozygote samples, leading to PV or IMF like diseases. Homozygous form of this single-point mutation is found in at least 30% of PV patients, probably due to mitotic recombination [36, 40]. On the other

**Figure 3.** *JAK2 point mutations.*

hand, heterozygous mice might drive the ET phenotype. In fact, data point out ET as the most heterogeneous MPD. The second hypothesis suggests that the precedence or upcoming sequence of nonidentified mutations following JAK2V617F may drive the acquisition of one or another phenotype [40, 42], thus showing genetic heterogeneity [35]. Finally, Funakoshi and colleagues proposed that cellular contextspecific receptor's interaction with JAK2V617F expression levels would determine the activated phenotype [48]. From another perspective, JAK2V617F mutation in Ph-negative MPDs leads to constitutive phosphorylation of JAK2 in the absence of EPO [36]. This event is closely linked to downstream STAT3/5 proteins phosphorylation. PV patients exhibit high STAT5 and STAT3 phosphorylation; ET patients exhibit high STAT3 but low STAT5 phosphorylation; and myelofibrosis patients exhibit both low STAT5 and STAT3 phosphorylation. Different STAT3/5 phosphorylation patterns allow the discrimination among Ph-negative MPDs [49]. As we can see, constitutive activation of JAK2–STAT5 or JAK2–STAT3 signaling is a major driver of PV, ET, and IMF [36, 49]. In short, JAK/STAT signaling pathway is demonstrated to be essential for hematologic stem cells differentiation. Focusing on JAK2 as a therapeutically valid target remains an attractive option for MPDs treatment.

#### *4.1.2 JAK2K539L mutation (exon 12 mutations) in polycythemia vera*

The JAK2V617F mutation discovery was followed by other different JAK2 gene gain-of-function mutations identification [33, 38, 50–52]. As we have already described above, most PV patients express JAK2V617F [36, 40, 52]. Nevertheless, less frequently (3–5%) PV cases harbor several exon 12 JAK2 mutations present in the linking region of JH2 and JH3 domains, encompassing a highly conserved amino acid region F537–E543 in the absence of V617F mutation. This leads to a distinct clinical syndrome with isolated erythrocytosis [43, 53]. Three of the cluster of different JAK2 exon 12 mutations [43, 51, 52] included a substitution of leucine for lysine at position 539 (539L) of JAK2 in JAK2V617F-negative PV patients or idiopathic erythrocytosis: F537-K539delinsL, H538QK539L, and K539L. They are reported to be acquired, thus explaining why they appeared in peripheral-blood granulocytes but are absent in T lymphocytes [43, 51]. Functionally, K539L exon 12 mutations modify JH2 domain, resulting in aberrant growth factor responses in Ba/F3 cells *in vitro*. This cell line was able to proliferate without the addition of IL-3 and demonstrated to have an increased phosphorylation of JAK2, ERK1/2, and STAT5, in comparison to murine cells transduced by wild-type JAK2 or V617F JAK2 [39, 51, 54]. Furthermore, these mutations discharged a myeloproliferative phenotype in a murine model, resulting in higher levels of phosphorylated JAK2 compared to those with the V617F mutation. The described consequences as well as kinetics exhibited by K539L mutations were not distinguishable from those observed for cells with the V617F mutation [51]. From a genetic point of view, unlike JAK2V617F-positive PV patients, JAK2 exon 12-mutated PV patients are often heterozygous. However, they share a similar clinical outcome [39, 51].

#### *4.1.3 JAK2T875N mutation in acute megakaryoblastic myeloid leukemia*

Acute megakaryoblastic myeloid leukemia (AMKL) is a rare subtype of acute myeloid leukemia (AML) that presents different genetic characteristics and morphological phenotypes. AMKL appears frequently in childhood but is also common in adults in their 50s or 60s [55]. Some cases are developed after chemotherapy or are the result of leukemic transformation of chronic myeloproliferative neoplasms [56]. Diverse cytogenic abnormalities are associated to AMKL that differs between children and adults. The most commonly seen aberrations in adulthood are inv(3)(q21;q26), deletions of chromosomes 5 and 7, and t(9;22)(q34;q11) [55]. Children that develop

**53**

*JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

this disease are subdivided in two groups. The first one presents constitutional trisomy 21 (Down syndrome) associated to a somatic mutation in *GATA-1* [57]. The second is represented by 1(1:22)(p13:q13) translocation that encodes a fusion protein OTT-MAL (RBM15-MKL1) [58]. Despite all these genetic factors, the fact that DS children spontaneously experiment disease remission, in most cases [57] together with the fact that models of GATA1 mutation fail in reproduce AMKL leukemogenesis [59], suggests that there should be several mechanisms contributing to AMKL promotion.

Merchel et al. were interested in the STAT5 hyperactivation observed in AML, which, in most cases, is the result of activating mutations in tyrosine kinases. In 2006, they identified a novel mutation in JAK2 studying AMKL cell lines such as CHRF-288-11, M07e, or UT7. DNA sequencing of all JAK family members in CHRF-288-11 detected a single homozygous JAK2C2624A allele. This mutation leads to a substitution of a threonine for an asparagine at position 875 of the JAK2 JH1 kinase domain (**Figure 3**). Based on the crystal structure of JAK2, T875 lies within the loop between strands β2 and β3, which could alter JH1-JH2 interface [56]. However, studying full-length JAK2 crystal structure is necessary to better comprehend the mechanism of constitutive activation of JAK2 mutants [60]. The other cell lines studied, M07e and UT7 (6-month-old and 64-year-old AMKL patients, respectively), did not express hyperactivated STAT5, which is consistent with the heterogeneity of this disorder [56]. Although the frequency of this mutation in patients remains unknown, everything points to an important role of JAK2T875N in AMKL. Indeed, this mutation constitutively activates JAK2 kinase and its downstream effectors in naturally carrying JAK2T875N mutation cells *in vitro* [56] and Ba/F3 cells transduced with EpoR or TpoR. Interestingly, this mutation conferred Ba/F3 cells the capacity of IL-3 independent growth [56, 60]. Moreover, comparative studies of Ba/F3 stably expressing JAK2 wild type or JAK2V617F, JAK2K539L, JAK2T875N mutations showed that the highest kinase activity is associated with JAK2T875N mutation followed by JAK2V617F [60]. Also, JAK2T875N expression was accompanied by significantly increased activation of pathways induced by cytokines and growth factors compared with the other mutations [60]. However, these differences were not detected in HEK293 cells expressing the same JAK2 mutants, which could be result of differences in the transduced cell type [61]. Surprisingly, the higher activation of JAK2-associated JAK2T875N mutant was not linked with the capacity of transforming erythroid progenitors in bone marrow, which showed to be the lowest among the other JAK2 mutations [60]. Moreover, expression of JAK2T875N in a murine bone marrow transplant model was able to reproduce myeloproliferative disease with some AMKL characteristics, except thrombocytosis, insinuating that other genetic

events could be involved in the promotion of the disease [56].

*4.1.4 JAK2 deletion of IREED (682–686) in acute lymphoblastic leukemia*

Children with Down syndrome have an increased risk of developing ALL apart from AMKL, but unlike AMKL favorable outcomes, Down syndrome-ALL undergo higher toxicity of chemotherapy, leading to increased morbidity and mortality compared with non-Down syndrome ALL patients [62]. Activating JAK2 mutations are detected in approximately 20% of Down syndrome-ALL patients [63]. For this reason, Malinge et al. analyzed 90 cases of acute leukemia of myeloid or B-cell origin to screen activating gene mutations based on high level gene expression. This technique allowed them to discover a novel JAK2 mutation in a Down syndrome 4-year-old patient with B-cell precursor acute lymphoblastic leukemia (BCP-ALL). This JAK2 mutation encodes a protein that lacks five amino acids

(682–686), JAK2∆IREED. They confirmed constitutive activation of JAK-STAT, ERK, and AKT signaling pathways in Ba/F3 cells artificially harboring JAK2∆IREED and

#### *JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

*Tyrosine Kinases as Druggable Targets in Cancer*

hand, heterozygous mice might drive the ET phenotype. In fact, data point out ET as the most heterogeneous MPD. The second hypothesis suggests that the precedence or upcoming sequence of nonidentified mutations following JAK2V617F may drive the acquisition of one or another phenotype [40, 42], thus showing genetic heterogeneity [35]. Finally, Funakoshi and colleagues proposed that cellular contextspecific receptor's interaction with JAK2V617F expression levels would determine the activated phenotype [48]. From another perspective, JAK2V617F mutation in Ph-negative MPDs leads to constitutive phosphorylation of JAK2 in the absence of EPO [36]. This event is closely linked to downstream STAT3/5 proteins phosphorylation. PV patients exhibit high STAT5 and STAT3 phosphorylation; ET patients exhibit high STAT3 but low STAT5 phosphorylation; and myelofibrosis patients exhibit both low STAT5 and STAT3 phosphorylation. Different STAT3/5 phosphorylation patterns allow the discrimination among Ph-negative MPDs [49]. As we can see, constitutive activation of JAK2–STAT5 or JAK2–STAT3 signaling is a major driver of PV, ET, and IMF [36, 49]. In short, JAK/STAT signaling pathway is demonstrated to be essential for hematologic stem cells differentiation. Focusing on JAK2 as a therapeutically valid target remains an attractive option for MPDs treatment.

*4.1.2 JAK2K539L mutation (exon 12 mutations) in polycythemia vera*

The JAK2V617F mutation discovery was followed by other different JAK2 gene

described above, most PV patients express JAK2V617F [36, 40, 52]. Nevertheless, less frequently (3–5%) PV cases harbor several exon 12 JAK2 mutations present in the linking region of JH2 and JH3 domains, encompassing a highly conserved amino acid region F537–E543 in the absence of V617F mutation. This leads to a distinct clinical syndrome with isolated erythrocytosis [43, 53]. Three of the cluster of different JAK2 exon 12 mutations [43, 51, 52] included a substitution of leucine for lysine at position 539 (539L) of JAK2 in JAK2V617F-negative PV patients or idiopathic erythrocytosis: F537-K539delinsL, H538QK539L, and K539L. They are reported to be acquired, thus explaining why they appeared in peripheral-blood granulocytes but are absent in T lymphocytes [43, 51]. Functionally, K539L exon 12 mutations modify JH2 domain, resulting in aberrant growth factor responses in Ba/F3 cells *in vitro*. This cell line was able to proliferate without the addition of IL-3 and demonstrated to have an increased phosphorylation of JAK2, ERK1/2, and STAT5, in comparison to murine cells transduced by wild-type JAK2 or V617F JAK2 [39, 51, 54]. Furthermore, these mutations discharged a myeloproliferative phenotype in a murine model, resulting in higher levels of phosphorylated JAK2 compared to those with the V617F mutation. The described consequences as well as kinetics exhibited by K539L mutations were not distinguishable from those observed for cells with the V617F mutation [51]. From a genetic point of view, unlike JAK2V617F-positive PV patients, JAK2 exon 12-mutated PV patients are

gain-of-function mutations identification [33, 38, 50–52]. As we have already

often heterozygous. However, they share a similar clinical outcome [39, 51].

Acute megakaryoblastic myeloid leukemia (AMKL) is a rare subtype of acute myeloid leukemia (AML) that presents different genetic characteristics and morphological phenotypes. AMKL appears frequently in childhood but is also common in adults in their 50s or 60s [55]. Some cases are developed after chemotherapy or are the result of leukemic transformation of chronic myeloproliferative neoplasms [56]. Diverse cytogenic abnormalities are associated to AMKL that differs between children and adults. The most commonly seen aberrations in adulthood are inv(3)(q21;q26), deletions of chromosomes 5 and 7, and t(9;22)(q34;q11) [55]. Children that develop

*4.1.3 JAK2T875N mutation in acute megakaryoblastic myeloid leukemia*

**52**

this disease are subdivided in two groups. The first one presents constitutional trisomy 21 (Down syndrome) associated to a somatic mutation in *GATA-1* [57]. The second is represented by 1(1:22)(p13:q13) translocation that encodes a fusion protein OTT-MAL (RBM15-MKL1) [58]. Despite all these genetic factors, the fact that DS children spontaneously experiment disease remission, in most cases [57] together with the fact that models of GATA1 mutation fail in reproduce AMKL leukemogenesis [59], suggests that there should be several mechanisms contributing to AMKL promotion.

Merchel et al. were interested in the STAT5 hyperactivation observed in AML, which, in most cases, is the result of activating mutations in tyrosine kinases. In 2006, they identified a novel mutation in JAK2 studying AMKL cell lines such as CHRF-288-11, M07e, or UT7. DNA sequencing of all JAK family members in CHRF-288-11 detected a single homozygous JAK2C2624A allele. This mutation leads to a substitution of a threonine for an asparagine at position 875 of the JAK2 JH1 kinase domain (**Figure 3**). Based on the crystal structure of JAK2, T875 lies within the loop between strands β2 and β3, which could alter JH1-JH2 interface [56]. However, studying full-length JAK2 crystal structure is necessary to better comprehend the mechanism of constitutive activation of JAK2 mutants [60]. The other cell lines studied, M07e and UT7 (6-month-old and 64-year-old AMKL patients, respectively), did not express hyperactivated STAT5, which is consistent with the heterogeneity of this disorder [56]. Although the frequency of this mutation in patients remains unknown, everything points to an important role of JAK2T875N in AMKL. Indeed, this mutation constitutively activates JAK2 kinase and its downstream effectors in naturally carrying JAK2T875N mutation cells *in vitro* [56] and Ba/F3 cells transduced with EpoR or TpoR. Interestingly, this mutation conferred Ba/F3 cells the capacity of IL-3 independent growth [56, 60]. Moreover, comparative studies of Ba/F3 stably expressing JAK2 wild type or JAK2V617F, JAK2K539L, JAK2T875N mutations showed that the highest kinase activity is associated with JAK2T875N mutation followed by JAK2V617F [60]. Also, JAK2T875N expression was accompanied by significantly increased activation of pathways induced by cytokines and growth factors compared with the other mutations [60]. However, these differences were not detected in HEK293 cells expressing the same JAK2 mutants, which could be result of differences in the transduced cell type [61]. Surprisingly, the higher activation of JAK2-associated JAK2T875N mutant was not linked with the capacity of transforming erythroid progenitors in bone marrow, which showed to be the lowest among the other JAK2 mutations [60]. Moreover, expression of JAK2T875N in a murine bone marrow transplant model was able to reproduce myeloproliferative disease with some AMKL characteristics, except thrombocytosis, insinuating that other genetic events could be involved in the promotion of the disease [56].

