**2.3 Polymerase chain reaction (PCR)**

*Cytogenetics - Classical and Molecular Strategies for Analysing Heredity Material*

malities, and recently updated risk stratification systems.

the old technic saw improved their sensibility and specificity.

transcriptome sequencing, and epigenomics [5].

aspiration in sterile way with heparine filled probes.

**2.2 Fluorescence in situ hybridization (FISH)**

malities using molecular technology.

it can also be performed from fixed and sectioned tissue [3].

Sometimes, katyotype can be realised by peripheral blood.

**2.1 Conventional cytogenetic**

required time [2].

resource limited situation.

revolution on haematology [4].

genesis of these diseases.

**2. Cytogenetic technic**

neoplasms.

haematological diseases genetics.

Over the last few years, more powerful technologies were developed, the most remarkable one is was the Next-generation sequencing (NGS) witch was a real

In this chapter, we describe the techniques commonly employed for elucidating chromosomal aberrations, prognostic impact of recurrent chromosomal abnor-

We will summaries the chromosomal abnormalities recently identified on many of haematological diseases such acute myeloid leukaemia, acute lymphoid leukaemia, myelodysplasic syndrome, multiple myeloma, meyloproliferative disease and clarify their impacts on clinical phenotype and prognosis, as well as their role in the patho-

The aim of this chapter is to provide a brief overview of the recent progresses in

Cytogenetic study plays a crucial role on haematology, its the main outil for making diagnosis for almost all haematological malignancies, and its have an important impact prognosis for those diseases, and cytogenetic abnormalities are included on almost all prognosis score and risk stratifications for haematological

The past several years' remarkable efforts were deployed for better understanding of genetics and genome biology, many new technologies were developed, and

Veritable revolution was seen thanks to NGS, this technic offers possibility of broad analysis of a genome by whole-genome sequencing (WGS), exome sequencing,

Cytogenetic analyse in hematologicl neoplasms is performed by bone marrow

The Celle of aspirated bonne marrow are cultured in vitro, then microscopic slides with metaphases chromosomes and/or interphase nuclei is performed. Karyotype needs many metaphases cell (20 to 30) to be significatif, so its

Conventional cytogenetics still be the most frequently ordered genetic test for various leukaemias, most prominently chronic myelogenous leukaemia (CML) in a

FISH is the best alternative to karyotype, it is rapid technic, with high level of specificity and sensibility. It can be realised from bone marrow or peripheral blood,

FISH constitues a big step for studying somatic chromosomal mosaicism and molecular cytogenetic detection of chromosomal variations in interphase

FISH is a molecular cytogenetic technique, it identifies chromosomal abnor-

**90**

nuclei [6, 7].

PCR is a technique that involves amplification of a desired segment of DNA by using primers, nucleotides and enzymes like reverse transcriptase and DNA polymerases. It represents the most used molecular technique on haematology. Different types of PCRs exist. Reverse transcriptase PCR (RT-PCR), Real time PCR (RQ-PCR).

In clinical practice RQ PCR is commonly used for viral copies detection and specific gene detection like Bcr-abl and PML-RARA for assessment of treatment response [1] new technique was developted: Digital PCR it s used for DNA/RNA detection and quantification. It is emerging as an alternative to conventional RQ-PCR for quantification and low abundance mutation detection [8].

PCR have many applications in hematologic malignancies include. it s used for detection of fusion genes and mutations. Its also performed for analysing of post transplant chimerism, and can be realised for determination of lymphoid clonality.

#### **2.4 Genome-wide arrays**

Microarray based testing such as array comparative genomic hybridization (CGH) and single nucleotide polymorphisms (SNP) arrays are now more used in routine diagnostics for haematological malignancies.

The copy numbers of DNA sequences in the test and reference samples are quantified by assessment of relative fluorescence intensities detected by digital imaging systems.

## **2.5 Gene expression profiling**

This technique is based on DNA microarray which utilises plates which have various complementary genetic sequence covalently attached to them.

