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Chapter 56 Conventional and Molecular Cytogenomic Basis of Hematologic Malignancies 839

in both the API2 and MALT1 genomic sequences require more
precise molecular cytogenetic methods. Genetic Testing for Non-Hodgkin Lymphoma
t(1;14)(p22;q32) and its variant t(1;2)(p22;q12) occur in less than
5% of MALT lymphomas, and are associated with an advanced stage FISH testing can be performed on touch preparations, paraffin-
embedded tissue, or bone marrow cells if involved in disease. Periph-
of the disease. This translocation relocates the entire BCL10 gene eral blood cells are not appropriate for genetic testing of non-Hodgkin
from 1p22 to chromosome 14, bringing it under the control of an lymphoma. Bone marrow cytogenetics at diagnosis is not mandatory
IGH enhancer. Currently, a specific probe for detecting BCL10-IGH but is important for staging in some cases. Fig. 56.42 lists the most
fusion is not commercially available, but an IGH dual-color break- useful FISH probes for detection of genomic defects in non-Hodgkin
apart FISH probe may determine IGH rearrangements without lymphoma. They include CCND1-IGH, IGH-BCL2, MYC-IGH, BCL6,
identifying the partner chromosome. The recurrent t(3;14)(p13;q32) MYC for detection of Burkitt variant not associated with t(8;14), and
abnormality, which results in fusion of the FOXP1F gene on chromo- MALT1 for detection of t(11;18). For patients with suspected anaplastic
some 3 to IGH, is a rare rearrangement that causes FOXP1F overex- large cell lymphoma, ALK probe should be informative.
pression. Its significance in lymphoma remains unknown. The
t(14;18)(q32;q21), which occurs in 2% to 18% of all MALT lym-
phomas, results in fusion of IGH-MALT. The 18q21 breakpoint most frequent mutation in WM. It is found in over 91% of cases,
involving the MALT1 gene is 5 Mb centromeric from the BCL2 including those with a gain of chromosome 3 (approximately 12%).
breakpoint on chromosome 18, which is associated with FL. In This mutation may be considered as the first detectable genetic hit
contrast to API2-MALT1 cases, patients with IGH-MALT1 fusion in WM that promotes NF-κB and JAK-STAT signaling pathways.
have disease outside the gastrointestinal tract, usually presenting with MYD88 L265P mutation is also seen in SMZL and MALT lym-
ocular, skin, liver, or salivary gland tumors. FISH studies are useful phoma. MYD88 L265P cannot be used to differentiate between WM
for detecting the IGH-MALT fusion in paraffin-embedded lymph and IgM MGUS. Patients with mutated MYD88 have a shorter
node biopsies. Compelling evidence links these translocations to survival as compared with patients with unmutated MYD88. The
constitutive activation of the NF-κB pathway. CXCR4 (2q22.1 locus), one of the main regulators of B-cell homing
Splenic marginal zone lymphoma (SMZL) lacks recurrent chro- is mutated in almost 30% of patients with WM and 20% of patients
mosome translocations although approximately 30% of patients have with IgM MGUS. Collectively these results indicate that CXCR4 is
heterozygous 7q deletions. Studies utilizing whole-exome sequencing an activating mutation in WM and has a critical role in WM
have identified a novel and recurrent inactivating mutations in pathogenesis.
Krupel-like factor 2 (KLF2) in 42% of patients. These mutations are Little progress has been made in delineating recurrent chromo-
rarely observed in other B-cell lymphomas. Different KLF2 muta- somal abnormalities in Hodgkin lymphoma (HL) (see Chapter 75).