#### *4.1.4 JAK2 deletion of IREED (682–686) in acute lymphoblastic leukemia*

Children with Down syndrome have an increased risk of developing ALL apart from AMKL, but unlike AMKL favorable outcomes, Down syndrome-ALL undergo higher toxicity of chemotherapy, leading to increased morbidity and mortality compared with non-Down syndrome ALL patients [62]. Activating JAK2 mutations are detected in approximately 20% of Down syndrome-ALL patients [63]. For this reason, Malinge et al. analyzed 90 cases of acute leukemia of myeloid or B-cell origin to screen activating gene mutations based on high level gene expression. This technique allowed them to discover a novel JAK2 mutation in a Down syndrome 4-year-old patient with B-cell precursor acute lymphoblastic leukemia (BCP-ALL). This JAK2 mutation encodes a protein that lacks five amino acids (682–686), JAK2∆IREED. They confirmed constitutive activation of JAK-STAT, ERK, and AKT signaling pathways in Ba/F3 cells artificially harboring JAK2∆IREED and

JAK2V216F mutations. As observed for other JAK2 mutations, EpoR expression was necessary for JAK2∆IREED to transform Ba/F3 cells to growth factor independency. Remarkably, these cells were sensitive to the JAK inhibitor I. In addition, a bone marrow transplant in mice revealed that this mutation promoted MPD in the model, with increased platelet, granulocytic, and red blood cell counts. Intriguingly, EpoR, myeloproliferative leukemia (MPL), and G-CSF receptor are not transcribed in the patient's cells. Hence, which cytokine receptor chain expressed in the leukemic cells is likely to associate with the mutated JAK2 is still unclear [64]. Another important source of information was the study performed by Bercovich et al. that analyzed JAK2 DNA mutations on diagnostic bone marrow samples of 88 Down syndrome-ALL patients and 216 patients with sporadic ALL. They identified acquired somatic mutations of JAK2 in 18% of Down syndrome-ALL patients. Five different alleles were detected, affecting the same evolutionary conserved arginine residue (R683), which is predicted to be located at the pseudokinase to Src homology 2 domain interface. These mutations presented associated genotype-phenotype specificity. Jak2 mutant expression in Ba/F3 EpoR and TpoR cells conferred cytokine independent growth and constitutive activation of JAK2 and STAT5. They also described pro-B cells transduced with the R683S JAK2 as sensitive to pharmacological inhibition of JAK/STAT pathway [63]. Supporting these findings, another group recently performed a genetic study of 83 BCP-ALL cell lines, detecting activating JAK2 mutations in YCUB-5 cell line (JAK2 R683I) and KOPN49 cell line (JAK2 R683G) accompanied by RAS mutations, which point out the involvement of RAS pathway apart from JAK/STAT in the progression of the disease [65]. Furthermore, some reports showed that JAK2 and P2RY8-CRLF2 (cytokine receptor-like factor 2) mutations are rare in Japanese non-Down syndrome ALL and Down syndrome-ALL patients, while in Western countries, CRLF2 is overexpressed in approximately 50–60% of Down syndrome-ALL patients. JAK2 mutations and CRLF2 seem to act in conjunction in leukemogenesis. For this reason, it is being suggested that these genetic aberrations are related to ethnicity [63].

#### **4.2 JAK3 mutations**

As we mentioned above, JAK3 is involved in lymphocyte development and function, and to carry out its functions, JAK3 interacts with the common gamma chain of some interleukin receptors, including interleukin (IL)–2, IL-4, IL-7, IL-9, IL-15, and IL-21 [5, 66]. Recently, JAK3-activating mutations have been reported in different lymphoproliferative disorders [66–68]. Mutations within the FERM domain, essential for binding of JAK to its receptor, and defects in gamma chain of receptors involved in JAK3 signaling pathway are associated with severe combined immunodeficiency (SCID) [5] and X-linked SCID (XSCID) [69], respectively. There are several activating mutations of JAK3, which have been validated in Ba/F3 cells, including P132T, L156P, R172Q , E183G, Q501H, M511I, A572V, A573V, R657Q , and V722I [67]. Among these transforming mutations, some of them have been more extensively studied because of their frequency and pathological consequences.

#### *4.2.1 JAK3A572V, V722I, P132T mutations in acute megakaryoblastic leukemia*

In acute megakaryoblastic leukemia (AMKL), AMKL cells present constitutive STAT5 phosphorylation, which indicates an upstream tyrosine kinase activation. The identified candidate responsible of STAT5 activation was JAK3, which carried an A572V mutation in the pseudokinase JH2 [70] that negatively regulates the JH1 kinase activity. Analysis of the entire coding sequence of JAK3 in AMKL patients allowed

**55**

*JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

for the identification of two additional JAK3 mutations: V722I substitution in the JH2 pseudokinase domain and P132T change in the JH6 domain of the receptor-binding region. These mutations resulted in constitutive activation of JAK3 and phosphorylation of STAT5 and made Ba/F3 hematopoietic cell line cytokine grow independently [66, 70, 71]. However, JAK3 A572V summarized some, but not all, of the phenotypic characteristics of AMKL in a murine bone marrow transplant model, suggesting that

Natural killer/T cell lymphoma (NKTCL) is a localized (areas of Asian and South America) aggressive subtype of non-Hodgkin lymphoma, with molecular characteristics and pathogenesis quite unknown. JAK3A572V and JAK3A573V mutations, located at exon 12 in the JH2 domain, have been described associated to this disease [67, 72]. NK cells need interleukin (IL)-2 to proliferate and be activated and this cytokine mediates JAK1 and JAK3 phosphorylation. In NKTCL, JAK3A572V and JAK3A573V mutations were identified in NK-S1 and MEC04 cell lines [67, 72]. These mutations were shown to trigger constitutive phosphorylation of JAK3, STAT3 [72], and STAT5 [67], respectively, in these cell lines and the ability

AML is associated with different karyotype anomalies, and these aberrations are determinant of prognosis. An array-based analysis of human leukemia exemplars could identify the JAK3 M511I mutation [73]. It is located between the SH2 domain and the pseudokinase domain of JAK3. When JAK3M511I is introduced in 32D mouse cell line, which depends on interleukin-3 (IL-3) to grow, cells are able to survive in the absence of the cytokine and they do not differentiate in the presence of G-CSF [73]. Moreover, mice with hematopoietic stem cells infected with retrovirus encoding JAK3M511I showed a marked lymphocytosis in peripheral blood and

Considering its important role in lymphopoiesis, JAK1-activating mutations have also been described in several lymphoid neoplasms, with highest frequency (7–27%) in T-ALL, but also in B-ALL and T cell prolymphocytic leukemia, and more rarely in ALL and AML [9]. Most of these mutations occur within the pseudokinase domain of JAK1. Certainly, the oncogenic potential of JAK1 pseudokinase domain disruption had been previously predicted since introduction of a V658F mutation in JAK1 (homologous to the V617F mutation in JAK2) led to its constitutive activation [75]. Recently, the mutation JAK1A634D was identified in adult T-ALL, and it was shown to lead to constitutive JAK1 activation when overexpressed in JAK1-deficient cell lines. Furthermore, A634D was shown to induce the autonomous growth of the cytokine-dependent Ba/F3 cell line, whereas it protected the murine ALL cell line BW5147 from dexamethasone-induced apoptosis. A recent study discovered another JAK1 mutation called JAK1S646P, showing that it is an activating mutation both *in vitro* and *in vivo* in ALL [76]. The first group in reporting somatic JAK1 mutations in AML (JAK1T478S and JAK1V623A) exposed that these mutations may function as disease-modifying mutations in AML, since they do not directly induce cell transformation, but rather modify the activation of downstream signaling

other mutations may cooperate in complete AMKL transformation [70].

*4.2.2 JAK3A572V and A573V mutations in natural killer/T cell lymphoma*

of IL-2 to independently proliferate in cell culture [67].

*4.2.3 JAK3M511I mutation in AML*

spleen expansion, developing T-ALL [73, 74].

pathways in response to external stimuli [77].

**4.3 JAK1 mutations**

#### *JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

*Tyrosine Kinases as Druggable Targets in Cancer*

are related to ethnicity [63].

pathological consequences.

**4.2 JAK3 mutations**

JAK2V216F mutations. As observed for other JAK2 mutations, EpoR expression was necessary for JAK2∆IREED to transform Ba/F3 cells to growth factor independency. Remarkably, these cells were sensitive to the JAK inhibitor I. In addition, a bone marrow transplant in mice revealed that this mutation promoted MPD in the model, with increased platelet, granulocytic, and red blood cell counts. Intriguingly, EpoR, myeloproliferative leukemia (MPL), and G-CSF receptor are not transcribed in the patient's cells. Hence, which cytokine receptor chain expressed in the leukemic cells is likely to associate with the mutated JAK2 is still unclear [64]. Another important source of information was the study performed by Bercovich et al. that analyzed JAK2 DNA mutations on diagnostic bone marrow samples of 88 Down syndrome-ALL patients and 216 patients with sporadic ALL. They identified acquired somatic mutations of JAK2 in 18% of Down syndrome-ALL patients. Five different alleles were detected, affecting the same evolutionary conserved arginine residue (R683), which is predicted to be located at the pseudokinase to Src homology 2 domain interface. These mutations presented associated genotype-phenotype specificity. Jak2 mutant expression in Ba/F3 EpoR and TpoR cells conferred cytokine independent growth and constitutive activation of JAK2 and STAT5. They also described pro-B cells transduced with the R683S JAK2 as sensitive to pharmacological inhibition of JAK/STAT pathway [63]. Supporting these findings, another group recently performed a genetic study of 83 BCP-ALL cell lines, detecting activating JAK2 mutations in YCUB-5 cell line (JAK2 R683I) and KOPN49 cell line (JAK2 R683G) accompanied by RAS mutations, which point out the involvement of RAS pathway apart from JAK/STAT in the progression of the disease [65]. Furthermore, some reports showed that JAK2 and P2RY8-CRLF2 (cytokine receptor-like factor 2) mutations are rare in Japanese non-Down syndrome ALL and Down syndrome-ALL patients, while in Western countries, CRLF2 is overexpressed in approximately 50–60% of Down syndrome-ALL patients. JAK2 mutations and CRLF2 seem to act in conjunction in leukemogenesis. For this reason, it is being suggested that these genetic aberrations

As we mentioned above, JAK3 is involved in lymphocyte development and function, and to carry out its functions, JAK3 interacts with the common gamma chain of some interleukin receptors, including interleukin (IL)–2, IL-4, IL-7, IL-9, IL-15, and IL-21 [5, 66]. Recently, JAK3-activating mutations have been reported in different lymphoproliferative disorders [66–68]. Mutations within the FERM domain, essential for binding of JAK to its receptor, and defects in gamma chain of receptors involved in JAK3 signaling pathway are associated with severe combined immunodeficiency (SCID) [5] and X-linked SCID (XSCID) [69], respectively. There are several activating mutations of JAK3, which have been validated in Ba/F3 cells, including P132T, L156P, R172Q , E183G, Q501H, M511I, A572V, A573V, R657Q , and V722I [67]. Among these transforming mutations, some of them have been more extensively studied because of their frequency and

*4.2.1 JAK3A572V, V722I, P132T mutations in acute megakaryoblastic leukemia*

In acute megakaryoblastic leukemia (AMKL), AMKL cells present constitutive STAT5 phosphorylation, which indicates an upstream tyrosine kinase activation. The identified candidate responsible of STAT5 activation was JAK3, which carried an A572V mutation in the pseudokinase JH2 [70] that negatively regulates the JH1 kinase activity. Analysis of the entire coding sequence of JAK3 in AMKL patients allowed

**54**

for the identification of two additional JAK3 mutations: V722I substitution in the JH2 pseudokinase domain and P132T change in the JH6 domain of the receptor-binding region. These mutations resulted in constitutive activation of JAK3 and phosphorylation of STAT5 and made Ba/F3 hematopoietic cell line cytokine grow independently [66, 70, 71]. However, JAK3 A572V summarized some, but not all, of the phenotypic characteristics of AMKL in a murine bone marrow transplant model, suggesting that other mutations may cooperate in complete AMKL transformation [70].

#### *4.2.2 JAK3A572V and A573V mutations in natural killer/T cell lymphoma*

Natural killer/T cell lymphoma (NKTCL) is a localized (areas of Asian and South America) aggressive subtype of non-Hodgkin lymphoma, with molecular characteristics and pathogenesis quite unknown. JAK3A572V and JAK3A573V mutations, located at exon 12 in the JH2 domain, have been described associated to this disease [67, 72]. NK cells need interleukin (IL)-2 to proliferate and be activated and this cytokine mediates JAK1 and JAK3 phosphorylation. In NKTCL, JAK3A572V and JAK3A573V mutations were identified in NK-S1 and MEC04 cell lines [67, 72]. These mutations were shown to trigger constitutive phosphorylation of JAK3, STAT3 [72], and STAT5 [67], respectively, in these cell lines and the ability of IL-2 to independently proliferate in cell culture [67].

#### *4.2.3 JAK3M511I mutation in AML*

AML is associated with different karyotype anomalies, and these aberrations are determinant of prognosis. An array-based analysis of human leukemia exemplars could identify the JAK3 M511I mutation [73]. It is located between the SH2 domain and the pseudokinase domain of JAK3. When JAK3M511I is introduced in 32D mouse cell line, which depends on interleukin-3 (IL-3) to grow, cells are able to survive in the absence of the cytokine and they do not differentiate in the presence of G-CSF [73]. Moreover, mice with hematopoietic stem cells infected with retrovirus encoding JAK3M511I showed a marked lymphocytosis in peripheral blood and spleen expansion, developing T-ALL [73, 74].

#### **4.3 JAK1 mutations**

Considering its important role in lymphopoiesis, JAK1-activating mutations have also been described in several lymphoid neoplasms, with highest frequency (7–27%) in T-ALL, but also in B-ALL and T cell prolymphocytic leukemia, and more rarely in ALL and AML [9]. Most of these mutations occur within the pseudokinase domain of JAK1. Certainly, the oncogenic potential of JAK1 pseudokinase domain disruption had been previously predicted since introduction of a V658F mutation in JAK1 (homologous to the V617F mutation in JAK2) led to its constitutive activation [75]. Recently, the mutation JAK1A634D was identified in adult T-ALL, and it was shown to lead to constitutive JAK1 activation when overexpressed in JAK1-deficient cell lines. Furthermore, A634D was shown to induce the autonomous growth of the cytokine-dependent Ba/F3 cell line, whereas it protected the murine ALL cell line BW5147 from dexamethasone-induced apoptosis. A recent study discovered another JAK1 mutation called JAK1S646P, showing that it is an activating mutation both *in vitro* and *in vivo* in ALL [76]. The first group in reporting somatic JAK1 mutations in AML (JAK1T478S and JAK1V623A) exposed that these mutations may function as disease-modifying mutations in AML, since they do not directly induce cell transformation, but rather modify the activation of downstream signaling pathways in response to external stimuli [77].