At present availability of GEP is limited to few research centers only limiting its wide use in clinical practice.

#### **2.6 New generation sequencing**

Over the past few years, an important increasing of the use of NGS on haematology have been shown, new platforms are available and are very helpful to identify the genetic basis of haematological neoplasms and genome biology.

Next-generation sequencing (NGS) encompasses several different methodologies that allow the investigation of genomics, transcriptomics and epigenomics [4].

Application of NGS in hematologic malignancies has confirmed presence of a lot of mutation of certain genes like TP53, ATM, RAS etc. the inconvenient for NGS, its the cost, this technic still expensive and can not be used on large spectre today especially for limited resource's country.

## **3. Haematological malignancies applications**

Haematological neoplasms benefits from progress of biological technology, the use of new platforms helps to approve the performance of identification of genetic abnormalities.

This knowledge is crucial and have clinical utility, it can also improve diagnostics, prognosis, monitoring of minimal residual disease and it can be helpful to target of dysregulated signalling pathways by specific therapeutic targets [4].

We will summaries the chromosomal abnormalities recently identified on many of haematological diseases such acute myeloid leukaemia, acute lymphoid leukaemia, myelodysplasic syndrome, multiple myeloma, chronic lymphoblastic leukaemia, meyloproliferative disease and clarify their impacts on clinical phenotype and prognosis, as well as their role in the pathogenesis of these diseases.

#### **3.1 Acute myeloid leukaemia (AML)**

Cytogenetic abnormalities are frequently reported in the literature describing the presence of chromosomal rearrangements in important cases of acute myeloid leukaemia (AML): the rate can reach 50–60% of cases of AML [9].

It has been proved that AML is a complex and evaluative disease [10, 11]. There are many leukaemia genes, most of which are infrequently mutated, and patients typically have more than one driver mutation. The AML evolved over time, with multiple competing clones coexisting at any time [10, 11].

Over the few recent years, genome biology have seen a veritable revolution of technology, including chromosome banding, with fluorescence/chromosome in situ hybridization, or other analyses like array comparative network genomic hybridization, genome breakpoints cloning and Sanger Sequencing of candidate genes and profiling of single nucleotide polymorphism, and even whole-genome sequencing (WGS), whole-exome sequencing (WES), and RNA sequencing have all contributed to incremental improvements in understanding the genetic basis of the AML.

The whole-genome sequencing for AML showed that it is an evaluative and complex disease. There are many leukemia genes, most of which are infrequently mutated, and patients typically have many driver mutations. The evolution is characterized by emergence of many competing clones which can coexist at any time.

In fact, it has been proved that different genes and clones coexisting in the same patient, **Figure 1** illustrates that clearly [12].

200 AML patients has been analysed by The Cancer Genome Atlas (TCGA) consortium, they use whole-genome or whole-exome sequencing and they identified 23 genes as "significantly mutated" at a higher-than-expected frequency [13].

Conventional cytogenetics is very important on AML, it identifies chromosomal abnormalities, it can be balanced translocations, inversions, insertions, monosomies, and trisomies, which are present in approximately 55% of adult cases and 80% of children with AML. These are the strongest prognostic factors for response to treatment and survival in multivariate analysis. The 2008 WHO classification categorized AML based on cytogenetic or molecular abnormalities [14, 15].

The WHO 2008 and 2016 classifications incorporated modifications that allowed for a greater number of patients to be classified into the category of AML [16, 17].

Even patient with normal karyotype AML, it has been proved recently with certitude that those patients constituted very heterogeneous group; new technology helps to identified many gene mutations in normal karyotype AML by cutting-edge next-generation sequencing NGS technology, like FLT3-ITD, NPM1, CEBPA, and other additional mutations.

The most important predictors of shorter overall survival in AML patients aged less than 60 years are represented by DNMT3A and RUNX1 mutations especially those with intermediate-risk cytogenetic.