tions compromise the ability of KLF2 gene to suppress NF-κB activa- Less than 1% of the cells in HL are Reed-Sternberg cells, of B-cell
tion, leading to an altered gene expression pattern favoring B-cell origin. The most specific chromosomal abnormalities in HL are
homing to the marginal zone. KLF2 inactivation alone is insufficient hyperdiploidy/tetraploidy with tremendous variations in chromo-
for malignant transformation and requires cooperating genetic events. some number, indicating heterogeneity from patient to patient. Even
Other common mutations found in SMZL include NOTCH2, with use of nine different centromeric probes, no specific numerical
TRAF3, TNFAIP3, and CARD11. These mutations with and without chromosomal abnormality has been identified. Deletions of 1p, 4q,
KLF2 mutations indicate that KLF2 mutation identifies a subset of 6q, and 7q are recurrent, and JAK2, located on 9p21, frequently is
patients with SMZL with distinct genotype. NGS has demonstrated amplified in patients with HL. Another gene, REL, on chromosome
that 40% of patients with SMZL have mutations in fours genes: 2, band region p14–p15, is amplified in 50% of patients. Of note,
TP53, KLF2, NOTCH2, and TRAF3, of which TP53 mutations were REL is under the influence of NF-κB transcription factor, and con-
associated with a shorter survival whereas mutations in MYB gene stitutive NF-κB activation is a critical prerequisite for Reed-Sternberg
were predictors of a longer OS. cell survival and proliferation.
Lymphoplasmacytoid lymphoma (LPL) is a small lymphocytic The genome of Reed-Sternberg cells was largely unexplored
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lymphoma with plasmacytoid differentiation (CD5 CD10 ) charac- because of the technical difficulties in isolating these cells. This dif-
terized by t(9;14)(p13;q32) in approximately 50% of cases. As a ficulty was recently overcome and preliminary results of full-exome
result of this translocation, the paired homeobox 5 (PAX5) gene on deep sequencing of 10 patients revealed inactivating mutations in
9p13 moves to the IGH locus on der(14), causing dysregulation of β2-microglobulin gene (B2M, localized at 15q21.1) in 7 of 10
PAX5. Molecular characterization of t(9;14) has revealed that the patients. These mutations lead to the loss of major histocompatibility
coding region of the PAX5 gene remains intact in some patients. complex class I (MHC1) expression. A lack of MHC1 expression in
t(9;14) should be considered a regulatory mutation in which the Reed-Sternberg cells was previously reported as an independent
PAX5 gene is brought under the control of the IGH locus. In other adverse prognostic factor. Therefore molecular mechanisms leading
cases, molecular studies of t(9;14) reveal that the breakpoint occurs to MHC1 downregulation in Hodgkin lymphoma is through inacti-
upstream of the PAX5 promoter, leading to insertion of the IGH vating mutations in B2M gene.
enhancer upstream of the PAX5 gene. Recurrent mutations in
MYD88, specifically L265P mutation, have been identified in over
96% of patients with LPL whereas mutations in chemokine receptor, HAIRY CELL LEUKEMIA
CXCR4, has been observed in about 36% of cases. These mutations
identify a subgroup of LPL with aggressive disease. No recurrent chromosomal aberrations have been identified in hairy
Waldenström macroglobulinemia (WM) (see Chapter 87) is cell leukemia (HCL) although as shown in Fig. 56.59 some patients
characterized by a lymphoplasmacytic clonal expansion in the marrow. may have a complex bone marrow karyotype at diagnosis. HCL is a
Historically recurrent chromosomal abnormalities include deletion of mature B-cell malignancy with the bone marrow, spleen and liver
the long arms of chromosome 6 in 21% to 55% of cases and trisomy infiltrated by leukemic B cells that have abundant cytoplasm with
4 present in approximately 20% of cases. In contrast to other B-cell hairy-looking projections and unique immunophenotyping features.
disorders, abnormalities of IGH at 14q32 as detected by FISH are High-density genome-wide SNP genotyping have shown a remark-
rarely observed. Whole-genome sequencing of WM lymphoplasma- able balanced genomic profile. In 2011, a BRAF-V600E mutation
cytic cells has led to the identification of a recurring sequence variant was described for the first time in every one of 47 patients. The
at position 38182641 in chromosome 3p22.2. A single-nucleotide BRAF-V600E mutation defines HCL, is present in virtually all
change from T to C in the MYD88 gene resulted in a leucine-to- patients with HCL, and is absent in other B-cell malignancies except
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proline change at amino acid position 265. MYD88 (3p22 locus) a small number of patients with MM. BRAF is a kinase within the
mutations are detected by PCR in the peripheral blood of untreated RAS-RAF-MEK-ERK pathway that plays a pivotal role in regulating
patients. The MYB88 L265P mutation is highly specific and is the hematopoietic stem cells from patients with HCL. Quantitative