**Table 3.**

*Most common JAK2 fusion proteins in hematological cancer.*

#### **4.4 JAK fusion proteins**

Historically, the identification of oncogenic fusion proteins involving JAK kinase domain entailed the first evidence of the key role of JAK kinases in HCs [31]. After this finding, acquired lesions involving JAK1, JAK2, and JAK3 (but not TYK2) have been reported in both AML and ALL. Interestingly, artificial chimeric TEL-JAK1, TEL-JAK3, and TEL-TYK2 proteins are able to sustain cytokine-independent growth in Ba/F3 cells [78] in which the expression of TEL-JAK2 protects Ba/F3 cells from IL-3 withdrawal-induced apoptotic cell death and leads to IL-3-independent growth. Furthermore, mice transplanted with bone marrow cells containing the ETV6-JAK2 fusion have been shown to develop leukemia [79]. There is no patientderived chromosomal translocation that fuses the kinase domain of JAK1, JAK3, or TYK2 to a dimerizer described so far. This is probably related to an intrinsic genetic instability of the JAK2 locus, which can otherwise also be subject to amplifications in 30–50% of Hodgkin lymphomas and primary peripheral B-cell lymphomas [80–82]. The chromosomal translocation [t(9;12) (p24;p13)] is associated with T cell childhood ALL and results in the production of the fusion protein TEL-JAK2 (also known as ETV6-JAK2), which contains the JAK2 catalytic domain (JH1) and the oligomerization domain of TEL, one of the Ets transcription factor family members [31, 83]. The TEL subunit facilitates homodimerization of TEL-JAK2 molecules, thus facilitating transphosphorylation and activation of the JAK2 kinase domains. Several analogous JAK2 fusion proteins have since been described in ALLs or AMLs, including PCM1-JAK2 [84], BCR-JAK2 [85], RPN1-JAK2 [86], SSBP2-JAK2 [87], and PAX5-JAK2 [88] (**Table 3**). In all cases, the mechanism of JAK2 activation is thought to be similar, with the JAK2 fusion partner promoting dimerization and constitutive activation of the JAK2 tyrosine kinase component of the fusion protein, which constitutively triggers several downstream signal transduction pathways, such as STAT3, STAT5 [31, 89, 90], MAP kinase [91], PI3-kinase/Akt [92, 93], and NF-kB [94] independent of the presence of anchoring receptors.

#### **5. JAK inhibitors and hematological cancer treatment**

The starting point for the development of JAK inhibitors is located in 2005 when the JAK2V617F mutation was identified as the main cause of the majority of BCR-ABL1-negative myeloproliferative neoplasms (MPNs). Subsequently, the search for JAK inhibitors, and its development, continued with the discovery of other driver mutations (calreticulin (CALR) and myeloproliferative leukemia (MPL) virus oncogene) that also produce a constitutive JAK2 activation and, thus, aberrant JAK-STAT

**57**

*JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

the present book chapter will focus on them.

**5.1 Type I inhibitors**

*5.1.1 Ruxolitinib*

signaling [35, 36, 40, 95–97]. JAK inhibitors could be classified into different groups depending on their mechanism of action/region targeted in JAK: type I (they target the ATP-binding site of JAKs in the active conformation of the kinase domain), type II (they target the ATP-binding pocket of kinase domain in inactive conformation), and allosteric inhibitors (they bind to a different site from the ATP-binding site) [98]. All JAK inhibitors that have been clinically tested are type I, so this section of

These compounds may be differentiated according to their specificity for each JAK. Most often they target JAK2, JAK1, and other kinases, such as TYK2 (e.g., ruxolitinib and momelotinib (CYT-387) or JAK3 and JAK1 (tofacitinib)). Some of them can inhibit all JAKs (e.g., gandotinib and peficitinib) and less frequently they specifically target JAK2 (e.g., pacritinib, NS-018 and CEP-33779), JAK1 (e.g., filgotinib and itacitinib) or JAK3 (e.g., decernotinib and JANEX 1) [98–100]. Type I JAK2 inhibitors are commonly used in MPNs, such as myelofibrosis (MF), polycythemia vera (PV), and essential thrombocythemia (ET) [101–104]. However, type I JAK inhibitors that target JAK1 and/or JAK3 are utilized to treat inflammation and autoimmune diseases [105]. Toxicity of type I inhibitors is also related to their specificity for the different JAKs: hematologic dyscrasia/immune suppression for JAK2 inhibitors [106] and immune suppression for JAK1 and JAK3 inhibitors [107]. At this point, it should be kept in mind that JAK2 cannot be completely long term inhibited because this will produce a severe cytopenia and even lead to aplastic anemia, since wild-type JAK2 (WT-JAK2) is indispensable for normal hematopoiesis. Thus, these inhibitors may be

therapeutically used because they only partially inhibit JAK2 *in vivo*.

Nowadays, ruxolitinib is the only type I JAK2 inhibitor that has been approved by the US Food and Drug Administration (FDA) to be used in the treatment of MF and hydroxyurea (HU)-resistant or HU-intolerant PVs [101, 108, 109]. Approval for MF was due to the two key phase 3 studies: Controlled Myelofibrosis study with Oral JAK inhibitor Treatment I and II (COMFORT-I and II) [108, 109]. In both studies, ruxolitinib was very effective in reducing spleen size and improving MF-general symptoms with dose-dependent anemia and thrombocytopenia, due to JAK2 inhibition, as the most frequent hematological side effect. However, anemia was well managed with dose adjustments and/or red blood cell transfusions [108]. Moreover, in both trials, ruxolitinib significantly reduced the risk of death [110]. In HU-refractory PVs, ruxolitinib effectively controls hematocrit, reduces spleen volume, and decreases JAK2V617F allele burden [101, 111]. Combined therapy with ruxolitinib and other JAK2 inhibitors may provide novel therapeutic strategies for the treatment of MPNs. Notably, it has been recently reported that combinations of ruxolitinib and vorinostat, a histone deacetylase (HDAC) inhibitor that downregulates JAK2 expression, acted synergistically to reduce tumor growth in several hematological cancer cell lines (B cell lymphoma, multiple myeloma, anaplastic cell lymphoma, chronic B cell leukemia, and Hodgkin lymphoma) [112]. Moreover, this synergic effect on tumor cell growth was related to reduced glucose metabolism and induced ROS production and apoptosis [112]. These findings provide the rationale to support future clinical trials evaluating ruxolitinib-vorinostat combinations in patients. This combinatorial strategy has proved effective even in CML (BCR-ABL+ myeloproliferative neoplasm). Thus, it has been shown that synergic combinations of ruxolitinib and nilotinib (a direct BCR-ABL inhibitor) profoundly inhibit JAK2

signaling [35, 36, 40, 95–97]. JAK inhibitors could be classified into different groups depending on their mechanism of action/region targeted in JAK: type I (they target the ATP-binding site of JAKs in the active conformation of the kinase domain), type II (they target the ATP-binding pocket of kinase domain in inactive conformation), and allosteric inhibitors (they bind to a different site from the ATP-binding site) [98]. All JAK inhibitors that have been clinically tested are type I, so this section of the present book chapter will focus on them.

#### **5.1 Type I inhibitors**

*Tyrosine Kinases as Druggable Targets in Cancer*

**Fusion proteins Disease** TEL-JAK2 T-ALL BCD-JAK2 Atypical CML PCM1-JAK2 AML, T-ALL RPN1-JAK2 PMF SSBP2-JAK2 B-ALL PAX5-JAK2 B-ALL

*Most common JAK2 fusion proteins in hematological cancer.*

**4.4 JAK fusion proteins**

**Table 3.**

Historically, the identification of oncogenic fusion proteins involving JAK kinase domain entailed the first evidence of the key role of JAK kinases in HCs [31]. After this finding, acquired lesions involving JAK1, JAK2, and JAK3 (but not TYK2) have been reported in both AML and ALL. Interestingly, artificial chimeric TEL-JAK1, TEL-JAK3, and TEL-TYK2 proteins are able to sustain cytokine-independent growth in Ba/F3 cells [78] in which the expression of TEL-JAK2 protects Ba/F3 cells from IL-3 withdrawal-induced apoptotic cell death and leads to IL-3-independent growth. Furthermore, mice transplanted with bone marrow cells containing the ETV6-JAK2 fusion have been shown to develop leukemia [79]. There is no patientderived chromosomal translocation that fuses the kinase domain of JAK1, JAK3, or TYK2 to a dimerizer described so far. This is probably related to an intrinsic genetic instability of the JAK2 locus, which can otherwise also be subject to amplifications in 30–50% of Hodgkin lymphomas and primary peripheral B-cell lymphomas [80–82]. The chromosomal translocation [t(9;12) (p24;p13)] is associated with T cell childhood ALL and results in the production of the fusion protein TEL-JAK2 (also known as ETV6-JAK2), which contains the JAK2 catalytic domain (JH1) and the oligomerization domain of TEL, one of the Ets transcription factor family members [31, 83]. The TEL subunit facilitates homodimerization of TEL-JAK2 molecules, thus facilitating transphosphorylation and activation of the JAK2 kinase domains. Several analogous JAK2 fusion proteins have since been described in ALLs or AMLs, including PCM1-JAK2 [84], BCR-JAK2 [85], RPN1-JAK2 [86], SSBP2-JAK2 [87], and PAX5-JAK2 [88] (**Table 3**). In all cases, the mechanism of JAK2 activation is thought to be similar, with the JAK2 fusion partner promoting dimerization and constitutive activation of the JAK2 tyrosine kinase component of the fusion protein, which constitutively triggers several downstream signal transduction pathways, such as STAT3, STAT5 [31, 89, 90], MAP kinase [91], PI3-kinase/Akt [92, 93], and NF-kB

[94] independent of the presence of anchoring receptors.

**5. JAK inhibitors and hematological cancer treatment**

The starting point for the development of JAK inhibitors is located in 2005 when the JAK2V617F mutation was identified as the main cause of the majority of BCR-ABL1-negative myeloproliferative neoplasms (MPNs). Subsequently, the search for JAK inhibitors, and its development, continued with the discovery of other driver mutations (calreticulin (CALR) and myeloproliferative leukemia (MPL) virus oncogene) that also produce a constitutive JAK2 activation and, thus, aberrant JAK-STAT

**56**

These compounds may be differentiated according to their specificity for each JAK. Most often they target JAK2, JAK1, and other kinases, such as TYK2 (e.g., ruxolitinib and momelotinib (CYT-387) or JAK3 and JAK1 (tofacitinib)). Some of them can inhibit all JAKs (e.g., gandotinib and peficitinib) and less frequently they specifically target JAK2 (e.g., pacritinib, NS-018 and CEP-33779), JAK1 (e.g., filgotinib and itacitinib) or JAK3 (e.g., decernotinib and JANEX 1) [98–100]. Type I JAK2 inhibitors are commonly used in MPNs, such as myelofibrosis (MF), polycythemia vera (PV), and essential thrombocythemia (ET) [101–104]. However, type I JAK inhibitors that target JAK1 and/or JAK3 are utilized to treat inflammation and autoimmune diseases [105]. Toxicity of type I inhibitors is also related to their specificity for the different JAKs: hematologic dyscrasia/immune suppression for JAK2 inhibitors [106] and immune suppression for JAK1 and JAK3 inhibitors [107]. At this point, it should be kept in mind that JAK2 cannot be completely long term inhibited because this will produce a severe cytopenia and even lead to aplastic anemia, since wild-type JAK2 (WT-JAK2) is indispensable for normal hematopoiesis. Thus, these inhibitors may be therapeutically used because they only partially inhibit JAK2 *in vivo*.

#### *5.1.1 Ruxolitinib*

Nowadays, ruxolitinib is the only type I JAK2 inhibitor that has been approved by the US Food and Drug Administration (FDA) to be used in the treatment of MF and hydroxyurea (HU)-resistant or HU-intolerant PVs [101, 108, 109]. Approval for MF was due to the two key phase 3 studies: Controlled Myelofibrosis study with Oral JAK inhibitor Treatment I and II (COMFORT-I and II) [108, 109]. In both studies, ruxolitinib was very effective in reducing spleen size and improving MF-general symptoms with dose-dependent anemia and thrombocytopenia, due to JAK2 inhibition, as the most frequent hematological side effect. However, anemia was well managed with dose adjustments and/or red blood cell transfusions [108]. Moreover, in both trials, ruxolitinib significantly reduced the risk of death [110]. In HU-refractory PVs, ruxolitinib effectively controls hematocrit, reduces spleen volume, and decreases JAK2V617F allele burden [101, 111]. Combined therapy with ruxolitinib and other JAK2 inhibitors may provide novel therapeutic strategies for the treatment of MPNs. Notably, it has been recently reported that combinations of ruxolitinib and vorinostat, a histone deacetylase (HDAC) inhibitor that downregulates JAK2 expression, acted synergistically to reduce tumor growth in several hematological cancer cell lines (B cell lymphoma, multiple myeloma, anaplastic cell lymphoma, chronic B cell leukemia, and Hodgkin lymphoma) [112]. Moreover, this synergic effect on tumor cell growth was related to reduced glucose metabolism and induced ROS production and apoptosis [112]. These findings provide the rationale to support future clinical trials evaluating ruxolitinib-vorinostat combinations in patients. This combinatorial strategy has proved effective even in CML (BCR-ABL+ myeloproliferative neoplasm). Thus, it has been shown that synergic combinations of ruxolitinib and nilotinib (a direct BCR-ABL inhibitor) profoundly inhibit JAK2

and STAT5 phosphorylation and induce apoptosis in primary CML CD34<sup>+</sup> cells. These effects contribute to an effective elimination of these cells *in vitro* and *in vivo* and support the current utilization of ruxolitinib/nilotinib combinations in clinical trials to eradicate persistent disease in CML patients [113]. In fact, a phase I and a phase I/II clinical studies are already underway to evaluate the potential synergic effects of ruxolitinib-tyrosine kinase inhibitors combinations, such as nilotinib/ imatinib, on eradicating CML stem/progenitor cells (ClinicalTrials.gov identifiers: NCT01702064 and NCT01751425).

#### *5.1.2 Momelotinib*

Given its potential clinical relevance, there are other type I JAK inhibitors that should be highlighted: momelotinib (CYT38) is a dual JAK1/2 inhibitor that, similar to ruxolitinib, reduces spleen size and MPN general related symptoms in intermediate or high-risk MF patients [114, 115]. Relevant, momelotinib has been shown to reduce anemia, which is a major issue in MF, so this drug might be an alternative to ruxolitinib for MPN patients with anemia. However, two phase-3 studies, SIMPLIFY-1 and SIMPLIFY-2, have reported that momelotinib does not seem to have major advantages over ruxolitinib, although it was related to less transfusion requirement [116, 117]. These findings have caused that momelotinib development has been stopped.