**93**

*Haematological Malignancies: Overview of the Recent Progresses in Genetics*

NPM1 mutations were also considered as important molecular prognosticators of Overall Survivor, particularly in the absence of FLT3-ITD, mutated TP53, and

Actually, for treating patient with AML, it is indispensable to perform the research of these gene mutations. It s important for diagnosis and it can be helpful as molecular marker of prognosis, and its necessary to predictive for response of

*Molecular classes of AML and concurrent gene mutations in adult patients up to the age of ~65 years. For each AML class denoted in the pie chart, frequent co-occurring mutations are shown in the respective boxes. Data on the frequency of genetic lesions are compiled from the databases of the British Medical Research Council (MRC) and the German-Austrian AML study group (AMLSG) and from selected studies. It indicates cohesin genes including RAD21 (10%), SMC1A (5%), and SMC3 (5%); inv(16)(p13.1q22) or t(16;16)(p13.1;q22); CBFB-MYH11; and inv(3)(q21.3q26.2) or t(3;3)(q21.3;q26.2); GATA2,MECOM(EVI1),* 

*and TP53 mutations are found in 45% and complex karyotypes in 70% of this class.*

Studies demonstrated that patients with cytogenetically normal AML or intermediate-risk abnormalities have more additional gene mutations than patients with favourable or unfavourable abnormal cytogenetic and especially those with

Recent research showed that aged patients have more driver gene mutations than younger patients. Its seems that elderly patients have more alterations in specific genes including TET2, RUNX1, ASXL1, and SRSF2. All this gene has recently been implicated in age-related clonal haematopoiesis. These found participate to improve our understanding of knowledge in AML biology between younger and older

The application for biological technology such NSG are multiple, for exemple, there are a number of FLT3 inhibitors at various stages of clinical development were produced, such as PKC412 (midostaurin), CEP-701 (lestaurtinib), or MLN518

TKIs are promising agents in the treatment of AML patients with an FLT3-ITD

Acute lymphoblastic leukaemia (ALL) is the most often childhood neoplasm

mutation, especially when they are combined with chemotherapy [20].

*DOI: http://dx.doi.org/10.5772/intechopen.96913*

bi-allelic CEBPA mutations.

**Figure 1.**

balanced translocations [18].

patients [19].

(tandutinib).

treatment, and used also for disease monitoring.

**3.2 Acute lymphoblastic leukaemia (ALL)**

occuring about 30% of all cancer.

*Haematological Malignancies: Overview of the Recent Progresses in Genetics DOI: http://dx.doi.org/10.5772/intechopen.96913*

#### **Figure 1.**

*Cytogenetics - Classical and Molecular Strategies for Analysing Heredity Material*

prognosis, as well as their role in the pathogenesis of these diseases.

leukaemia (AML): the rate can reach 50–60% of cases of AML [9].

multiple competing clones coexisting at any time [10, 11].

patient, **Figure 1** illustrates that clearly [12].

**3.1 Acute myeloid leukaemia (AML)**

This knowledge is crucial and have clinical utility, it can also improve diagnostics, prognosis, monitoring of minimal residual disease and it can be helpful to target of dysregulated signalling pathways by specific therapeutic targets [4].

We will summaries the chromosomal abnormalities recently identified on many of haematological diseases such acute myeloid leukaemia, acute lymphoid leukaemia, myelodysplasic syndrome, multiple myeloma, chronic lymphoblastic leukaemia, meyloproliferative disease and clarify their impacts on clinical phenotype and

Cytogenetic abnormalities are frequently reported in the literature describing the presence of chromosomal rearrangements in important cases of acute myeloid

It has been proved that AML is a complex and evaluative disease [10, 11]. There are many leukaemia genes, most of which are infrequently mutated, and patients typically have more than one driver mutation. The AML evolved over time, with

