MYD88 L265P Mutation in Waldenstrom's Macroglogulinemia

Abstract

1307 Background. Mutation of MYD88 gene has recently been identified in activated B-cell like diffuse B-cell lymphoma, and enhanced JAK STAT and NF-kB signalling pathways. Whole exome sequencing study in Waldenstrom macroglobulinemia (WM) suggested a high frequency of MYD88 L265P mutation in WM. Although the genetic background is not fully deciphered in WM, the role of NF-kB and JAK STAT pathways has been demonstrated in WM; which underlying mechanisms of deregulation remain to be elucidated. We aimed to analyze MYD88 mutation in exon 5 and to characterize the clinical significance of this genetic alteration in 67 WM. Method. 67 patients (42 males, 25 females) diagnosed with WM were included in this study, along with 9 patients with chronic lymphocytic leukemia (CLL), 4 multiple myeloma (MM) and 9 marginal zone lymphoma (MZL) were also studied. Patients were untreated at time of BM collection and gave informed consent prior to research sampling. Clinical features, immunophenotypic markers using flow cytometry (Matutes score panel, CD38, CD138, CD27, CD80), conventional cytogenetic, FISH and SNP array data (n = 46) were analysed. B cells from bone marrow and T cells from blood were isolated respectively using B cell isolation kit and Pan T isolation kit (Myltenyi Biotech). For DNA sequencing of exon 5 of MYD88, the exon 5 of MYD88 gene was amplified from genomic DNA by PCR. The purified PCR products were directly sequenced in both directions using BigDye® Terminator Cycle Sequencing Kit (Applied Biosystems, CA, USA) and analyzed on the Applied Biosystems 3130xl Genetic Analyzer. Data were analyzed with SeqScape software version 2.5 (Applied Biosystems). Results. MYD88 L265P mutation (MYDmut) was observed in 79% of patients, including homozygous mutation in two patients (3%). MYD88 mutation was not identified in T lymphocytes isolated from 4 WM patients that confirmed MYD88 mutation was acquired in the tumoral cells. We haven't observed any other mutation on exon 5. We then sought for other mechanisms of MYD88 gene alteration, such as copy number alteration (CNA) and copy neutral –loss of heterozygosity (CN-LOH) also considered as an acquired UPD (uniparental disomy) at MYD88 locus. We found an UPD at MYD88 locus in solely one patient (2%), and haven't identified any deletion at 3p22. On the contrary, we observed a gain on chromosome 3 at 3p22 locus (including MYD88 gene) in 7/57 (12%) patients. Taking together, we identified alteration of the MYD88 locus in 85% of patients with WM, by either gain-of-function mutation (79%) or CNA (12%). Interestingly, we found gain on chromosome 3 more frequently in the MYDwildgroup than in the MYDmutgroup (p=0.02). Twenty one percent of the patients with WM had no mutation of MYD (MYDwild), and were characterized with a female predominance, a splenomegaly, gain of chromosome 3 and CD27 expression. We did not observed difference in terms of survival according to the MYD88 mutation status. MYD88 mutation was not related to deletion 6q, gain of 4, deletion 11q, deletion 17p, deletion 13q14 in our study. Interestingly, deletion 7q, a frequent cytogenetic aberration in marginal zone lymphoma, was rare in our series (4/57; 7%) and was independent of MYD88 mutation status (2 in the MYDwild and 2 in the MYDmut) (p=ns). No MYD88 L265P mutation was observed in CLL and MM. In MZL, 1/9 patient without M monoclonal component had a MYDL265p mutation. Conclusion. These results confirm a high frequency of MYD88 L265P mutation in WM that may become a useful biomarker for diagnostic in WM and may help better understand the physiopathogeny of WM. Disclosures: No relevant conflicts of interest to declare.

Targetable activating mutations are very frequent in GCB and ABC diffuse large B‐cell lymphoma

“…The patients with MYD88 mutations were significantly older than the patients without MYD88 mutations, which is consistent with the higher frequency of these mutations in ABC DLBCL cases and the increase in the proportion of ABC DLBCL cases with age (Mareschal et al, ). If the MYD88 L265P mutation can be considered the hallmark of lymphoplasmacytic lymphoma (Treon et al, ; Gachard et al, ; Poulain et al, ), the higher frequency of MYD88 mutations in the ABC subtype that primarily expresses an IgM isotype is logical (Ruminy et al, ; Jardin et al, ). We identified somatically acquired mutations in the ITAM motifs of CD79A or CD79B preferentially in ABC and rarely in GCB.…”

The bendamustine plus rituximab regimen is active against primary nodal marginal zone B‐cell lymphoma

“…DNA was extracted from lymph node biopsies using the QIAmp kit (Sigma‐Aldrich Co, Lyon, France) according to manufacturer's recommendations, and the presence of the MYD88 L265P mutation in tumor samples from patients with an M component was assessed using a commercially available assay that combines allele‐specific amplification with amplification refractory mutation system technology and hydrolysis probe detection (Biomarker Somatic Mutation Assay, SABiosciences, Qiagen, Hilden, Germany) (quantitative polymerase chain reaction) as previously described, with slight modifications . We evaluated the sensitivity of the method by analyzing the DNA from a WM cell line containing the MYD88 L265P mutation (BCWM1) (data not shown) …”

MYD88 L265P and CXCR4 mutations in lymphoplasmacytic lymphoma identify cases with high disease activity

“…Activated MYD88 recruits IL1 receptor‐associated kinase 1 (IRAK1) and IRAK4, as well as Bruton tyrosine kinase (BTK) (Yang et al , ), ultimately leading to activation of NF‐κB (NFKB1). Inhibition of the MYD88 signalling pathway blocks the translocation of NF‐κB into the nucleus in cells expressing MYD88 L265P, and it was shown that WM cells depend on NF‐κB signalling for growth and survival (Leleu et al , ; Treon et al , ; Poulain et al , ).…”