#### *5.1.3 Pacritinib*

Pacritinib (SB1518) is a JAK2-selective inhibitor (it does not inhibit JAK1) that also inhibits FLT3 (FMS-like tyrosine kinase 3, a key target in the therapeutics of acute myeloid leukemia), colony-stimulating factor 1 receptor (CSF1R) and interleukin-1 receptor-associated kinase 1 (IRAK1) [118]. In phase I/II studies, pacritinib, at a recommended dose of 400 mg/day, showed a good activity in MF patients with gastrointestinal alterations being the most frequent side effect [119, 120]. After these promising results, two phase-3 clinical trials (PERSIST 1 and 2) were initiated testing different pacritinib concentrations [121]. However, in 2016, FDA carried out a full clinical hold on these trials due to a suspected excess of mortality in treated patients caused by intracranial hemorrhage and cardiac events. This clinical hold was lifted by the FDA on January 2017 [121] and subsequently CTI Biopharma announced PAC203, a new trial in which different doses of pacritinib are being evaluated in MF patients with thrombocytopenia.

#### *5.1.4 NS-108*

NS-108 is a potent JAK2-selective inhibitor that also inhibits Src kinases [122]. This compound showed selectivity and high potency for JAK2V617F mutant in mouse models without producing anemia or thrombocytopenia [122]. NS-108 has been tested in a phase II trial at a recommended dose of 300 mg/day in MF patients. As previously described for other JAK2 inhibitors, NS-108 significantly reduced spleen size and improved general MF-related symptoms. However, this product was not able to significantly reduce the amount of JAK2V617F mutant cells [123].

#### *5.1.5 Gandotinib*

Gandotinib (LY2784544) is a selective and potent inhibitor of JAK2V617F [124]. This drug has been evaluated in a phase I trial for safety, tolerability, and pharmacokinetic parameters in patients with MF, PV, and ET. Treatment with this compound

**59**

*JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

**5.2 Type II inhibitors**

development and clinical use.

allosteric inhibitor in clinical development.

**5.3 Allosteric inhibitors**

**6. Conclusions**

**Acknowledgements**

Program and ULPGC, respectively.

at 120 mg/day (oral recommended phase II dose) was associated with an acceptable safety/tolerability and with clinical improvements in MPN JAK2V617F patients [103]. These findings provide rationale for further gandotinib clinical testing.

Two type II JAK2 inhibitors (NVP-BBT594 and NVP-CHZ868) have been developed. NVP-BBT594 was effective in MPN cellular models [125] and NVP-CHZ868 in preclinical mouse MPN models. However, JAK2 inhibition caused by type II inhibitors is more effective and powerful than that produced by type I inhibitors, which in turn may induce profound cytopenia, limiting its future

In this group are the so-called type III (they bind to a site close to the ATPbinding site, e.g., LS104 [126]) and type IV inhibitors (they bind to an allosteric site distant from the ATP-binding site, e.g., ON044580 [127]). Since these inhibitors do not target the ATP pocket, hypothetically, they are more specific than type I/II JAK inhibitors due to the high homology shown by the ATP-binding sites. Taking this into account, JAK allosteric inhibitors would be particularly indicated to treat MPNs related to JAK mutations (especially JAK2V617F) as an efficient inhibition of WT-JAK2 will always produce a profound cytopenia. Nowadays, there is no a JAK

In summary, JAK kinases are key proteins in the development of hematological malignancies, since different genetic alterations including fusion protein formation, gene amplification, and point mutations have been discovered in a wide array of hematological malignancies. Particularly, JAK somatic point mutations have been detected in a high proportion of HC patients. Furthermore, detection of JAK mutations is beginning to provide prognostic information. For all these reasons, manipulating JAK activity currently constitutes an interesting therapeutic strategy and an interesting biomarker in hematological cancer. A great effort has been made by researchers in the last decade to find and characterize novel JAK inhibitors that might be clinically used, and, in fact, some of them have already reached clinical evaluation. However, more efforts are needed in order to identify more JAK mutations that lead to develop more accurate therapies against specific malignancies.

We thank all the authors who contributed to the understanding of the cross-talk between the JAK kinase and hematological cancer. We apologize to those whose work deserves to be cited but unfortunately are not quoted because of space restriction. The research program in the author's lab was supported by grants-in-aid from the Spanish Ministry of Economy and Competitivity (MINECO) with the funding of European Regional Development Fund-European Social Fund (SAF2015-65113- C2-2-R), and Alfredo Martin-Reyes Foundation (Arehucas)-FICIC. CR and MMG were supported by postdoctoral grants-in-aid from the MINECO-Juan de la Cierva

at 120 mg/day (oral recommended phase II dose) was associated with an acceptable safety/tolerability and with clinical improvements in MPN JAK2V617F patients [103]. These findings provide rationale for further gandotinib clinical testing.

#### **5.2 Type II inhibitors**

*Tyrosine Kinases as Druggable Targets in Cancer*

NCT01702064 and NCT01751425).

evaluated in MF patients with thrombocytopenia.

*5.1.2 Momelotinib*

has been stopped.

*5.1.3 Pacritinib*

*5.1.4 NS-108*

*5.1.5 Gandotinib*

and STAT5 phosphorylation and induce apoptosis in primary CML CD34<sup>+</sup>

These effects contribute to an effective elimination of these cells *in vitro* and *in vivo* and support the current utilization of ruxolitinib/nilotinib combinations in clinical trials to eradicate persistent disease in CML patients [113]. In fact, a phase I and a phase I/II clinical studies are already underway to evaluate the potential synergic effects of ruxolitinib-tyrosine kinase inhibitors combinations, such as nilotinib/ imatinib, on eradicating CML stem/progenitor cells (ClinicalTrials.gov identifiers:

Given its potential clinical relevance, there are other type I JAK inhibitors that should be highlighted: momelotinib (CYT38) is a dual JAK1/2 inhibitor that, similar to ruxolitinib, reduces spleen size and MPN general related symptoms in intermediate or high-risk MF patients [114, 115]. Relevant, momelotinib has been shown to reduce anemia, which is a major issue in MF, so this drug might be an alternative to ruxolitinib for MPN patients with anemia. However, two phase-3 studies, SIMPLIFY-1 and SIMPLIFY-2, have reported that momelotinib does not seem to have major advantages over ruxolitinib, although it was related to less transfusion requirement [116, 117]. These findings have caused that momelotinib development

Pacritinib (SB1518) is a JAK2-selective inhibitor (it does not inhibit JAK1) that also inhibits FLT3 (FMS-like tyrosine kinase 3, a key target in the therapeutics of acute myeloid leukemia), colony-stimulating factor 1 receptor (CSF1R) and interleukin-1 receptor-associated kinase 1 (IRAK1) [118]. In phase I/II studies, pacritinib, at a recommended dose of 400 mg/day, showed a good activity in MF patients with gastrointestinal alterations being the most frequent side effect [119, 120]. After these promising results, two phase-3 clinical trials (PERSIST 1 and 2) were initiated testing different pacritinib concentrations [121]. However, in 2016, FDA carried out a full clinical hold on these trials due to a suspected excess of mortality in treated patients caused by intracranial hemorrhage and cardiac events. This clinical hold was lifted by the FDA on January 2017 [121] and subsequently CTI Biopharma announced PAC203, a new trial in which different doses of pacritinib are being

NS-108 is a potent JAK2-selective inhibitor that also inhibits Src kinases [122]. This compound showed selectivity and high potency for JAK2V617F mutant in mouse models without producing anemia or thrombocytopenia [122]. NS-108 has been tested in a phase II trial at a recommended dose of 300 mg/day in MF patients. As previously described for other JAK2 inhibitors, NS-108 significantly reduced spleen size and improved general MF-related symptoms. However, this product was not able to significantly reduce the amount of JAK2V617F mutant cells [123].

Gandotinib (LY2784544) is a selective and potent inhibitor of JAK2V617F [124]. This drug has been evaluated in a phase I trial for safety, tolerability, and pharmacokinetic parameters in patients with MF, PV, and ET. Treatment with this compound

cells.

**58**

Two type II JAK2 inhibitors (NVP-BBT594 and NVP-CHZ868) have been developed. NVP-BBT594 was effective in MPN cellular models [125] and NVP-CHZ868 in preclinical mouse MPN models. However, JAK2 inhibition caused by type II inhibitors is more effective and powerful than that produced by type I inhibitors, which in turn may induce profound cytopenia, limiting its future development and clinical use.

#### **5.3 Allosteric inhibitors**

In this group are the so-called type III (they bind to a site close to the ATPbinding site, e.g., LS104 [126]) and type IV inhibitors (they bind to an allosteric site distant from the ATP-binding site, e.g., ON044580 [127]). Since these inhibitors do not target the ATP pocket, hypothetically, they are more specific than type I/II JAK inhibitors due to the high homology shown by the ATP-binding sites. Taking this into account, JAK allosteric inhibitors would be particularly indicated to treat MPNs related to JAK mutations (especially JAK2V617F) as an efficient inhibition of WT-JAK2 will always produce a profound cytopenia. Nowadays, there is no a JAK allosteric inhibitor in clinical development.

### **6. Conclusions**

In summary, JAK kinases are key proteins in the development of hematological malignancies, since different genetic alterations including fusion protein formation, gene amplification, and point mutations have been discovered in a wide array of hematological malignancies. Particularly, JAK somatic point mutations have been detected in a high proportion of HC patients. Furthermore, detection of JAK mutations is beginning to provide prognostic information. For all these reasons, manipulating JAK activity currently constitutes an interesting therapeutic strategy and an interesting biomarker in hematological cancer. A great effort has been made by researchers in the last decade to find and characterize novel JAK inhibitors that might be clinically used, and, in fact, some of them have already reached clinical evaluation. However, more efforts are needed in order to identify more JAK mutations that lead to develop more accurate therapies against specific malignancies.

### **Acknowledgements**

We thank all the authors who contributed to the understanding of the cross-talk between the JAK kinase and hematological cancer. We apologize to those whose work deserves to be cited but unfortunately are not quoted because of space restriction. The research program in the author's lab was supported by grants-in-aid from the Spanish Ministry of Economy and Competitivity (MINECO) with the funding of European Regional Development Fund-European Social Fund (SAF2015-65113- C2-2-R), and Alfredo Martin-Reyes Foundation (Arehucas)-FICIC. CR and MMG were supported by postdoctoral grants-in-aid from the MINECO-Juan de la Cierva Program and ULPGC, respectively.

### **Conflict of interest**

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

#### **Author details**

Carlota Recio\*, Haidée Aranda-Tavío, Miguel Guerra-Rodríguez, Mercedes de Mirecki-Garrido, Patricia Martín-Rodríguez, Borja Guerra and Leandro Fernández-Pérez Institute for Research in Biomedicine and Health (IUIBS), University of Las Palmas de Gran Canaria (ULPGC), Las Palmas de Gran Canaria, Spain

\*Address all correspondence to: carlota.recio@ulpgc.es

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

**61**

2016;**32**:29-33

*JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

[1] O'Shea JJ, Schwartz DM, Villarino AV, Gadina M, McInnes IB, Laurence A. The JAK-STAT pathway: Impact on human disease and therapeutic intervention.

[11] Banerjee S, Biehl A, Gadina M, Hasni S, Schwartz DM. JAK-STAT signaling as a target for inflammatory and autoimmune diseases: Current and future prospects. Drugs.

2017;**77**(5):521-546

[12] Vainchenker W, Dusa A,

Constantinescu SN. JAKs in pathology: Role of Janus kinases in hematopoietic malignancies and immunodeficiencies. Seminars in Cell & Developmental Biology. 2008;**19**(4):385-393

[13] Schindler C, Levy DE, Decker T. JAK-STAT signaling: From interferons to cytokines. The Journal of Biological Chemistry. 2007;**282**(28):20059-20063

[14] Ehret GB, Reichenbach P, Schindler U, Horvath CM, Fritz S, Nabholz M, et al. DNA binding specificity of different STAT proteins. Comparison of in vitro specificity with natural target sites. The Journal of Biological Chemistry. 2001;**276**(9):6675-6688

[15] Stark GR, Darnell JE. The JAK-STAT pathway at twenty. Immunity.

[16] Schindler C, Plumlee C. Inteferons pen the JAK-STAT pathway. Seminars in Cell & Developmental Biology.

Pharmaceutical intervention in the JAK/ STAT signaling pathway. Oncogene.

Mechanisms of disease: Insights into the emerging role of signal transducers and activators of transcription in cancer. Nature Clinical Practice. Oncology.

[19] Braunstein J, Brutsaert S, Olson R, Schindler C. STATs dimerize in the absence of phosphorylation. The

[17] Seidel HM, Lamb P, Rosen J.

[18] Haura EB, Turkson J, Jove R.

2012;**36**(4):503-514

2008;**19**(4):311-318

2000;**19**(21):2645-2656

2005;**2**(6):315-324

Annual Review of Medicine.

[2] Levy DE, Darnell JE. Stats:

Biology. 2002;**3**(9):651-662

Biology. 2012;**4**:a011205

2009;**228**(1):273-287

2012;**30**(2):88-106

2013;**12**(8):611-629

Janus kinases in immune cell signaling. Immunological Reviews.

[6] Kiu H, Nicholson SE. Biology and significance of the JAK/STAT signalling pathways. Growth Factors.

[8] Senkevitch E, Durum S. The promise of Janus kinase inhibitors in the treatment of hematological malignancies. Cytokine. 2017;**98**:33-41

[9] Springuel L, Renauld JC, Knoops L. JAK kinase targeting in hematologic malignancies: A sinuous pathway from identification of genetic

alterations towards clinical indications. Haematologica. 2015;**100**(10):1240-1253

[10] Yamaoka K. Janus kinase inhibitors for rheumatoid arthritis. Current Opinion in Chemical Biology.

[7] Miklossy G, Hilliard TS, Turkson J. Therapeutic modulators of STAT signalling for human diseases. Nature Reviews. Drug Discovery.

Transcriptional control and biological impact. Nature Reviews. Molecular Cell

[3] Leonard WJ, O'Shea JJ. Jaks and STATs: Biological implications. Annual Review of Immunology. 1998;**16**:293-322

[4] Harrison DA. The Jak/STAT pathway. Cold Spring Harbor Perspectives in

[5] Ghoreschi K, Laurence A, O'Shea JJ.

**References**

2015;**66**:311-328

*JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

#### **References**

*Tyrosine Kinases as Druggable Targets in Cancer*

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of

**Conflict of interest**

interest.

**60**

**Author details**

Leandro Fernández-Pérez

provided the original work is properly cited.