Over the few recent years, genome biology have seen a veritable revolution of technology, including chromosome banding, with fluorescence/chromosome in situ hybridization, or other analyses like array comparative network genomic hybridization, genome breakpoints cloning and Sanger Sequencing of candidate genes and profiling of single nucleotide polymorphism, and even whole-genome sequencing (WGS), whole-exome sequencing (WES), and RNA sequencing have all contributed to incremental improvements in understanding the genetic basis of the AML. The whole-genome sequencing for AML showed that it is an evaluative and complex disease. There are many leukemia genes, most of which are infrequently mutated, and patients typically have many driver mutations. The evolution is characterized by emergence of many competing clones which can coexist at any time. In fact, it has been proved that different genes and clones coexisting in the same

200 AML patients has been analysed by The Cancer Genome Atlas (TCGA) consortium, they use whole-genome or whole-exome sequencing and they identified 23 genes as "significantly mutated" at a higher-than-expected fre-

Conventional cytogenetics is very important on AML, it identifies chromosomal abnormalities, it can be balanced translocations, inversions, insertions, monosomies, and trisomies, which are present in approximately 55% of adult cases and 80% of children with AML. These are the strongest prognostic factors for response to treatment and survival in multivariate analysis. The 2008 WHO classification categorized AML based on cytogenetic or molecular abnormalities [14, 15]. The WHO 2008 and 2016 classifications incorporated modifications that allowed for a greater number of patients to be classified into the category of AML

Even patient with normal karyotype AML, it has been proved recently with certitude that those patients constituted very heterogeneous group; new technology helps to identified many gene mutations in normal karyotype AML by cutting-edge next-generation sequencing NGS technology, like FLT3-ITD, NPM1, CEBPA, and

The most important predictors of shorter overall survival in AML patients aged less than 60 years are represented by DNMT3A and RUNX1 mutations especially

**92**

quency [13].

[16, 17].

other additional mutations.

those with intermediate-risk cytogenetic.

*Molecular classes of AML and concurrent gene mutations in adult patients up to the age of ~65 years. For each AML class denoted in the pie chart, frequent co-occurring mutations are shown in the respective boxes. Data on the frequency of genetic lesions are compiled from the databases of the British Medical Research Council (MRC) and the German-Austrian AML study group (AMLSG) and from selected studies. It indicates cohesin genes including RAD21 (10%), SMC1A (5%), and SMC3 (5%); inv(16)(p13.1q22) or t(16;16)(p13.1;q22); CBFB-MYH11; and inv(3)(q21.3q26.2) or t(3;3)(q21.3;q26.2); GATA2,MECOM(EVI1), and TP53 mutations are found in 45% and complex karyotypes in 70% of this class.*

NPM1 mutations were also considered as important molecular prognosticators of Overall Survivor, particularly in the absence of FLT3-ITD, mutated TP53, and bi-allelic CEBPA mutations.

Actually, for treating patient with AML, it is indispensable to perform the research of these gene mutations. It s important for diagnosis and it can be helpful as molecular marker of prognosis, and its necessary to predictive for response of treatment, and used also for disease monitoring.

Studies demonstrated that patients with cytogenetically normal AML or intermediate-risk abnormalities have more additional gene mutations than patients with favourable or unfavourable abnormal cytogenetic and especially those with balanced translocations [18].

Recent research showed that aged patients have more driver gene mutations than younger patients. Its seems that elderly patients have more alterations in specific genes including TET2, RUNX1, ASXL1, and SRSF2. All this gene has recently been implicated in age-related clonal haematopoiesis. These found participate to improve our understanding of knowledge in AML biology between younger and older patients [19].

The application for biological technology such NSG are multiple, for exemple, there are a number of FLT3 inhibitors at various stages of clinical development were produced, such as PKC412 (midostaurin), CEP-701 (lestaurtinib), or MLN518 (tandutinib).

TKIs are promising agents in the treatment of AML patients with an FLT3-ITD mutation, especially when they are combined with chemotherapy [20].

#### **3.2 Acute lymphoblastic leukaemia (ALL)**

Acute lymphoblastic leukaemia (ALL) is the most often childhood neoplasm occuring about 30% of all cancer.

Abnormalities in chromosome number as well as structural rearrangements (translocations) are detected in 60–80% of patients with ALL, whereas the remaining 20–40% have a normal karyotype [21, 22].

