Detection of the MYD88 L265P mutation in Waldenström’s macroglobulinemia using a highly sensitive allele-specific PCR assay.

Abstract

8042 Background: Waldenström’s macroglobulinemia (WM) is a B-cell malignancy characterized by bone marrow (BM) infiltration with lymphoplasmacytic cells and production of an IgM paraprotein. By whole genome sequencing, we recently identified a somatic mutation (L265P) in the MYD88 gene in 27/30 (90%) WM patients (Treon et al, ASH 2011). To expand this finding for possible diagnostic testing, we developed an allele-specific PCR assay for MYD88 L265P and evaluated this assay in a large cohort of WM patients. Methods: An allele-specific PCR assay was developed with a threshold of detection of 0.125% for MYD88 L265P. DNA from bone marrow aspirates from 99 patients with the clinicopathological diagnosis of WM was used for assessment of MYD88 L265P expression by both allele-specific PCR and Sanger sequencing. Findings were correlated with clinical parameters using ANOVA. Results: We observed that 85/99 (86%) WM patients were positive for MYD88 L265P using the allele-specific PCR assay. Of the 85 allele-specific PCR positive patients, 81 demonstrated a detectable mutation peak by Sanger sequencing. All 14 allele-specific PCR negative patients remained negative by Sanger sequencing. By the allele-specific PCR assay, MYD88 L265P positive patients showed greater bone marrow involvement, higher serum IgM and lower serum IgA and IgG levels versus MYD88 L265P negative patients (p<0.008). Conclusions: MYD88 L265P is highly expressed in BM samples of WM patients using an allele-specific PCR assay, and is associated with greater bone marrow disease burden and serum IgM levels. Use of allele-specific PCR provides a simple and sensitive diagnostic tool for detection of the MYD88 L265P mutation.