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

Institute for Research in Biomedicine and Health (IUIBS), University of Las Palmas

Carlota Recio\*, Haidée Aranda-Tavío, Miguel Guerra-Rodríguez,

de Gran Canaria (ULPGC), Las Palmas de Gran Canaria, Spain

\*Address all correspondence to: carlota.recio@ulpgc.es

Mercedes de Mirecki-Garrido, Patricia Martín-Rodríguez, Borja Guerra and

[1] O'Shea JJ, Schwartz DM, Villarino AV, Gadina M, McInnes IB, Laurence A. The JAK-STAT pathway: Impact on human disease and therapeutic intervention. Annual Review of Medicine. 2015;**66**:311-328

[2] Levy DE, Darnell JE. Stats: Transcriptional control and biological impact. Nature Reviews. Molecular Cell Biology. 2002;**3**(9):651-662

[3] Leonard WJ, O'Shea JJ. Jaks and STATs: Biological implications. Annual Review of Immunology. 1998;**16**:293-322

[4] Harrison DA. The Jak/STAT pathway. Cold Spring Harbor Perspectives in Biology. 2012;**4**:a011205

[5] Ghoreschi K, Laurence A, O'Shea JJ. Janus kinases in immune cell signaling. Immunological Reviews. 2009;**228**(1):273-287

[6] Kiu H, Nicholson SE. Biology and significance of the JAK/STAT signalling pathways. Growth Factors. 2012;**30**(2):88-106

[7] Miklossy G, Hilliard TS, Turkson J. Therapeutic modulators of STAT signalling for human diseases. Nature Reviews. Drug Discovery. 2013;**12**(8):611-629

[8] Senkevitch E, Durum S. The promise of Janus kinase inhibitors in the treatment of hematological malignancies. Cytokine. 2017;**98**:33-41

[9] Springuel L, Renauld JC, Knoops L. JAK kinase targeting in hematologic malignancies: A sinuous pathway from identification of genetic alterations towards clinical indications. Haematologica. 2015;**100**(10):1240-1253

[10] Yamaoka K. Janus kinase inhibitors for rheumatoid arthritis. Current Opinion in Chemical Biology. 2016;**32**:29-33

[11] Banerjee S, Biehl A, Gadina M, Hasni S, Schwartz DM. JAK-STAT signaling as a target for inflammatory and autoimmune diseases: Current and future prospects. Drugs. 2017;**77**(5):521-546

[12] Vainchenker W, Dusa A, Constantinescu SN. JAKs in pathology: Role of Janus kinases in hematopoietic malignancies and immunodeficiencies. Seminars in Cell & Developmental Biology. 2008;**19**(4):385-393

[13] Schindler C, Levy DE, Decker T. JAK-STAT signaling: From interferons to cytokines. The Journal of Biological Chemistry. 2007;**282**(28):20059-20063

[14] Ehret GB, Reichenbach P, Schindler U, Horvath CM, Fritz S, Nabholz M, et al. DNA binding specificity of different STAT proteins. Comparison of in vitro specificity with natural target sites. The Journal of Biological Chemistry. 2001;**276**(9):6675-6688

[15] Stark GR, Darnell JE. The JAK-STAT pathway at twenty. Immunity. 2012;**36**(4):503-514

[16] Schindler C, Plumlee C. Inteferons pen the JAK-STAT pathway. Seminars in Cell & Developmental Biology. 2008;**19**(4):311-318

[17] Seidel HM, Lamb P, Rosen J. Pharmaceutical intervention in the JAK/ STAT signaling pathway. Oncogene. 2000;**19**(21):2645-2656

[18] Haura EB, Turkson J, Jove R. Mechanisms of disease: Insights into the emerging role of signal transducers and activators of transcription in cancer. Nature Clinical Practice. Oncology. 2005;**2**(6):315-324

[19] Braunstein J, Brutsaert S, Olson R, Schindler C. STATs dimerize in the absence of phosphorylation. The

Journal of Biological Chemistry. 2003;**278**(36):34133-34140

[20] Dawson MA, Bannister AJ, Göttgens B, Foster SD, Bartke T, Green AR, et al. JAK2 phosphorylates histone H3Y41 and excludes HP1alpha from chromatin. Nature. 2009;**461**(7265):819-822

[21] Yang S, Park K, Turkson J, Arteaga CL. Ligand-independent phosphorylation of Y869 (Y845) links mutant EGFR signaling to stat-mediated gene expression. Experimental Cell Research. 2008;**314**(2):413-419

[22] Hu X, Dutta P, Tsurumi A, Li J, Wang J, Land H, et al. Unphosphorylated STAT5A stabilizes heterochromatin and suppresses tumor growth. Proceedings of the National Academy of Sciences of the United States of America. 2013;**110**(25):10213-10218

[23] Shuai K, Liu B. Regulation of JAK-STAT signalling in the immune system. Nature Reviews. Immunology. 2003;**3**(11):900-911

[24] Espert L, Dusanter-Fourt I, Chelbi-Alix MK. [Negative regulation of the JAK/STAT: Pathway implication in tumorigenesis]. Bulletin du Cancer. 2005;**92**(10):845-857

[25] Larsen L, Röpke C. Suppressors of cytokine signalling: SOCS. APMIS. 2002;**110**(12):833-844

[26] Trengove MC, Ward AC. SOCS proteins in development and disease. American Journal of Clinical and Experimental Immunology. 2013;**2**(1):1-29

[27] Ward AC, Touw I, Yoshimura A. The Jak-Stat pathway in normal and perturbed hematopoiesis. Blood. 2000;**95**(1):19-29

[28] Bromberg J. Stat proteins and oncogenesis. The Journal of Clinical Investigation. 2002;**109**(9):1139-1142 [29] Groner B, Hennighausen L. The versatile regulation of cellular events by Jak-Stat signaling: From transcriptional control to microtubule dynamics and energy metabolism. Hormone Molecular Biology and Clinical Investigation. 2012;**10**(1):193-200

[30] Rani A, Murphy JJ. STAT5 in cancer and immunity. Journal of Interferon & Cytokine Research. 2016;**36**(4):226-237

[31] Lacronique V, Boureux A, Valle VD, Poirel H, Quang CT, Mauchauffé M, et al. A TEL-JAK2 fusion protein with constitutive kinase activity in human leukemia. Science. 1997;**278**(5341):1309-1312

[32] Hammarén HM, Virtanen AT, Raivola J, Silvennoinen O. The regulation of JAKs in cytokine signaling and its breakdown in disease. Cytokine. 2018;pii:S1043-4666 (18) 30127-3

[33] Constantinescu SN, Girardot M, Pecquet C. Mining for JAK-STAT mutations in cancer. Trends in Biochemical Sciences. 2008;**33**(3):122-131

[34] Vardiman JW, Harris NL, Brunning RD. The World Health Organization (WHO) classification of the myeloid neoplasms. Blood. 2002;**100**(7):2292-2302

[35] Baxter EJ, Scott LM, Campbell PJ, East C, Fourouclas N, Swanton S, et al. Acquired mutation of the tyrosine kinase JAK2 in human myeloproliferative disorders. Lancet. 2005;**365**(9464):1054-1061

[36] James C, Ugo V, Le Couédic JP, Staerk J, Delhommeau F, Lacout C, et al. A unique clonal JAK2 mutation leading to constitutive signalling causes polycythaemia vera. Nature. 2005;**434**(7037):1144-1148

[37] Zhao R, Xing S, Li Z, Fu X, Li Q, Krantz SB, et al. Identification of an acquired JAK2 mutation

**63**

*JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

[38] Butcher CM, Hahn U, To LB, Gecz J, Wilkins EJ, Scott HS, et al. Two novel JAK2 exon 12 mutations in JAK2V617Fnegative polycythaemia vera patients. Leukemia. 2008;**22**(4):870-873

of a homodimeric type I cytokine receptor is required for JAK2V617Fmediated transformation. Proceedings of the National Academy of Sciences of the United States of America. 2005;**102**(52):18962-18967

[47] Lacout C, Pisani DF, Tulliez M, Gachelin FM, Vainchenker W, Villeval JL. JAK2V617F expression in murine hematopoietic cells leads to MPD mimicking human PV with secondary myelofibrosis. Blood.

[48] Funakoshi-Tago M, Pelletier S, Matsuda T, Parganas E, Ihle JN. Receptor specific downregulation

autophosphorylation in the FERM domain of Jak2. The EMBO Journal.

[49] Teofili L, Martini M, Cenci T, Petrucci G, Torti L, Storti S, et al. Different STAT-3 and STAT-5

diseases and is independent of the V617F JAK-2 mutation. Blood.

Blood. 2007;**110**(7):2768-2769

2007;**21**(9):1960-1963

[51] Pardanani A, Lasho TL, Finke C, Hanson CA, Tefferi A. Prevalence and clinicopathologic correlates of JAK2 exon 12 mutations in JAK2V617Fnegative polycythemia vera. Leukemia.

[52] Williams DM, Kim AH, Rogers O, Spivak JL, Moliterno AR. Phenotypic variations and new mutations in JAK2 V617F-negative polycythemia vera, erythrocytosis, and idiopathic myelofibrosis. Experimental Hematology. 2007;**35**(11):1641-1646

2007;**110**(1):354-359

phosphorylation discriminates among Ph-negative chronic myeloproliferative

[50] Colaizzo D, Amitrano L, Tiscia GL, Grandone E, Guardascione MA, Margaglione M. A new JAK2 gene mutation in patients with polycythemia vera and splanchnic vein thrombosis.

2006;**108**(5):1652-1660

of cytokine signaling by

2006;**25**(20):4763-4772

[39] Saeidi K. Myeloproliferative neoplasms: Current molecular biology and genetics. Critical Reviews in Oncology/Hematology. 2016;**98**:375-389

[40] Kralovics R, Passamonti F, Buser AS, Teo SS, Tiedt R, Passweg JR, et al. A gain-of-function mutation of JAK2 in myeloproliferative disorders. The New England Journal of Medicine.

2005;**352**(17):1779-1790

[41] Larsen TS, Christensen JH, Hasselbalch HC, Pallisgaard N. The JAK2 V617F mutation involves B- and T-lymphocyte lineages in a subgroup of patients with Philadelphia-chromosome negative chronic myeloproliferative

disorders. British Journal of Haematology. 2007;**136**(5):745-751

2008;**59**:213-222

2013;**368**(2):161-170

2007;**26**(47):6738-6749

2007;**14**(1):43-47

[42] Morgan KJ, Gilliland DG. A role for JAK2 mutations in myeloproliferative diseases. Annual Review of Medicine.

[43] O'Shea JJ, Holland SM, Staudt LM.

[44] Van Etten RA. Aberrant cytokine signaling in leukemia. Oncogene.

[45] Levine RL, Gilliland DG. JAK-2 mutations and their relevance to myeloproliferative disease. Current Opinion in Hematology.

[46] Lu X, Levine R, Tong W, Wernig G, Pikman Y, Zarnegar S, et al. Expression

JAKs and STATs in immunity, immunodeficiency, and cancer. The New England Journal of Medicine.

in polycythemia vera. The Journal of Biological Chemistry. 2005;**280**(24):22788-22792

*JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

in polycythemia vera. The Journal of Biological Chemistry. 2005;**280**(24):22788-22792

*Tyrosine Kinases as Druggable Targets in Cancer*

[20] Dawson MA, Bannister AJ, Göttgens B, Foster SD, Bartke T, Green AR, et al. JAK2 phosphorylates histone H3Y41 and excludes HP1alpha from chromatin. Nature. 2009;**461**(7265):819-822

[29] Groner B, Hennighausen L. The versatile regulation of cellular events by Jak-Stat signaling: From transcriptional control to microtubule dynamics and energy metabolism. Hormone Molecular Biology and Clinical Investigation.

[30] Rani A, Murphy JJ. STAT5 in cancer and immunity. Journal of Interferon & Cytokine Research. 2016;**36**(4):226-237

[31] Lacronique V, Boureux A, Valle VD, Poirel H, Quang CT, Mauchauffé M, et al. A TEL-JAK2 fusion protein with constitutive kinase activity in human leukemia. Science. 1997;**278**(5341):1309-1312

[32] Hammarén HM, Virtanen AT, Raivola J, Silvennoinen O. The regulation

of JAKs in cytokine signaling and its breakdown in disease. Cytokine. 2018;pii:S1043-4666 (18) 30127-3

[33] Constantinescu SN, Girardot M, Pecquet C. Mining for JAK-STAT mutations in cancer. Trends in

[34] Vardiman JW, Harris NL, Brunning RD. The World Health Organization (WHO) classification of the myeloid neoplasms. Blood.

2002;**100**(7):2292-2302

2005;**365**(9464):1054-1061

2005;**434**(7037):1144-1148

[36] James C, Ugo V, Le Couédic JP, Staerk J, Delhommeau F, Lacout C, et al. A unique clonal JAK2 mutation leading to constitutive signalling causes polycythaemia vera. Nature.

[37] Zhao R, Xing S, Li Z, Fu X, Li Q, Krantz SB, et al. Identification of an acquired JAK2 mutation

Biochemical Sciences. 2008;**33**(3):122-131

[35] Baxter EJ, Scott LM, Campbell PJ, East C, Fourouclas N, Swanton S, et al. Acquired mutation of the tyrosine kinase JAK2 in human myeloproliferative disorders. Lancet.

2012;**10**(1):193-200

Journal of Biological Chemistry. 2003;**278**(36):34133-34140

[21] Yang S, Park K, Turkson J, Arteaga CL. Ligand-independent phosphorylation of Y869 (Y845) links mutant EGFR signaling to stat-mediated gene expression. Experimental Cell Research. 2008;**314**(2):413-419

[22] Hu X, Dutta P, Tsurumi A, Li J, Wang J, Land H, et al.

Unphosphorylated STAT5A stabilizes heterochromatin and suppresses tumor growth. Proceedings of the National Academy of Sciences of the United States of America. 2013;**110**(25):10213-10218

[23] Shuai K, Liu B. Regulation of JAK-STAT signalling in the immune system. Nature Reviews. Immunology.

[24] Espert L, Dusanter-Fourt I, Chelbi-Alix MK. [Negative regulation of the JAK/STAT: Pathway implication in tumorigenesis]. Bulletin du Cancer.

[25] Larsen L, Röpke C. Suppressors of cytokine signalling: SOCS. APMIS.

[26] Trengove MC, Ward AC. SOCS proteins in development and disease.

[27] Ward AC, Touw I, Yoshimura A. The Jak-Stat pathway in normal and perturbed hematopoiesis. Blood.

[28] Bromberg J. Stat proteins and oncogenesis. The Journal of Clinical Investigation. 2002;**109**(9):1139-1142

American Journal of Clinical and Experimental Immunology.