Improvement of cytogenetic, FISH, and reverse transcription polymerase chain reaction (RT-PCR) analyses permits to identify subgroups of acute lymphoblastic leukaemia with specific chromosome abnormalities and allow determining treatment strategy for childhood ALL, especially when specific aberrations are present [23, 24].

Recurrent genetic abnormalities have been identified on ALL, including balanced translocations and aneuploidies. Based on the World Health Organization (WHO) classification, BCP-ALL is categorized into ALL with hyperdiploidy (>50 chromosomes), ALL with hypodiploidy (<44 chromosomes), and ALL with translocation t(9;22) (q34;q11.2) encoding BCR–ABL1, t(12;21) (p13;q22) encoding TEL–AML1, t(1;19) (q23;p13.3) encoding E2A–PBX1, t(5;14) (q31;q32) encoding IL3–IGH, and rearrangement of MLL at 11q23, with a diverse range of partner genes [25, 26].

Concerning T- ALL, common alterations include rearrangement of the T-cell receptor gene loci to transcription factor genes including TLX1, TLX3, LYL1, TAL1, and MLL [27].

ALL genomes are not static but exhibit acquisition of new chromosomal abnormalities over time. Single-nucleotide polymorphism microarray profiling studies of matched diagnosis–relapse ALL samples show that most ALL cases exhibit changes in the patterns of structural genomic alterations from diagnosis to relapse and that many relapse-acquired lesions, including those targeting genes associated with high-risk ALL (IKZF1, IKZF2, CDKN2A, and CDKN2B), are detectable at the diagnosis [28, 29].

#### **3.3 Myelodysplasic syndrome**

Myelodysplasia syndromes (MDS) are defined by a heterogeneous group of myeloid malignancies characterised by peripheral blood cytopenia and dishematopoiesis and frequently progress to acute myeloid leukaemia.

The 2016 revision defines 10 MDS subtypesas follows:


Conventional cytogenetic allow the identification of abnormalities in approximately 50% of MDS. Some of cytogenetic abnormalities are characteristic of MDS, they may be considered as specific to MDS if the clinical context is appropriate such del(5q).

**95**

*Haematological Malignancies: Overview of the Recent Progresses in Genetics*

Majority of of MDS (90%) presents somatic mutations, those mutations identify molecular pathways that drive the pathogenesis of MDS. Even low abundance mutations can have prognostic value as they identify emerging clones before they

Recent studies demonstrated that 65% of MDS patients harboured mutations in RNA splicing (SF3B1, SRSF2, U2AF1, ZRSR2) [28], 47% harbouring mutations in DNA methylation genes (DNMT3A, IDH1/2, TET2) [29, 30] and 28% in histone

Mutations in isocitrate dehydrogenase 1 or 2 (IDH1 and IDH2) are important to identify at the time of diagnosis of high- or very high-risk MDS. These particular mutations lead to abnormal leukemogenesis. Mutated IDH1 or IDH2 are not common and are only found in approximately 4% to 12% of patients with MDS. Those gene mutations have treatment impact. Recently, two IDH inhibitors, specifically ivosidenib targeting IDH1 and enasidenib for IDH2, are approved by the United States Food and Drug Administration (FDA) for use in AML, but not in MDS [30, 31]. At present, NGS is rarely incorporated into clinical guidelines although an increasing number of studies have demonstrated the benefit of using NGS in the

Multiple myeloma is a malignant disease characterised by proliferation of monoclonal plasma cells leading to clinical features that include hypercalcaemia, renal dysfunction, anaemia, and bone disease (frequently referred to by the

Recent studies have confirmed that myeloma is an heterogeneous disease composed of multiple molecularly-defined subtypes each with varying clinicopatho-

Chromosomal translocations account for 40–50% of primary events in myeloma

Karyotypes are complex, hyperploidy can be seen in2/3 of cases, karyotypes can

Fluorescence in situ hybridization (FISH) ssems to be more adequate for recognising specific chromosomal changes in quiescent cells and increases the proportion of detection of chromosomal abnormalities in MM up to more than 90% [35].

