**4. miRNA expression and its role in the differential diagnosis of acute lymphoblastic leukemia subtypes**

Acute lymphoblastic leukemia (ALL) is characterized by clonal proliferation of early B- and T-lymphocyte progenitors that result in the accumulation of lymphoblasts in the bone marrow and various extramedullary sites. ALL is also the hematology neoplasia most commonly observed in the pediatric population, while it is relatively **less common than AML** in adults [36]. Around 75% of childhood ALL cases contain at least one alteration chromosomic, have lymphoid maturation arrest in distinct stages, and involve B- or T-lineages to leaving different immunophenotypes with different miRNA signatures [37].

microRNAs participate in different physiological and cellular processes, such as development and tissue differentiation, cell identity, cell cycle progression, and programmed cell death [38]. Nowadays, it is known that the distinct stages of lymphopoiesis and the direction of lymphoid precursor maturation are influenced by miRNA expression differentially. However, an aberrant expression of miRNAs is related with malignant lymphopoiesis, characteristic that can be utilized as signature to diagnosis and classification diagnosis of acute lymphoblastic leukemia [18]. Interestingly, miRNA groups that can clearly differentiate ALL of its normal counterpart, B-ALL versus T-ALL and ALL subtypes with specific genetic abnormalities have been reported. De Oliveira and collaborators reported miRNA-128a and miRNA-181b overexpressed and miRNA-100, miRNA-196b, and let-7e with lower level when compared the miRNAs expression in normal pediatric bone morrow (BM) samples and BM samples of pediatric ALL. The authors point out miR-196b as a miRNA highly expressed in T-ALL, while miR-100 was related with the presence of t(12;21) [39].

A study in Brazilian children with T- or B-cell acute lymphoblastic leukemia (T-ALL or B-ALL) evaluated a bone marrow miRNAs profile that may be used for distinguishing childhood lymphoblastic leukemia subtypes [40]. The authors mention that miR-708-5p, miR-497-5p, miR-151a-5p, miR-151b, miR-371b-5p, miR-455-5p, miR-195-5p, miR-1266-5p, miR-574-5p, miR-425-5p downregulated and miR-450b-5p, miR-450a-5p, miR-542-5p, miR-424-5p, miR-629-5p, miR-29c-5p upregulated in childhood T-ALL may be used for distinguishing childhood T- and B-ALL subtypes. However, a machine learning analysis showed that miR-29c-5p, which is involved in calcium signaling, is critical for B-cell lymphocyte fate. So, it is the best discriminator between childhood T- and B-ALL [40].

In a series of adult ALL cases, the expression profile of 470 miRNAs was measured by microarray analysis; 3 miRNAs (miR-148, miR-151, and miR-424) were identified as discriminative of T-lineage versus B-lineage ALL; and miR-151 dramatically downmodulated an miR-148a and miR-424 with higher expression in patients with T-ALL [41]. Furthermore, in the B-lineage ALL cases with special molecular lesions, those with BCR/ABL, E2A/PBX1, MLL/AF4 rearrangements and cases lacking known genetic abnormalities can be differentiated by a set of six miRNA, which was highlighted by one-way analysis of variance [41]. These miRNAs were preferentially expressed in each chromosomic rearrangement; miR-425-5p, miR-191, and miR-128 were expressed in the E2A/PBX1-positive case, miR-629 was highly expressed in cases harboring MLL/AF4 rearrangement, while high levels of miR-146b and miR-126 were observed in the BCR/ABL-positive cases [41]. Other study in pediatric ALL showed

**61**

*miRNAs in Acute Lymphoblastic Leukemia: Diagnosis, Prognosis and Target Therapeutic*

450a-5p, miR-542-5p, miR-424-5p, miR-629-5p,

miR-29c-5p

miRNA-708

miR-424

miR-128

miR-629

miR-146b, miR-126

miR-425-5p, miR-191,

**ALL subtype Upregulated expression Downregulated expression References**

MLL-rearranged, T-ALL miR-196b [41]

miR-708-5p, miR-497-5p, miR-151a-5p, miR-151b, miR-371b-5p, miR-455-5p, miR-195-5p, miR-1266-5p, miR-574-5p, miR-425-5p

miR-151 [40]