2003;**3**(11):900-911

2005;**92**(10):845-857

2002;**110**(12):833-844

2013;**2**(1):1-29

2000;**95**(1):19-29

**62**

[38] Butcher CM, Hahn U, To LB, Gecz J, Wilkins EJ, Scott HS, et al. Two novel JAK2 exon 12 mutations in JAK2V617Fnegative polycythaemia vera patients. Leukemia. 2008;**22**(4):870-873

[39] Saeidi K. Myeloproliferative neoplasms: Current molecular biology and genetics. Critical Reviews in Oncology/Hematology. 2016;**98**:375-389

[40] Kralovics R, Passamonti F, Buser AS, Teo SS, Tiedt R, Passweg JR, et al. A gain-of-function mutation of JAK2 in myeloproliferative disorders. The New England Journal of Medicine. 2005;**352**(17):1779-1790

[41] Larsen TS, Christensen JH, Hasselbalch HC, Pallisgaard N. The JAK2 V617F mutation involves B- and T-lymphocyte lineages in a subgroup of patients with Philadelphia-chromosome negative chronic myeloproliferative disorders. British Journal of Haematology. 2007;**136**(5):745-751

[42] Morgan KJ, Gilliland DG. A role for JAK2 mutations in myeloproliferative diseases. Annual Review of Medicine. 2008;**59**:213-222

[43] O'Shea JJ, Holland SM, Staudt LM. JAKs and STATs in immunity, immunodeficiency, and cancer. The New England Journal of Medicine. 2013;**368**(2):161-170

[44] Van Etten RA. Aberrant cytokine signaling in leukemia. Oncogene. 2007;**26**(47):6738-6749

[45] Levine RL, Gilliland DG. JAK-2 mutations and their relevance to myeloproliferative disease. Current Opinion in Hematology. 2007;**14**(1):43-47

[46] Lu X, Levine R, Tong W, Wernig G, Pikman Y, Zarnegar S, et al. Expression

of a homodimeric type I cytokine receptor is required for JAK2V617Fmediated transformation. Proceedings of the National Academy of Sciences of the United States of America. 2005;**102**(52):18962-18967

[47] Lacout C, Pisani DF, Tulliez M, Gachelin FM, Vainchenker W, Villeval JL. JAK2V617F expression in murine hematopoietic cells leads to MPD mimicking human PV with secondary myelofibrosis. Blood. 2006;**108**(5):1652-1660

[48] Funakoshi-Tago M, Pelletier S, Matsuda T, Parganas E, Ihle JN. Receptor specific downregulation of cytokine signaling by autophosphorylation in the FERM domain of Jak2. The EMBO Journal. 2006;**25**(20):4763-4772

[49] Teofili L, Martini M, Cenci T, Petrucci G, Torti L, Storti S, et al. Different STAT-3 and STAT-5 phosphorylation discriminates among Ph-negative chronic myeloproliferative diseases and is independent of the V617F JAK-2 mutation. Blood. 2007;**110**(1):354-359

[50] Colaizzo D, Amitrano L, Tiscia GL, Grandone E, Guardascione MA, Margaglione M. A new JAK2 gene mutation in patients with polycythemia vera and splanchnic vein thrombosis. Blood. 2007;**110**(7):2768-2769

[51] Pardanani A, Lasho TL, Finke C, Hanson CA, Tefferi A. Prevalence and clinicopathologic correlates of JAK2 exon 12 mutations in JAK2V617Fnegative polycythemia vera. Leukemia. 2007;**21**(9):1960-1963

[52] Williams DM, Kim AH, Rogers O, Spivak JL, Moliterno AR. Phenotypic variations and new mutations in JAK2 V617F-negative polycythemia vera, erythrocytosis, and idiopathic myelofibrosis. Experimental Hematology. 2007;**35**(11):1641-1646

[53] Scott LM, Tong W, Levine RL, Scott MA, Beer PA, Stratton MR, et al. JAK2 exon 12 mutations in polycythemia vera and idiopathic erythrocytosis. The New England Journal of Medicine. 2007;**356**(5):459-468

[54] Pietra D, Li S, Brisci A, Passamonti F, Rumi E, Theocharides A, et al. Somatic mutations of JAK2 exon 12 in patients with JAK2 (V617F)-negative myeloproliferative disorders. Blood. 2008;**111**(3):1686-1689

[55] Hahn AW, Li B, Prouet P, Giri S, Pathak R, Martin MG. Acute megakaryocytic leukemia: What have we learned. Blood Reviews. 2016;**30**(1):49-53

[56] Mercher T, Wernig G, Moore SA, Levine RL, Gu T-L, Fröhling S, et al. JAK2T875N is a novel activating mutation that results in myeloproliferative disease with features of megakaryoblastic leukemia in a murine bone marrow transplantation model. Blood. 2006;**108**(8):2770-2779

[57] Hitzler JK, Cheung J, Li Y, Scherer SW, Zipursky A. GATA1 mutations in transient leukemia and acute megakaryoblastic leukemia of Down syndrome. Blood. 2003;**101**(11):4301-4304

[58] Mercher T, Coniat MB, Monni R, Mauchauffe M, Nguyen Khac F, Gressin L, et al. Involvement of a human gene related to the Drosophila spen gene in the recurrent t(1;22) translocation of acute megakaryocytic leukemia. Proceedings of the National Academy of Sciences of the United States of America. 2001;**98**(10):5776-5779

[59] Li Z, Godinho FJ, Klusmann J-H, Garriga-Canut M, Yu C, Orkin SH. Developmental stage–selective effect of somatically mutated leukemogenic transcription factor GATA1. Nature Genetics. 2005;**37**:613

[60] Zou H, Yan D, Mohi G. Differential biological activity of disease-associated JAK2 mutants. FEBS Letters. 2011;**585**(7):1007-1013

[61] Haan S, Wuller S, Kaczor J, Rolvering C, Nocker T, Behrmann I, et al. SOCS-mediated downregulation of mutant Jak2 (V617F, T875N and K539L) counteracts cytokineindependent signaling. Oncogene. 2009;**28**(34):3069-3080

[62] Lee P, Bhansali R, Izraeli S, Hijiya N, Crispino JD. The biology, pathogenesis and clinical aspects of acute lymphoblastic leukemia in children with Down syndrome. Leukemia. 2016;**30**(9):1816-1823

[63] Bercovich D, Ganmore I, Scott LM, Wainreb G, Birger Y, Elimelech A, et al. Mutations of JAK2 in acute lymphoblastic leukaemias associated with Down's syndrome. Lancet. 2008;**372**(9648):1484-1492

[64] Malinge S, Ben-Abdelali R, Settegrana C, Radford-Weiss I, Debre M, Beldjord K, et al. Novel activating JAK2 mutation in a patient with Down syndrome and B-cell precursor acute lymphoblastic leukemia. Blood. 2007;**109**(5):2202-2204

[65] Tomoyasu C, Imamura T, Tomii T, Yano M, Asai D, Goto H, et al. Copy number abnormality of acute lymphoblastic leukemia cell lines based on their genetic subtypes. International Journal of Hematology. 2018;**108**(3):312-318

[66] Cornejo MG, Kharas MG, Werneck MB, Le Bras S, Moore SA, Ball B, et al. Constitutive JAK3 activation induces lymphoproliferative syndromes in murine bone marrow transplantation models. Blood. 2009;**113**(12):2746-2754

[67] Koo GC, Tan SY, Tang T, Poon SL, Allen GE, Tan L, et al. Janus kinase 3-activating mutations identified in

**65**

*JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

natural killer/T-cell lymphoma. Cancer

leukemia and lymphoma. Immunity.

[76] Li Q, Li B, Hu L, Ning H, Jiang M, Wang D, et al. Identification of a novel functional JAK1 S646P mutation in acute lymphoblastic leukemia. Oncotarget. 2017;**8**(21):34687-34697

[77] Xiang Z, Zhao Y, Mitaksov V, Fremont DH, Kasai Y, Molitoris A, et al. Identification of somatic JAK1 mutations in patients with acute myeloid leukemia. Blood.

[78] Lacronique V, Boureux A, Monni R, Dumon S, Mauchauffe M, Mayeux P, et al. Transforming properties of chimeric TEL-JAK proteins in Ba/F3 cells. Blood. 2000;**95**(6):2076-2083

[79] Schwaller J, Frantsve J, Aster J, Williams IR, Tomasson MH, Ross TS, et al. Transformation of hematopoietic cell lines to growth-factor independence and induction of a fatal myelo- and lymphoproliferative disease in mice by retrovirally transduced TEL/JAK2 fusion genes. The EMBO Journal.

[80] Joos S, Kupper M, Ohl S, von Bonin F, Mechtersheimer G, Bentz M, et al. Genomic imbalances including amplification of the tyrosine kinase gene JAK2 in CD30+ Hodgkin cells. Cancer Research. 2000;**60**(3):549-552

[81] Rosenwald A, Wright G, Leroy K, Yu X, Gaulard P, Gascoyne RD, et al. Molecular diagnosis of primary

mediastinal B cell lymphoma identifies

a clinically favorable subgroup of diffuse large B cell lymphoma related to Hodgkin lymphoma. The Journal of Experimental Medicine.

[82] Lenz G, Wright GW, Emre NC, Kohlhammer H, Dave SS, Davis RE, et al. Molecular subtypes of diffuse large B-cell lymphoma arise by distinct

2003;**198**(6):851-862

2008;**111**(9):4809-4812

1998;**17**(18):5321-5333

2012;**36**(4):529-541

[69] Russell SM, Tayebi N, Nakajima H, Riedy MC, Roberts JL, Aman MJ, et al. Mutation of Jak3 in a patient with SCID: Essential role of Jak3 in lymphoid development. Science.

[70] Walters DK, Mercher T, Gu TL, O' Hare T, Tyner JW, Loriaux M, et al. Activating alleles of JAK3 in acute megakaryoblastic leukemia. Cancer

[71] Malinge S, Ragu C, Della-Valle V, Pisani D, Constantinescu SN, Perez C, et al. Activating mutations in human acute megakaryoblastic leukemia. Blood. 2008;**112**(10):4220-4226

[72] Bouchekioua A, Scourzic L, de Wever O, Zhang Y, Cervera P, Aline-Fardin A, et al. JAK3 deregulation by activating mutations confers invasive growth advantage in extranodal nasaltype natural killer cell lymphoma. Leukemia. 2014;**28**(2):338-348

[73] Yamashita Y, Yuan J, Suetake I, Suzuki H, Ishikawa Y, Choi YL, et al. Array-based genomic resequencing of human leukemia. Oncogene.

[74] Degryse S, Bornschein S, de Bock CE, Leroy E, Vanden Bempt M, Demeyer S, et al. Mutant JAK3 signaling

is increased by loss of wild-type JAK3 or by acquisition of secondary JAK3 mutations in T-ALL. Blood.

[75] Chen E, Staudt LM, Green AR. Janus kinase deregulation in

2010;**29**(25):3723-3731

2018;**131**(4):421-425

Discovery. 2012;**2**(7):591-597

[68] Bellanger D, Jacquemin V, Chopin M, Pierron G, Bernard OA, Ghysdael J, et al. Recurrent JAK1 and JAK3 somatic mutations in T-cell prolymphocytic leukemia. Leukemia.

2014;**28**(2):417-419

1995;**270**(5237):797-800

Cell. 2006;**10**(1):65-75

*JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

*Tyrosine Kinases as Druggable Targets in Cancer*

[53] Scott LM, Tong W, Levine RL, Scott MA, Beer PA, Stratton MR, et al. JAK2 exon 12 mutations in polycythemia vera and idiopathic erythrocytosis. The New England Journal of Medicine.

[60] Zou H, Yan D, Mohi G. Differential biological activity of disease-associated

[62] Lee P, Bhansali R, Izraeli S, Hijiya N, Crispino JD. The biology, pathogenesis

[63] Bercovich D, Ganmore I, Scott LM, Wainreb G, Birger Y, Elimelech A, et al. Mutations of JAK2 in acute lymphoblastic leukaemias associated with Down's syndrome. Lancet. 2008;**372**(9648):1484-1492

[64] Malinge S, Ben-Abdelali R, Settegrana C, Radford-Weiss I, Debre M, Beldjord K, et al. Novel activating JAK2 mutation in a patient with Down syndrome and B-cell precursor acute lymphoblastic leukemia. Blood.

[65] Tomoyasu C, Imamura T, Tomii T,

[66] Cornejo MG, Kharas MG, Werneck MB, Le Bras S, Moore SA, Ball B, et al. Constitutive JAK3 activation induces lymphoproliferative syndromes in murine bone marrow transplantation models. Blood. 2009;**113**(12):2746-2754

[67] Koo GC, Tan SY, Tang T, Poon SL, Allen GE, Tan L, et al. Janus kinase 3-activating mutations identified in

Yano M, Asai D, Goto H, et al. Copy number abnormality of acute lymphoblastic leukemia cell lines based on their genetic subtypes. International Journal of Hematology.

2007;**109**(5):2202-2204

2018;**108**(3):312-318

JAK2 mutants. FEBS Letters. 2011;**585**(7):1007-1013

[61] Haan S, Wuller S, Kaczor J, Rolvering C, Nocker T, Behrmann I, et al. SOCS-mediated downregulation

of mutant Jak2 (V617F, T875N and K539L) counteracts cytokineindependent signaling. Oncogene.

2009;**28**(34):3069-3080

and clinical aspects of acute lymphoblastic leukemia in children with Down syndrome. Leukemia.

2016;**30**(9):1816-1823

[54] Pietra D, Li S, Brisci A, Passamonti F, Rumi E, Theocharides A, et al. Somatic mutations of JAK2 exon 12 in patients with JAK2 (V617F)-negative myeloproliferative disorders. Blood.

[55] Hahn AW, Li B, Prouet P, Giri S, Pathak R, Martin MG. Acute megakaryocytic leukemia: What have we learned. Blood Reviews.

[56] Mercher T, Wernig G, Moore SA, Levine RL, Gu T-L, Fröhling S, et al. JAK2T875N is a novel activating mutation that results in myeloproliferative disease with features of megakaryoblastic leukemia in a murine bone marrow transplantation model. Blood. 2006;**108**(8):2770-2779

[57] Hitzler JK, Cheung J, Li Y, Scherer SW, Zipursky A. GATA1 mutations in transient leukemia and acute megakaryoblastic

2003;**101**(11):4301-4304

leukemia of Down syndrome. Blood.