IG rearrangements: translocations involving 14q32 are found in at least 65–70% of patients, most of them result from short segments exchange and are detected

The (4; 14) is present in 15% of myeloma cases and has been associated with a poor prognosis in a variety of clinical settings such as those receiving high dose

The (11; 14) is observed in approximately 17% of myeloma patients and also

The (6; 14) is a rare translocation occurring in 2% of myeloma patients which

Other translocations with IGH involving are reported, but they are rare and it

Like other haematological neoplasm, multiple myelom benefits from the development of molecular technic like NGS, the knowledge about pathogenesis and the progression of disease has been improved, with apparition of a new concept called subclonality. In fact, NGS characterised the the wide molecular heterogeneity of the disease and the frequent occurrence of some supposedly "driver" mutations only in

subclones. Those found are important for the targeted future therapies [38].

change from normal to abnormal during evolution of multiple myoloma.

acronym CRAB) which represent evidence of end organ failure.

*DOI: http://dx.doi.org/10.5772/intechopen.96913*

modification genes (ASXL1,BCOR, EZH2).

clinical management of MDS patients [32].

logical features and disease outcomes [33].

and strongly influence disease phenotype [34].

therapy with autologous stem cell transplant (ASCT).

directly up regulates a cyclin D gene in the form CCND1.

results in the direct up regulation of the CCND3 gene [36].

**3.4 Multiple myeloma**

quite exclusively by FISH.

seems that they are secondary [37].

impact clinical parameters.

Majority of of MDS (90%) presents somatic mutations, those mutations identify molecular pathways that drive the pathogenesis of MDS. Even low abundance mutations can have prognostic value as they identify emerging clones before they impact clinical parameters.

Recent studies demonstrated that 65% of MDS patients harboured mutations in RNA splicing (SF3B1, SRSF2, U2AF1, ZRSR2) [28], 47% harbouring mutations in DNA methylation genes (DNMT3A, IDH1/2, TET2) [29, 30] and 28% in histone modification genes (ASXL1,BCOR, EZH2).

Mutations in isocitrate dehydrogenase 1 or 2 (IDH1 and IDH2) are important to identify at the time of diagnosis of high- or very high-risk MDS. These particular mutations lead to abnormal leukemogenesis. Mutated IDH1 or IDH2 are not common and are only found in approximately 4% to 12% of patients with MDS. Those gene mutations have treatment impact. Recently, two IDH inhibitors, specifically ivosidenib targeting IDH1 and enasidenib for IDH2, are approved by the United States Food and Drug Administration (FDA) for use in AML, but not in MDS [30, 31].

At present, NGS is rarely incorporated into clinical guidelines although an increasing number of studies have demonstrated the benefit of using NGS in the clinical management of MDS patients [32].

#### **3.4 Multiple myeloma**

*Cytogenetics - Classical and Molecular Strategies for Analysing Heredity Material*

ing 20–40% have a normal karyotype [21, 22].

Abnormalities in chromosome number as well as structural rearrangements (translocations) are detected in 60–80% of patients with ALL, whereas the remain-

Improvement of cytogenetic, FISH, and reverse transcription polymerase chain reaction (RT-PCR) analyses permits to identify subgroups of acute lymphoblastic leukaemia with specific chromosome abnormalities and allow determining treatment strategy for childhood ALL, especially when specific aberrations are present [23, 24]. Recurrent genetic abnormalities have been identified on ALL, including balanced translocations and aneuploidies. Based on the World Health Organization (WHO) classification, BCP-ALL is categorized into ALL with hyperdiploidy (>50 chromosomes), ALL with hypodiploidy (<44 chromosomes), and ALL with translocation t(9;22) (q34;q11.2) encoding BCR–ABL1, t(12;21) (p13;q22) encoding TEL–AML1, t(1;19) (q23;p13.3) encoding E2A–PBX1, t(5;14) (q31;q32) encoding IL3–IGH, and rearrangement of MLL at 11q23, with a diverse range of partner genes

