Novel identification of t(14;18)(q32;q21)/IGH::MALT1 in chronic lymphocytic leukemia

Abstract

Chronic lymphocytic leukemia (CLL) is a clonal B-cell malignancy and remains a chronic disease despite improvements in clinical outcomes since the use of targeted therapies. Both clinical and biological parameters are important for determining prognosis. Unlike other mature B-cell lymphomas, translocations involving the immunoglobulin heavy chain (IGH) locus are uncommon in CLL. There have been few case reports of CLL harboring t(14;18)/IGH::BCL2 and t(14;19)/IGH::BCL3. Here we describe the first two cases of patients with CLL with documented t(14;18)(q32;q21)/IGH::MALT1. Both cases in this report were associated with lower-risk biological parameters. Thus, FISH testing for MALT1 in cases with unknown IGH translocation partners in the setting of CLL should be implemented in clinical practice to better define such cases.

Chronic lymphocytic leukemia (CLL) is a B-cell neoplasm composed of a clonal proliferation of mature B lymphocytes involving the peripheral blood (PB), bone marrow (BM), and lymphoid tissues. Despite significant improvements in clinical outcomes since the advent of targeted therapies, CLL remains a chronic disease.¹ Clinical and biological parameters are important for determining prognosis. Biological parameters that influence prognosis include cytogenetics studies for structural variants and copy number alterations, mutation analysis (particularly for TP53), and assessment of immunoglobulin heavy variable (IGHV) gene mutational status. For example, deletion of the long arm of chromosome 13, del(13q) is associated with an indolent course if present in isolation and TP53 deletion and/or mutation predicts adverse outcome.^1–3 The presence or absence of somatic hypermutation (SHM) is also important for risk stratification in CLL, and unmutated IGHV gene is predictive of shorter progression-free survival and considered a high-risk genomic subgroup.^1,4–6

In ~80% of patients with CLL, at least one of four recurrent chromosomal aberrations can be identified by fluorescent in situ hybridization (FISH), including del(13q), trisomy 12, del(11q)/ATM, and del(17p)/TP53.⁷ However, unlike other mature B-cell lymphomas, translocations involving the immunoglobulin heavy chain (IGH) locus are uncommon in CLL. There have been few case reports of CLL harboring t(14;18)(q32;q21)/IGH::BCL2, a translocation frequently seen in follicular lymphoma.^8–10 CLL with t(14;19)(q32;q13)/IGH::BCL3 has also been occasionally described.¹ Here we describe the first two cases of patients with CLL with documented t(14;18)(q32;q21)/IGH::MALT1.

Patient 1 was a 54-year-old male who presented with bilateral neck swelling, loss of appetite, and 10–15 pounds unintentional weight loss. Laboratory studies upon presentation to our institution revealed leukocytosis with a white blood cell (WBC) count of 11.5 K/μL (reference range: 4–11 K/μL) along with relative and absolute lymphocytosis (59%, 6.8 K/μL, reference range: 12.2–47.4%, 0.9–3.2 K/μL). Computed tomography imaging demonstrated diffuse bilateral cervical and axillary adenopathy as well as mild adenopathy in the abdomen and pelvis.

A BM biopsy performed at an outside institution was reviewed and demonstrated an interstitial and nodular infiltrate composed of small, atypical lymphocytes with condensed chromatin and scant cytoplasm (Figure 1A), involving 15% of the marrow space. Flow cytometry performed on the PB showed an abnormal lambda restricted CD5-positive B-cell population expressing CD19, CD20-dim, CD22-dim, CD23, CD43, CD79b-partial dim, CD81-dim, CD200-bright, ROR1, and CD25-partial (Figure 1B). This population represented 45% (5.2 K/μL) of WBCs. Karyotyping analysis of a stimulated BM sample using IL2/DSP30 identified t(14;18)(q32;q21) in 17/20 metaphase cells.

Figure 1:

Histologic characteristics and flow cytometry data from Patient 1 with chronic lymphocytic leukemia. (A) Histologic sections of the BM core biopsy showed an interstitial and nodular infiltration of small size atypical lymphocytes with condensed chromatin and scant cytoplasm (H&E x20; inset bottom right x40). (B) Flow cytometry showed an abnormal lambda restricted CD5-positive B-cell population (cyan colored events) expressing CD19, CD20-dim, CD79b-partial dim, ROR1, CD23, CD200-bright, CD40, CD43, CD38, and CD81-dim, while negative for CD10. The blue colored events indicate background normal B-cells, also seen as blue and red events in the kappa and lambda plot (bottom right).