[39]

in seven major subtypes of pediatric ALL, which included: T-cell, MLL-rearranged, TEL-AML1-positive, E2A-PBX1-positive, hyperdiploid ALL, BCR-ABL-positive, and B-other ALLs, the differential miRNA expression. miRNA-708 was highly expressed in TEL-AML1, BCR-ABL, E2A-PBX1, hyperdiploid, and B-other cases than in the MLL-rearranged and T-ALL cases. On the other hand, the expression of miR-196b was higher in MLL-rearranged and T-ALL cases as compared with the expression level in the precursor B-ALL cases [42]. This information suggests that upregulated expression of miR-424 and downregulated expression of miR-151 might be good diagnostic

*Expression of miRNAs in children and adults to differentiate acute lymphoblastic leukemia subtypes.*

Malik and collaborators propose a novel miR-2909-KLF4 molecular axis to differentiate the pathogeneses of pediatric B- and T-cell ALLs that may represent a new diagnostic marker, through alterations in miRNA expression patterns and their respective targets. The authors demonstrate the ability of miR-2909 to repress KLF4 expression in pediatric B-ALL, but not T-ALL [43]. Another interesting work shows that miR-19b, miR-20a, miR-26a, miR-92, and miR-223 have cooperative effects on tumor suppressor genes implicated in the pathogenesis of T-ALL, including *IKAROS*, *PTEN*, *BIM, PHF6, NF1,* and *FBXW7*. Interestingly, these miRNAs are

MiRNAs are suggested as promising biomarkers not only in the diagnosis but also in the prognosis of ALL patients. Since they have been promising in identifying subgroups of patients with different clinical outcomes [45]. It has been observed that ectopic expression of miRNAs leads to the development of leukemia, such is the case of miR-125b, which has been reported in mice transplanted with fetal liver

markers to differentiate T-ALL regardless of age (**Table 1**).

capable of promoting T-ALL development in a mouse model [44].

**5. MicroRNAs as prognostic markers in ALL**

*DOI: http://dx.doi.org/10.5772/intechopen.84318*

T-ALL miR-450b-5p, miR-

T-ALL miR-148a,

**Children**

TEL-AML1 BCR-ABL, E2A-PBX1, hyperdiploid, and B-other

E2A/PBX1-positive

MLL/AF4-positive

BCR/ABL-positive

**Adults**

B-ALL

B-ALL

B-ALL

**Table 1.**

**ALL subtype Upregulated expression Downregulated expression References Children** T-ALL miR-450b-5p, miR-450a-5p, miR-542-5p, miR-424-5p, miR-629-5p, miR-29c-5p miR-708-5p, miR-497-5p, miR-151a-5p, miR-151b, miR-371b-5p, miR-455-5p, miR-195-5p, miR-1266-5p, miR-574-5p, miR-425-5p [39] MLL-rearranged, T-ALL miR-196b [41] TEL-AML1 BCR-ABL, E2A-PBX1, hyperdiploid, and B-other miRNA-708 **Adults** T-ALL miR-148a, miR-424 miR-151 [40] E2A/PBX1-positive B-ALL miR-425-5p, miR-191, miR-128 MLL/AF4-positive B-ALL miR-629 BCR/ABL-positive B-ALL miR-146b, miR-126

*miRNAs in Acute Lymphoblastic Leukemia: Diagnosis, Prognosis and Target Therapeutic DOI: http://dx.doi.org/10.5772/intechopen.84318*

#### **Table 1.**

*Advances in Hematologic Malignancies*

entiation of the Th2 cells [35].