Concerning T- ALL, common alterations include rearrangement of the T-cell receptor gene loci to transcription factor genes including TLX1, TLX3, LYL1, TAL1,

ALL genomes are not static but exhibit acquisition of new chromosomal abnormalities over time. Single-nucleotide polymorphism microarray profiling studies of matched diagnosis–relapse ALL samples show that most ALL cases exhibit changes in the patterns of structural genomic alterations from diagnosis to relapse and that many relapse-acquired lesions, including those targeting genes associated with high-risk ALL (IKZF1, IKZF2, CDKN2A, and CDKN2B), are detectable at the

Myelodysplasia syndromes (MDS) are defined by a heterogeneous group of myeloid malignancies characterised by peripheral blood cytopenia and dishemato-

• MDS with dysplasia in two or more myeloid lineages (MDS-MLD),

• MDS-SLD/MLD with ≥15% ring sideroblasts (RSs; MDS-MLD-RS),

• MDS with isolated deletion of chromosome 5q [del(5q)]

• MDS with an excess of blasts of up to 9% in bone marrow and up to 4% in

• MDS with 10%–19% bone marrow and 5%–19% blood blasts (MDS-EB-2),

• MDS unclassifiable (MDS-U) based on defining cytogenetic abnormality, MDS-U with SLD and pancytopenia and MDS-U with 1% blood blasts.

Conventional cytogenetic allow the identification of abnormalities in approximately 50% of MDS. Some of cytogenetic abnormalities are characteristic of MDS, they may be considered as specific to MDS if the clinical context is appropriate

poiesis and frequently progress to acute myeloid leukaemia. The 2016 revision defines 10 MDS subtypesas follows:

• MDS with single lineage dysplasia (MDS-SLD),

peripheral blood (MDSEB- 1),

**94**

such del(5q).

[25, 26].

and MLL [27].

diagnosis [28, 29].

**3.3 Myelodysplasic syndrome**

Multiple myeloma is a malignant disease characterised by proliferation of monoclonal plasma cells leading to clinical features that include hypercalcaemia, renal dysfunction, anaemia, and bone disease (frequently referred to by the acronym CRAB) which represent evidence of end organ failure.

Recent studies have confirmed that myeloma is an heterogeneous disease composed of multiple molecularly-defined subtypes each with varying clinicopathological features and disease outcomes [33].

Chromosomal translocations account for 40–50% of primary events in myeloma and strongly influence disease phenotype [34].

Karyotypes are complex, hyperploidy can be seen in2/3 of cases, karyotypes can change from normal to abnormal during evolution of multiple myoloma.

Fluorescence in situ hybridization (FISH) ssems to be more adequate for recognising specific chromosomal changes in quiescent cells and increases the proportion of detection of chromosomal abnormalities in MM up to more than 90% [35].

IG rearrangements: translocations involving 14q32 are found in at least 65–70% of patients, most of them result from short segments exchange and are detected quite exclusively by FISH.

The (4; 14) is present in 15% of myeloma cases and has been associated with a poor prognosis in a variety of clinical settings such as those receiving high dose therapy with autologous stem cell transplant (ASCT).

The (11; 14) is observed in approximately 17% of myeloma patients and also directly up regulates a cyclin D gene in the form CCND1.

The (6; 14) is a rare translocation occurring in 2% of myeloma patients which results in the direct up regulation of the CCND3 gene [36].

Other translocations with IGH involving are reported, but they are rare and it seems that they are secondary [37].

Like other haematological neoplasm, multiple myelom benefits from the development of molecular technic like NGS, the knowledge about pathogenesis and the progression of disease has been improved, with apparition of a new concept called subclonality. In fact, NGS characterised the the wide molecular heterogeneity of the disease and the frequent occurrence of some supposedly "driver" mutations only in subclones. Those found are important for the targeted future therapies [38].