FISH studies performed at our institution on the PB were negative for IGH::BCL2 fusion/t(14;18) and CCND1::IGH fusion/t(11;14). However, FISH analysis using a break-apart probe sets detected chromosomal translocation of IGH and MALT1 in 26.7% and 40%, respectively (Figure 2A and 2B). There was no evidence of TP53 (17p13.1) deletion or loss of chromosome 17. SNP-array analysis detected a small deletion of 3 Mb in segment 13q14.2-q14.3 including the RB1, SETDB2, MIR16–1, DLEU2 genes and a portion of the DLEU1 gene. Evaluation of IGHV SHM identified a clonal rearrangement involving IGH V3–23_J4 with a 2% mutation frequency, which qualifies as mutated status, although the percentage is close to the 98% cutoff and considered borderline-mutated.⁴ Overall, these findings supported a diagnosis of CLL.

Figure 2:

FISH analysis and MSK-IMPACT Heme assay results. (A) FISH studies showed a split signal indicating chromosomal translocation of IGH in the neoplastic cells (5’ IGH in green, 3’ IGH in red). FISH analysis showed a chromosomal translocation involving MALT1 (5’ MALT1 in red, 3’ MALT1 in green) in (B) Patient 1 and (C) Patient 2. MSK-IMPACT Heme assay results from Patient 1 depicting the DNA sequencing reads corresponding to the (D) IGH breakpoint and (E) the breakpoint located upstream of MALT1 visualized using Integrative Genomics Viewer (IGV) software with reads aligned to the human genome assembly GRCh37/hg19.

Targeted next-generation sequencing (NGS) based mutational analysis was performed on the PB using the Memorial Sloan Kettering (MSK) laboratory-developed hybrid-capture-based assay (MSK-IMPACT Heme). This assay uses a 468 gene panel that includes key oncogenes and tumor suppressor genes implicated in hematolymphoid malignancies. Mutations were called based on a paired analysis using the patient normal saliva sample, and only somatic variants were reported. Sequence reads were aligned to the human genome assembly GRCh37/hg19 and manually reviewed using the Integrative Genomics Viewer (IGV). Two somatic variants were identified: SF3B1 p.R775G (variant allele frequency [VAF] of 18.6%) and PRKCB p.D427N (VAF of 10.7%). In addition, a structural variant involving IGH was noted, corresponding to the alteration t(14;18)(q32.32;q21.31), resulting in a translocation with MALT1 (Figure 2D and 2E). Taken together with the cytogenetics results, NGS confirmed the presence of an IGH::MALT1 rearrangement. The patient was stable without symptoms and per the iwCLL guidelines, there were no indications to initiate treatment.⁵ The patient is continuing to be closely monitored given neck, axillary, intra-abdominal and inguinal adenopathy.

Patient 2 was a 57-year-old male who presented for follow up after a diagnosis of CLL two years prior who had remained on active observation. Recent imaging demonstrated progressive adenopathy and labs showed rising WBC of 58.8 K/μL along with progressive cytopenias (anemia, thrombocytopenia, neutropenia). Flow cytometry performed on the PB identified an abnormal kappa restricted CD5-positive B-cell population expressing CD19, CD20-dim, CD22-dim, CD23, CD43, CD79b-dim, CD81-dim, CD200-bright, and ROR1. This population represented 92% (54 K/μL) of WBC. An in-house BM biopsy showed extensive involvement by CLL, accounting for >90% of the marrow space. Chromosome analysis of the bone marrow sample detected an abnormal karyotype with t(14;18) in 6/20 metaphase cells, including one cell also with an interstitial deletion of the long arm of chromosome 13. FISH studies were negative for CCND1::IGH fusion/t(11;14). Chromosomal translocation of IGH and MALT1 were detected in more than 73% of cells (Figure 2C). There was no evidence of TP53 deletion or loss of chromosome 17. SNP-array analysis detected a deletion of ~38 Mb in 13q14.11-q14.2 and 13q21.1-q31.1. Evaluation of IGHV SHM identified a clonal rearrangement involving IGH V3–48_J4 with 8.2% mutation frequency, compatible with a mutated status. MSK-IMPACT Heme revealed two somatic variants: MALT1 p.E43A (VAF of 41.4%) and TSHR p.I622T (VAF of 14.4%). A structural variant involving IGH and MALT1 was identified, confirming the presence of an IGH::MALT1 rearrangement. Given the progressive lymphadenopathy, worsening lymphocytosis, and cytopenias, the patient initiated first-line treatment with venetoclax and obinutuzumab.