**lymphoblastic leukemia subtypes**

types with different miRNA signatures [37].

the presence of t(12;21) [39].

of Th1 differentiation [33]. Expression of miR-21 promotes Th2 differentiation in nonpolarized T cells [34]. miR-126 is another miRNA that also regulates the differ-

**4. miRNA expression and its role in the differential diagnosis of acute** 

Acute lymphoblastic leukemia (ALL) is characterized by clonal proliferation of early B- and T-lymphocyte progenitors that result in the accumulation of lymphoblasts in the bone marrow and various extramedullary sites. ALL is also the hematology neoplasia most commonly observed in the pediatric population, while it is relatively **less common than AML** in adults [36]. Around 75% of childhood ALL cases contain at least one alteration chromosomic, have lymphoid maturation arrest in distinct stages, and involve B- or T-lineages to leaving different immunopheno-

microRNAs participate in different physiological and cellular processes, such as development and tissue differentiation, cell identity, cell cycle progression, and programmed cell death [38]. Nowadays, it is known that the distinct stages of lymphopoiesis and the direction of lymphoid precursor maturation are influenced by miRNA expression differentially. However, an aberrant expression of miRNAs is related with malignant lymphopoiesis, characteristic that can be utilized as signature to diagnosis and classification diagnosis of acute lymphoblastic leukemia [18]. Interestingly, miRNA groups that can clearly differentiate ALL of its normal counterpart, B-ALL versus T-ALL and ALL subtypes with specific genetic abnormalities have been reported. De Oliveira and collaborators reported miRNA-128a and miRNA-181b overexpressed and miRNA-100, miRNA-196b, and let-7e with lower level when compared the miRNAs expression in normal pediatric bone morrow (BM) samples and BM samples of pediatric ALL. The authors point out miR-196b as a miRNA highly expressed in T-ALL, while miR-100 was related with

A study in Brazilian children with T- or B-cell acute lymphoblastic leukemia (T-ALL or B-ALL) evaluated a bone marrow miRNAs profile that may be used for distinguishing childhood lymphoblastic leukemia subtypes [40]. The authors mention that miR-708-5p, miR-497-5p, miR-151a-5p, miR-151b, miR-371b-5p, miR-455-5p, miR-195-5p, miR-1266-5p, miR-574-5p, miR-425-5p downregulated and miR-450b-5p, miR-450a-5p, miR-542-5p, miR-424-5p, miR-629-5p, miR-29c-5p upregulated in childhood T-ALL may be used for distinguishing childhood T- and B-ALL subtypes. However, a machine learning analysis showed that miR-29c-5p, which is involved in calcium signaling, is critical for B-cell lymphocyte fate. So, it is

In a series of adult ALL cases, the expression profile of 470 miRNAs was measured by microarray analysis; 3 miRNAs (miR-148, miR-151, and miR-424) were identified as discriminative of T-lineage versus B-lineage ALL; and miR-151 dramatically downmodulated an miR-148a and miR-424 with higher expression in patients with T-ALL [41]. Furthermore, in the B-lineage ALL cases with special molecular lesions, those with BCR/ABL, E2A/PBX1, MLL/AF4 rearrangements and cases lacking known genetic abnormalities can be differentiated by a set of six miRNA, which was highlighted by one-way analysis of variance [41]. These miRNAs were preferentially expressed in each chromosomic rearrangement; miR-425-5p, miR-191, and miR-128 were expressed in the E2A/PBX1-positive case, miR-629 was highly expressed in cases harboring MLL/AF4 rearrangement, while high levels of miR-146b and miR-126 were observed in the BCR/ABL-positive cases [41]. Other study in pediatric ALL showed

the best discriminator between childhood T- and B-ALL [40].

**60**

*Expression of miRNAs in children and adults to differentiate acute lymphoblastic leukemia subtypes.*

in seven major subtypes of pediatric ALL, which included: T-cell, MLL-rearranged, TEL-AML1-positive, E2A-PBX1-positive, hyperdiploid ALL, BCR-ABL-positive, and B-other ALLs, the differential miRNA expression. miRNA-708 was highly expressed in TEL-AML1, BCR-ABL, E2A-PBX1, hyperdiploid, and B-other cases than in the MLL-rearranged and T-ALL cases. On the other hand, the expression of miR-196b was higher in MLL-rearranged and T-ALL cases as compared with the expression level in the precursor B-ALL cases [42]. This information suggests that upregulated expression of miR-424 and downregulated expression of miR-151 might be good diagnostic markers to differentiate T-ALL regardless of age (**Table 1**).