[58] Mercher T, Coniat MB, Monni R, Mauchauffe M, Nguyen Khac F, Gressin L, et al. Involvement of a human gene related to the Drosophila spen gene in the recurrent t(1;22) translocation of acute megakaryocytic leukemia. Proceedings of the National Academy of Sciences of the United States of America. 2001;**98**(10):5776-5779

[59] Li Z, Godinho FJ, Klusmann J-H, Garriga-Canut M, Yu C, Orkin SH. Developmental stage–selective effect of somatically mutated leukemogenic transcription factor GATA1. Nature

Genetics. 2005;**37**:613

2007;**356**(5):459-468

2008;**111**(3):1686-1689

2016;**30**(1):49-53

**64**

natural killer/T-cell lymphoma. Cancer Discovery. 2012;**2**(7):591-597

[68] Bellanger D, Jacquemin V, Chopin M, Pierron G, Bernard OA, Ghysdael J, et al. Recurrent JAK1 and JAK3 somatic mutations in T-cell prolymphocytic leukemia. Leukemia. 2014;**28**(2):417-419

[69] Russell SM, Tayebi N, Nakajima H, Riedy MC, Roberts JL, Aman MJ, et al. Mutation of Jak3 in a patient with SCID: Essential role of Jak3 in lymphoid development. Science. 1995;**270**(5237):797-800

[70] Walters DK, Mercher T, Gu TL, O' Hare T, Tyner JW, Loriaux M, et al. Activating alleles of JAK3 in acute megakaryoblastic leukemia. Cancer Cell. 2006;**10**(1):65-75

[71] Malinge S, Ragu C, Della-Valle V, Pisani D, Constantinescu SN, Perez C, et al. Activating mutations in human acute megakaryoblastic leukemia. Blood. 2008;**112**(10):4220-4226

[72] Bouchekioua A, Scourzic L, de Wever O, Zhang Y, Cervera P, Aline-Fardin A, et al. JAK3 deregulation by activating mutations confers invasive growth advantage in extranodal nasaltype natural killer cell lymphoma. Leukemia. 2014;**28**(2):338-348

[73] Yamashita Y, Yuan J, Suetake I, Suzuki H, Ishikawa Y, Choi YL, et al. Array-based genomic resequencing of human leukemia. Oncogene. 2010;**29**(25):3723-3731

[74] Degryse S, Bornschein S, de Bock CE, Leroy E, Vanden Bempt M, Demeyer S, et al. Mutant JAK3 signaling is increased by loss of wild-type JAK3 or by acquisition of secondary JAK3 mutations in T-ALL. Blood. 2018;**131**(4):421-425

[75] Chen E, Staudt LM, Green AR. Janus kinase deregulation in

leukemia and lymphoma. Immunity. 2012;**36**(4):529-541

[76] Li Q, Li B, Hu L, Ning H, Jiang M, Wang D, et al. Identification of a novel functional JAK1 S646P mutation in acute lymphoblastic leukemia. Oncotarget. 2017;**8**(21):34687-34697

[77] Xiang Z, Zhao Y, Mitaksov V, Fremont DH, Kasai Y, Molitoris A, et al. Identification of somatic JAK1 mutations in patients with acute myeloid leukemia. Blood. 2008;**111**(9):4809-4812

[78] Lacronique V, Boureux A, Monni R, Dumon S, Mauchauffe M, Mayeux P, et al. Transforming properties of chimeric TEL-JAK proteins in Ba/F3 cells. Blood. 2000;**95**(6):2076-2083

[79] Schwaller J, Frantsve J, Aster J, Williams IR, Tomasson MH, Ross TS, et al. Transformation of hematopoietic cell lines to growth-factor independence and induction of a fatal myelo- and lymphoproliferative disease in mice by retrovirally transduced TEL/JAK2 fusion genes. The EMBO Journal. 1998;**17**(18):5321-5333

[80] Joos S, Kupper M, Ohl S, von Bonin F, Mechtersheimer G, Bentz M, et al. Genomic imbalances including amplification of the tyrosine kinase gene JAK2 in CD30+ Hodgkin cells. Cancer Research. 2000;**60**(3):549-552

[81] Rosenwald A, Wright G, Leroy K, Yu X, Gaulard P, Gascoyne RD, et al. Molecular diagnosis of primary mediastinal B cell lymphoma identifies a clinically favorable subgroup of diffuse large B cell lymphoma related to Hodgkin lymphoma. The Journal of Experimental Medicine. 2003;**198**(6):851-862

[82] Lenz G, Wright GW, Emre NC, Kohlhammer H, Dave SS, Davis RE, et al. Molecular subtypes of diffuse large B-cell lymphoma arise by distinct genetic pathways. Proceedings of the National Academy of Sciences of the United States of America. 2008;**105**(36):13520-13525

[83] Peeters P, Raynaud SD, Cools J, Wlodarska I, Grosgeorge J, Philip P, et al. Fusion of TEL, the ETS-variant gene 6 (ETV6), to the receptorassociated kinase JAK2 as a result of t(9;12) in a lymphoid and t(9;15;12) in a myeloid leukemia. Blood. 1997;**90**(7):2535-2540

[84] Reiter A, Walz C, Watmore A, Schoch C, Blau I, Schlegelberger B, et al. The t(8;9)(p22;p24) is a recurrent abnormality in chronic and acute leukemia that fuses PCM1 to JAK2. Cancer Research. 2005;**65**(7):2662-2667

[85] Griesinger F, Hennig H, Hillmer F, Podleschny M, Steffens R, Pies A, et al. A BCR-JAK2 fusion gene as the result of a t(9;22)(p24;q11.2) translocation in a patient with a clinically typical chronic myeloid leukemia. Genes, Chromosomes & Cancer. 2005;**44**(3):329-333

[86] Mark HF, Sotomayor EA, Nelson M, Chaves F, Sanger WG, Kaleem Z, et al. Chronic idiopathic myelofibrosis (CIMF) resulting from a unique 3;9 translocation disrupting the janus kinase 2 (JAK2) gene. Experimental and Molecular Pathology. 2006;**81**(3):217-223

[87] Poitras JL, Dal Cin P, Aster JC, Deangelo DJ, Morton CC. Novel SSBP2- JAK2 fusion gene resulting from a t(5;9) (q14.1;p24.1) in pre-B acute lymphocytic leukemia. Genes, Chromosomes & Cancer. 2008;**47**(10):884-889

[88] Nebral K, Denk D, Attarbaschi A, Konig M, Mann G, Haas OA, et al. Incidence and diversity of PAX5 fusion genes in childhood acute lymphoblastic leukemia. Leukemia. 2009;**23**(1):134-143

[89] Ho JM, Beattie BK, Squire JA, Frank DA, Barber DL. Fusion of the ets transcription factor TEL to Jak2 results in constitutive Jak-Stat signaling. Blood. 1999;**93**(12):4354-4364

[90] Carron C, Cormier F, Janin A, Lacronique V, Giovannini M, Daniel MT, et al. TEL-JAK2 transgenic mice develop T-cell leukemia. Blood. 2000;**95**(12):3891-3899

[91] Ho JM, Nguyen MH, Dierov JK, Badger KM, Beattie BK, Tartaro P, et al. TEL-JAK2 constitutively activates the extracellular signal-regulated kinase (ERK), stress-activated protein/Jun kinase (SAPK/JNK), and p38 signaling pathways. Blood. 2002;**100**(4):1438-1448

[92] Nguyen MH, Ho JM, Beattie BK, Barber DL. TEL-JAK2 mediates constitutive activation of the phosphatidylinositol 3′-kinase/ protein kinase B signaling pathway. The Journal of Biological Chemistry. 2001;**276**(35):32704-32713

[93] Santos SC, Lacronique V, Bouchaert I, Monni R, Bernard O, Gisselbrecht S, et al. Constitutively active STAT5 variants induce growth and survival of hematopoietic cells through a PI 3-kinase/Akt dependent pathway. Oncogene. 2001;**20**(17):2080-2090

[94] Malinge S, Monni R, Bernard O, Penard-Lacronique V. Activation of the NF-kappaB pathway by the leukemogenic TEL-Jak2 and TEL-Abl fusion proteins leads to the accumulation of antiapoptotic IAP proteins and involves IKKalpha. Oncogene. 2006;**25**(25):3589-3597

[95] Arber DA, Orazi A, Hasserjian R, Thiele J, Borowitz MJ, Le Beau MM, et al. The 2016 revision to the World Health Organization classification of myeloid neoplasms and acute leukemia. Blood. 2016;**127**(20):2391-2405

**67**

*JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

[96] Levine RL, Wadleigh M, Cools J, Ebert BL, Wernig G, Huntly BJ, et al. Activating mutation in the tyrosine kinase JAK2 in polycythemia vera, essential thrombocythemia, and myeloid metaplasia with myelofibrosis. (JAK2)(V617F) inhibitor, gandotinib (LY2784544), in patients with primary myelofibrosis, polycythemia vera, and essential thrombocythemia. Leukemia

Griesshammer M, Mesa RA, Brachmann CB, Kawashima J, et al. A phase 2 study of momelotinib, a potent JAK1 and JAK2 inhibitor, in patients with polycythemia vera or essential thrombocythemia. Leukemia Research. 2017;**60**:11-17

[105] Schwartz DM, Bonelli M, Gadina M, O'Shea JJ. Type I/II cytokines, JAKs, and new strategies for treating autoimmune diseases. Nature Reviews Rheumatology. 2016;**12**(1):25-36

[106] Tam CS, Kantarjian H, Cortes J, Lynn A, Pierce S, Zhou L, et al. Dynamic model for predicting death within 12 months in patients with primary or post-polycythemia vera/essential thrombocythemia myelofibrosis. Journal of Clinical Oncology. 2009;**27**(33):5587-5593

[107] Kleppe M, Spitzer MH, Li S, Hill CE, Dong L, Papalexi E, et al. Jak1 integrates cytokine sensing to regulate hematopoietic stem cell function and stress hematopoiesis. Cell Stem Cell.

[108] Harrison C, Kiladjian JJ, Al-Ali HK, Gisslinger H, Waltzman R, Stalbovskaya V, et al. JAK inhibition with ruxolitinib versus best available therapy for

myelofibrosis. The New England Journal of Medicine. 2012;**366**(9):787-798

[109] Verstovsek S, Mesa RA, Gotlib J, Levy RS, Gupta V, DiPersio JF, et al. A double-blind, placebo-controlled trial of ruxolitinib for myelofibrosis. The New England Journal of Medicine.

[110] Vannucchi AM, Kantarjian HM, Kiladjian JJ, Gotlib J, Cervantes F, Mesa RA, et al. A pooled analysis of

2017;**21**(4):489-501 e7

2012;**366**(9):799-807

Research. 2017;**61**:89-95

[104] Verstovsek S, Courby S,

Cancer Cell. 2005;**7**(4):387-397

neoplasm pathogenesis. Blood.

disorders. F1000Res. 2018;**7**:82

2014;**123**(22):e123-e133

2017;**18**(18):1929-1938

2015;**372**(5):426-435

2015;**125**(21):3352-3353

[101] Vannucchi AM, Kiladjian JJ, Griesshammer M, Masszi T, Durrant S,

Passamonti F, et al. Ruxolitinib versus standard therapy for the treatment of polycythemia vera. The New England Journal of Medicine.

[102] Pieri L, Pancrazzi A, Pacilli A, Rabuzzi C, Rotunno G, Fanelli T, et al. JAK2V617F complete molecular remission in polycythemia vera/ essential thrombocythemia patients treated with ruxolitinib. Blood.

[103] Verstovsek S, Mesa RA, Salama ME, Li L, Pitou C, Nunes FP, et al. A phase 1 study of the Janus kinase 2

[97] Rampal R, Al-Shahrour F, Abdel-Wahab O, Patel JP, Brunel JP, Mermel CH, et al. Integrated genomic analysis illustrates the central role of JAK-STAT pathway activation in myeloproliferative

[98] Vainchenker W, Leroy E, Gilles L, Marty C, Plo I, Constantinescu

SN. JAK inhibitors for the treatment of myeloproliferative neoplasms and other

[99] Bose P, Verstovsek S. Developmental therapeutics in myeloproliferative neoplasms. Clinical Lymphoma,

Myeloma & Leukemia. 2017;**17S**:S43-S52

[100] Griesshammer M, Sadjadian P. The BCR-ABL1-negative myeloproliferative neoplasms: A review of JAK inhibitors in the therapeutic armamentarium. Expert Opinion on Pharmacotherapy.

#### *JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

*Tyrosine Kinases as Druggable Targets in Cancer*

[89] Ho JM, Beattie BK, Squire JA, Frank DA, Barber DL. Fusion of the ets transcription factor TEL to Jak2 results in constitutive Jak-Stat signaling. Blood.

[90] Carron C, Cormier F, Janin A, Lacronique V, Giovannini M, Daniel MT, et al. TEL-JAK2 transgenic mice develop T-cell leukemia. Blood.

[91] Ho JM, Nguyen MH, Dierov JK, Badger KM, Beattie BK, Tartaro P, et al. TEL-JAK2 constitutively activates the extracellular signal-regulated kinase (ERK), stress-activated protein/Jun kinase (SAPK/JNK), and p38 signaling pathways. Blood.

[92] Nguyen MH, Ho JM, Beattie BK, Barber DL. TEL-JAK2 mediates constitutive activation of the phosphatidylinositol 3′-kinase/ protein kinase B signaling pathway. The Journal of Biological Chemistry.

[93] Santos SC, Lacronique V, Bouchaert I, Monni R, Bernard O, Gisselbrecht S, et al. Constitutively active STAT5 variants induce growth and survival of hematopoietic cells through a PI 3-kinase/Akt dependent pathway. Oncogene. 2001;**20**(17):2080-2090

[94] Malinge S, Monni R, Bernard O, Penard-Lacronique V. Activation of the NF-kappaB pathway by the leukemogenic TEL-Jak2 and TEL-Abl fusion proteins leads to the accumulation of antiapoptotic IAP proteins and involves IKKalpha. Oncogene. 2006;**25**(25):3589-3597

[95] Arber DA, Orazi A, Hasserjian R, Thiele J, Borowitz MJ, Le Beau MM, et al. The 2016 revision to the World Health Organization classification of myeloid neoplasms and acute leukemia.

Blood. 2016;**127**(20):2391-2405

1999;**93**(12):4354-4364

2000;**95**(12):3891-3899

2002;**100**(4):1438-1448

2001;**276**(35):32704-32713

genetic pathways. Proceedings of the National Academy of Sciences of the United States of America. 2008;**105**(36):13520-13525

[83] Peeters P, Raynaud SD, Cools J, Wlodarska I, Grosgeorge J, Philip P, et al. Fusion of TEL, the ETS-variant gene 6 (ETV6), to the receptorassociated kinase JAK2 as a result of t(9;12) in a lymphoid and t(9;15;12) in a myeloid leukemia. Blood.

[84] Reiter A, Walz C, Watmore A, Schoch C, Blau I, Schlegelberger B, et al. The t(8;9)(p22;p24) is a recurrent abnormality in chronic and acute leukemia that fuses PCM1 to JAK2. Cancer Research. 2005;**65**(7):2662-2667

[85] Griesinger F, Hennig H, Hillmer F, Podleschny M, Steffens R, Pies A, et al. A BCR-JAK2 fusion gene as the result of a t(9;22)(p24;q11.2) translocation

in a patient with a clinically typical chronic myeloid leukemia. Genes, Chromosomes & Cancer.

[86] Mark HF, Sotomayor EA, Nelson M, Chaves F, Sanger WG, Kaleem Z, et al. Chronic idiopathic myelofibrosis (CIMF) resulting from a unique 3;9 translocation disrupting the janus kinase 2 (JAK2) gene.

Experimental and Molecular Pathology.

[87] Poitras JL, Dal Cin P, Aster JC, Deangelo DJ, Morton CC. Novel SSBP2- JAK2 fusion gene resulting from a t(5;9) (q14.1;p24.1) in pre-B acute lymphocytic leukemia. Genes, Chromosomes & Cancer. 2008;**47**(10):884-889

[88] Nebral K, Denk D, Attarbaschi A, Konig M, Mann G, Haas OA, et al. Incidence and diversity of PAX5 fusion genes in childhood acute lymphoblastic leukemia. Leukemia.