To our knowledge, these would be the first two published cases of CLL harboring t(14;18)(q32;q21)/IGH::MALT1. Translocations involving MALT1 have been previously described in extranodal marginal zone lymphoma of the mucosa-associated lymphoid tissue (MALT) type, an indolent, primary extranodal B-cell lymphoma. Translocations described in this entity include t(11;18)(q21;q21)/BIRC3::MALT1 and t(14;18)(q32;q21)/IGH::MALT1.¹¹ The t(14;18)(q32;q21)/IGH::MALT1 translocation is rare and reported in MALT lymphoma of the ocular adnexa (7%) and lung (6%).¹² The IGH::MALT1 translocation brings the MALT1 gene under regulatory control of IGH, resulting in MALT1 overexpression. MALT1 forms a complex with CARD11 and BCL10, which activates NF-κB signaling and significantly upregulates NF-κB signaling through its protease function, promoting cell survival.¹² In addition, B-cell lymphomas, such as CLL, result in aberrant activation of the B cell receptor (BCR) pathway. Upon BCR stimulation, Bruton’s tyrosine kinase (BTK) is activated, which also regulates the NF-κB and PI3K/AKT/mTOR pathways.¹³ MALT1 hyperactivity may drive BTK inhibitor (BTKi) resistance by bypassing BTK/CARD11 upstream signaling in lymphoma patients.¹³ For patients with CLL, the role of MALT1 hyperactivity and response to covalent BTKi remains uncertain and requires additional study.

Both cases of CLL presented with a typical immunophenotype by flow cytometry with uniform expression of CD5, CD19, CD23, CD43, CD200, and ROR1. They also had dim expression of CD20, CD22, CD79b, and surface light chain restriction. In addition, Patient 1 also had a somatic SF3B1, which is one of the most frequently mutated genes in CLL.¹⁴ Notably, both patients were initially stable without adverse prognostic factors, including mutated IGHV status, wild-type TP53, and lack of cytopenia at presentation. These cases also demonstrated del(13q), which has been associated with an indolent course if present in isolation. Patient 2 ultimately developed progressive symptoms with cytopenia two years after initial diagnosis, requiring initiation of therapy.

Several studies have identified the occurrence of t(14;18)(q32;q21) in CLL. Since BCL2 and MALT1 are closely located on chromosome 18q21, differentiating between the presence of the two genes has not been well established. While cases of CLL harboring IGH translocations and partner genes including BCL2, BCL3, MYC, BCL11A^9,10,15 have been reported, MALT1 has not been previously described in the literature. Studies have shown 48–67% of CLL cases that harbor an IGH translocation have an unknown gene partner.^15,16 It is possible that several of these cases may be a result of translocations between IGH and MALT1. However, current cytogenetic testing using MALT1 break apart probes is not routinely performed in CLL and thus, evaluation for MALT1 rearrangements has not been well explored. Additionally, similar to our patients, the majority of reported CLL cases with t(14;18)(q32;q21) occur in male patients with a median age at diagnosis of 51 years, which is younger than the average age of CLL (~70 years).¹⁰ Overall, our findings suggest the need to better define such cases, which may identify a distinct subgroup of CLL.

In summary, we describe two unique cases of CLL with t(14;18)(q32;q21)/IGH::MALT1. Prognostic features of both cases include somatically mutated IGHV, deletion of 13q, and absence of TP53 deletion/alteration. Both patients were initially asymptomatic and stable at presentation, while Patient 2 required initiation of chemotherapy two years later due to progression of disease and cytopenia. Importantly, these characteristics suggest that CLL with t(14;18)/IGH::MALT1 may be associated with lower-risk biological parameters. Given the limited published literature evaluating IGH translocation status in CLL, further investigation is required to define its relative frequency. Accordingly, FISH testing for MALT1 in cases with unknown IGH translocation partners in the setting of CLL should be considered in clinical practice to better define these cases. Our findings highlight the value of future studies in a larger cohort of patients using whole genome sequencing to further characterize distinct subtypes of CLL that may improve risk stratification and targeted therapy.

Data availability statement:

The data that support the findings of this study are available from the corresponding author upon reasonable request.