Malik and collaborators propose a novel miR-2909-KLF4 molecular axis to differentiate the pathogeneses of pediatric B- and T-cell ALLs that may represent a new diagnostic marker, through alterations in miRNA expression patterns and their respective targets. The authors demonstrate the ability of miR-2909 to repress KLF4 expression in pediatric B-ALL, but not T-ALL [43]. Another interesting work shows that miR-19b, miR-20a, miR-26a, miR-92, and miR-223 have cooperative effects on tumor suppressor genes implicated in the pathogenesis of T-ALL, including *IKAROS*, *PTEN*, *BIM, PHF6, NF1,* and *FBXW7*. Interestingly, these miRNAs are capable of promoting T-ALL development in a mouse model [44].

#### **5. MicroRNAs as prognostic markers in ALL**

MiRNAs are suggested as promising biomarkers not only in the diagnosis but also in the prognosis of ALL patients. Since they have been promising in identifying subgroups of patients with different clinical outcomes [45]. It has been observed that ectopic expression of miRNAs leads to the development of leukemia, such is the case of miR-125b, which has been reported in mice transplanted with fetal liver

cells ectopically expressing miR-125b that showed an increase in white blood cell count, in particular in neutrophils and monocytes, associated with a macrocytic anemia. These mice developed B-cell acute lymphoblastic leukemia, T-cell acute lymphoblastic leukemia, or a myeloproliferative neoplasm, suggesting an important role for miR-125b in early hematopoiesis [46].

Patients group with high miR-21 expression was significantly associated with those aged <2 and > 10 years, lower platelets count, more incidence of central nervous system (CNS) infiltration, and poorer treatment outcome also; patients with high miR-21 showed a significantly poorer disease-free survival (DFS) and overall survival (OS) compared with those with low miR-21 expression group [47]. Also, miR-92a expression is significantly higher in ALL compared with peripheral blood mononuclear cells (PBMNCs) from healthy volunteers. Likewise, the expression levels of miR-99a, miR-100, and miR-128b correlated high-risk prognostic factors, including white blood cell (WBC) count, ALL subclassification (T-cell and B-cell ALL), the MLL-rearranged gene, and the BCR-ABL fusion gene, suggesting possible relation of miR-99a, miR-100, and miR-218b with prognosis [48, 49]. It has also been reported that mir-125b-2 is highly expressed in childhood ETV6/RUNX1 (TEL/AML1) leukemias and confers survival advantage to growth inhibitory signals independent of p53 [50].

More specifically, miR-9, miR-24, and miR-92a expression was significantly increased in a subset of ALL cells, and ALL patients with overexpressed miR-24 and miR-92a had poor prognoses [51–53]. Wang et al. (2010) observed that miR-146a, miR-181a/c, and miR-221 were significantly associated with overall survival of the ALL patients. Expression level of miR-146a and miR-181a/c was associated with a poor outcome (i.e., poor prognosis/short-term survival), whereas that of miR-221 was associated with a good outcome (i.e., good prognosis/long-term survival) [54], while that of miR-423-5p is associated with a poorer survival in patients with ALL [55]. Otherwise, the reduced expression of miR-155, miR-181b, miR-182, miR-143, miR-210, and miR-335 is associated with poor outcome of pediatric ALL [56–60]. Also, the expression of miRNAs miR-18a, miR-532, miR-218, miR-625, miR-193a, miR-638, miR-550, and miR-633 is associated with early relapse in childhood ALL, suggesting possible relation of these miRNAs with prognosis [61].

The high miR-16 expression is associated with hyperleukocytosis and poor cytogenetic groups. In B-cell ALL patients, the DFS was significantly shorter in patients with high miR-16 levels. While in T-cell ALL patients, for both DFS and overall survival, a significant trend was found with a survival shortening from the lowest to the highest miR-16 levels [62, 63]. Likewise, it was reported that the expression of miR-16 was upregulated in cases of T lymphoblastic lymphoma/leukemia (T-LBL/ ALL), and the high expression group of miR-16 was significantly correlated with longer over survival [64].

For instance, Gimenes-Teixeira et al. reported that T-ALL patients with high miR-221 expression had significantly lower 5-year overall survival (OS) rates compared with those with low miR-221 expression [65]. Oliveira et al. observed that lower levels of miR-29a were significantly associated with higher blast counts in the bone marrow and with increased disease-free survival in T-ALL patients [66].