2005;**44**(3):329-333

2006;**81**(3):217-223

2009;**23**(1):134-143

1997;**90**(7):2535-2540

**66**

[96] Levine RL, Wadleigh M, Cools J, Ebert BL, Wernig G, Huntly BJ, et al. Activating mutation in the tyrosine kinase JAK2 in polycythemia vera, essential thrombocythemia, and myeloid metaplasia with myelofibrosis. Cancer Cell. 2005;**7**(4):387-397

[97] Rampal R, Al-Shahrour F, Abdel-Wahab O, Patel JP, Brunel JP, Mermel CH, et al. Integrated genomic analysis illustrates the central role of JAK-STAT pathway activation in myeloproliferative neoplasm pathogenesis. Blood. 2014;**123**(22):e123-e133

[98] Vainchenker W, Leroy E, Gilles L, Marty C, Plo I, Constantinescu SN. JAK inhibitors for the treatment of myeloproliferative neoplasms and other disorders. F1000Res. 2018;**7**:82

[99] Bose P, Verstovsek S. Developmental therapeutics in myeloproliferative neoplasms. Clinical Lymphoma, Myeloma & Leukemia. 2017;**17S**:S43-S52

[100] Griesshammer M, Sadjadian P. The BCR-ABL1-negative myeloproliferative neoplasms: A review of JAK inhibitors in the therapeutic armamentarium. Expert Opinion on Pharmacotherapy. 2017;**18**(18):1929-1938

[101] Vannucchi AM, Kiladjian JJ, Griesshammer M, Masszi T, Durrant S, Passamonti F, et al. Ruxolitinib versus standard therapy for the treatment of polycythemia vera. The New England Journal of Medicine. 2015;**372**(5):426-435

[102] Pieri L, Pancrazzi A, Pacilli A, Rabuzzi C, Rotunno G, Fanelli T, et al. JAK2V617F complete molecular remission in polycythemia vera/ essential thrombocythemia patients treated with ruxolitinib. Blood. 2015;**125**(21):3352-3353

[103] Verstovsek S, Mesa RA, Salama ME, Li L, Pitou C, Nunes FP, et al. A phase 1 study of the Janus kinase 2

(JAK2)(V617F) inhibitor, gandotinib (LY2784544), in patients with primary myelofibrosis, polycythemia vera, and essential thrombocythemia. Leukemia Research. 2017;**61**:89-95

[104] Verstovsek S, Courby S, Griesshammer M, Mesa RA, Brachmann CB, Kawashima J, et al. A phase 2 study of momelotinib, a potent JAK1 and JAK2 inhibitor, in patients with polycythemia vera or essential thrombocythemia. Leukemia Research. 2017;**60**:11-17

[105] Schwartz DM, Bonelli M, Gadina M, O'Shea JJ. Type I/II cytokines, JAKs, and new strategies for treating autoimmune diseases. Nature Reviews Rheumatology. 2016;**12**(1):25-36

[106] Tam CS, Kantarjian H, Cortes J, Lynn A, Pierce S, Zhou L, et al. Dynamic model for predicting death within 12 months in patients with primary or post-polycythemia vera/essential thrombocythemia myelofibrosis. Journal of Clinical Oncology. 2009;**27**(33):5587-5593

[107] Kleppe M, Spitzer MH, Li S, Hill CE, Dong L, Papalexi E, et al. Jak1 integrates cytokine sensing to regulate hematopoietic stem cell function and stress hematopoiesis. Cell Stem Cell. 2017;**21**(4):489-501 e7

[108] Harrison C, Kiladjian JJ, Al-Ali HK, Gisslinger H, Waltzman R, Stalbovskaya V, et al. JAK inhibition with ruxolitinib versus best available therapy for myelofibrosis. The New England Journal of Medicine. 2012;**366**(9):787-798

[109] Verstovsek S, Mesa RA, Gotlib J, Levy RS, Gupta V, DiPersio JF, et al. A double-blind, placebo-controlled trial of ruxolitinib for myelofibrosis. The New England Journal of Medicine. 2012;**366**(9):799-807

[110] Vannucchi AM, Kantarjian HM, Kiladjian JJ, Gotlib J, Cervantes F, Mesa RA, et al. A pooled analysis of

overall survival in COMFORT-I and COMFORT-II, 2 randomized phase III trials of ruxolitinib for the treatment of myelofibrosis. Haematologica. 2015;**100**(9):1139-1145

[111] Passamonti F, Griesshammer M, Palandri F, Egyed M, Benevolo G, Devos T, et al. Ruxolitinib for the treatment of inadequately controlled polycythaemia vera without splenomegaly (RESPONSE-2): A randomised, openlabel, phase 3b study. The Lancet Oncology. 2017;**18**(1):88-99

[112] Civallero M, Cosenza M, Pozzi S, Sacchi S. Ruxolitinib combined with vorinostat suppresses tumor growth and alters metabolic phenotype in hematological diseases. Oncotarget. 2017;**8**(61):103797-103814

[113] Gallipoli P, Cook A, Rhodes S, Hopcroft L, Wheadon H, Whetton AD, et al. JAK2/STAT5 inhibition by nilotinib with ruxolitinib contributes to the elimination of CML CD34+ cells in vitro and in vivo. Blood. 2014;**124**(9):1492-1501

[114] Tefferi A, Barraco D, Lasho TL, Shah S, Begna KH, Al-Kali A, et al. Momelotinib therapy for myelofibrosis: A 7-year follow-up. Blood Cancer Journal. 2018;**8**(3):29

[115] Pardanani A, Gotlib J, Roberts AW, Wadleigh M, Sirhan S, Kawashima J, et al. Long-term efficacy and safety of momelotinib, a JAK1 and JAK2 inhibitor, for the treatment of myelofibrosis. Leukemia. 2018;**32**(4):1035-1038

[116] Mesa RA, Kiladjian JJ, Catalano JV, Devos T, Egyed M, Hellmann A, et al. SIMPLIFY-1: A phase III randomized trial of momelotinib versus ruxolitinib in janus kinase inhibitor-naive patients with myelofibrosis. Journal of Clinical Oncology. 2017;**35**(34):3844-3850

[117] Harrison CN, Vannucchi AM, Platzbecker U, Cervantes F, Gupta V, Lavie D, et al. Momelotinib versus best available therapy in patients with myelofibrosis previously treated with ruxolitinib (SIMPLIFY 2): A randomised, open-label, phase 3 trial. Lancet Haematology. 2018;**5**(2):e73-e81

[118] Poulsen A, William A, Blanchard S, Lee A, Nagaraj H, Wang H, et al. Structure-based design of oxygenlinked macrocyclic kinase inhibitors: Discovery of SB1518 and SB1578, potent inhibitors of Janus kinase 2 (JAK2) and Fms-like tyrosine kinase-3 (FLT3). Journal of Computer-Aided Molecular Design. 2012;**26**(4):437-450

[119] Verstovsek S, Odenike O, Singer JW, Granston T, Al-Fayoumi S, Deeg HJ. Phase 1/2 study of pacritinib, a next generation JAK2/FLT3 inhibitor, in myelofibrosis or other myeloid malignancies. Journal of Hematology & Oncology. 2016;**9**(1):137

[120] Komrokji RS, Seymour JF, Roberts AW, Wadleigh M, To LB, Scherber R, et al. Results of a phase 2 study of pacritinib (SB1518), a JAK2/JAK2(V617F) inhibitor, in patients with myelofibrosis. Blood. 2015;**125**(17):2649-2655

[121] Mesa RA, Vannucchi AM, Mead A, Egyed M, Szoke A, Suvorov A, et al. Pacritinib versus best available therapy for the treatment of myelofibrosis irrespective of baseline cytopenias (PERSIST-1): An international, randomised, phase 3 trial. Lancet Haematology. 2017;**4**(5):e225-ee36

[122] Nakaya Y, Shide K, Naito H, Niwa T, Horio T, Miyake J, et al. Effect of NS-018, a selective JAK2V617F inhibitor, in a murine model of myelofibrosis. Blood Cancer Journal. 2014;**4**:e174

[123] Verstovsek S, Talpaz M, Ritchie E, Wadleigh M, Odenike O, Jamieson C, et al. A phase I, open-label, doseescalation, multicenter study of the JAK2 inhibitor NS-018 in patients

**69**

*JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

[124] Ma L, Clayton JR, Walgren RA, Zhao B, Evans RJ, Smith MC, et al. Discovery and characterization of LY2784544, a small-molecule tyrosine kinase inhibitor of JAK2V617F. Blood

[125] Andraos R, Qian Z, Bonenfant D, Rubert J, Vangrevelinghe E, Scheufler C, et al. Modulation of activation-loop phosphorylation by JAK inhibitors is binding mode dependent. Cancer Discovery. 2012;**2**(6):512-523

[127] Jatiani SS, Cosenza SC, Reddy MV, Ha JH, Baker SJ, Samanta AK, et al. A non-ATP-competitive dual inhibitor of JAK2 and BCR-ABL kinases: Elucidation of a novel therapeutic spectrum based on substrate competitive inhibition. Genes & Cancer. 2010;**1**(4):331-345

with myelofibrosis. Leukemia.

Cancer Journal. 2013;**3**:e109

[126] Lipka DB, Hoffmann LS, Heidel F, Markova B, Blum MC, Breitenbuecher F, et al. LS104, a non-ATP-competitive small-molecule inhibitor of JAK2, is potently inducing apoptosis in JAK2V617F-positive cells. Molecular Cancer Therapeutics.

2008;**7**(5):1176-1184

2017;**31**(2):393-402

*JAK, an Oncokinase in Hematological Cancer DOI: http://dx.doi.org/10.5772/intechopen.84177*

with myelofibrosis. Leukemia. 2017;**31**(2):393-402

*Tyrosine Kinases as Druggable Targets in Cancer*

Lavie D, et al. Momelotinib versus best available therapy in patients with myelofibrosis previously treated with ruxolitinib (SIMPLIFY 2): A randomised, open-label, phase 3 trial. Lancet Haematology. 2018;**5**(2):e73-e81

[118] Poulsen A, William A, Blanchard S, Lee A, Nagaraj H, Wang H, et al. Structure-based design of oxygenlinked macrocyclic kinase inhibitors: Discovery of SB1518 and SB1578, potent inhibitors of Janus kinase 2 (JAK2) and Fms-like tyrosine kinase-3 (FLT3). Journal of Computer-Aided Molecular

Design. 2012;**26**(4):437-450

Oncology. 2016;**9**(1):137

2015;**125**(17):2649-2655

[120] Komrokji RS, Seymour JF, Roberts AW, Wadleigh M, To LB, Scherber R, et al. Results of a phase 2 study of pacritinib (SB1518), a JAK2/JAK2(V617F) inhibitor, in patients with myelofibrosis. Blood.

[121] Mesa RA, Vannucchi AM, Mead A, Egyed M, Szoke A, Suvorov A, et al. Pacritinib versus best available therapy for the treatment of myelofibrosis irrespective of baseline cytopenias (PERSIST-1): An international, randomised, phase 3 trial. Lancet Haematology. 2017;**4**(5):e225-ee36

[122] Nakaya Y, Shide K, Naito H, Niwa T, Horio T, Miyake J, et al. Effect of NS-018, a selective JAK2V617F inhibitor, in a murine model of myelofibrosis. Blood Cancer Journal. 2014;**4**:e174

[123] Verstovsek S, Talpaz M, Ritchie E, Wadleigh M, Odenike O, Jamieson C, et al. A phase I, open-label, doseescalation, multicenter study of the JAK2 inhibitor NS-018 in patients

[119] Verstovsek S, Odenike O, Singer JW, Granston T, Al-Fayoumi S, Deeg HJ. Phase 1/2 study of pacritinib, a next generation JAK2/FLT3 inhibitor, in myelofibrosis or other myeloid malignancies. Journal of Hematology &

overall survival in COMFORT-I and COMFORT-II, 2 randomized phase III trials of ruxolitinib for the treatment of myelofibrosis. Haematologica.

[111] Passamonti F, Griesshammer M, Palandri F, Egyed M, Benevolo G, Devos T, et al. Ruxolitinib for the treatment of inadequately controlled polycythaemia

(RESPONSE-2): A randomised, openlabel, phase 3b study. The Lancet Oncology. 2017;**18**(1):88-99

[112] Civallero M, Cosenza M, Pozzi S, Sacchi S. Ruxolitinib combined with vorinostat suppresses tumor growth and alters metabolic phenotype in hematological diseases. Oncotarget.

[113] Gallipoli P, Cook A, Rhodes S, Hopcroft L, Wheadon H, Whetton AD,

[114] Tefferi A, Barraco D, Lasho TL, Shah S, Begna KH, Al-Kali A, et al. Momelotinib therapy for myelofibrosis: A 7-year follow-up. Blood Cancer

[115] Pardanani A, Gotlib J, Roberts AW, Wadleigh M, Sirhan S, Kawashima J, et al. Long-term efficacy and safety of momelotinib, a JAK1 and JAK2 inhibitor, for the treatment of myelofibrosis. Leukemia. 2018;**32**(4):1035-1038

[116] Mesa RA, Kiladjian JJ, Catalano JV, Devos T, Egyed M, Hellmann A, et al. SIMPLIFY-1: A phase III randomized trial of momelotinib versus ruxolitinib in janus kinase inhibitor-naive patients with myelofibrosis. Journal of Clinical Oncology. 2017;**35**(34):3844-3850

[117] Harrison CN, Vannucchi AM, Platzbecker U, Cervantes F, Gupta V,

et al. JAK2/STAT5 inhibition by nilotinib with ruxolitinib contributes to the elimination of CML CD34+ cells in vitro and in vivo. Blood.

2015;**100**(9):1139-1145

vera without splenomegaly

2017;**8**(61):103797-103814

2014;**124**(9):1492-1501

Journal. 2018;**8**(3):29

**68**

[124] Ma L, Clayton JR, Walgren RA, Zhao B, Evans RJ, Smith MC, et al. Discovery and characterization of LY2784544, a small-molecule tyrosine kinase inhibitor of JAK2V617F. Blood Cancer Journal. 2013;**3**:e109

[125] Andraos R, Qian Z, Bonenfant D, Rubert J, Vangrevelinghe E, Scheufler C, et al. Modulation of activation-loop phosphorylation by JAK inhibitors is binding mode dependent. Cancer Discovery. 2012;**2**(6):512-523

[126] Lipka DB, Hoffmann LS, Heidel F, Markova B, Blum MC, Breitenbuecher F, et al. LS104, a non-ATP-competitive small-molecule inhibitor of JAK2, is potently inducing apoptosis in JAK2V617F-positive cells. Molecular Cancer Therapeutics. 2008;**7**(5):1176-1184

[127] Jatiani SS, Cosenza SC, Reddy MV, Ha JH, Baker SJ, Samanta AK, et al. A non-ATP-competitive dual inhibitor of JAK2 and BCR-ABL kinases: Elucidation of a novel therapeutic spectrum based on substrate competitive inhibition. Genes & Cancer. 2010;**1**(4):331-345

**71**

Section 3

Cancer Treatment by

Tyrosine Kinase Inhibitors

### Section 3
