Determination of MYD88L265P mutation fraction in IgM monoclonal gammopathies.

We here describe a novel method for the detection of MYD88L265P mutation using a competitive allele specific PCR (Cast-PCR) assay. This assay has a sensitivity of 1x10-3, is applicable in reactions containing very low amounts of DNA (as low as 20 pg) and allowed the detection of MYD88L265P somatic mutation in both tumor derived DNA (tDNA) and cell free DNA (cfDNA). In addition, using Cast-PCR assay we were able to determine the mutation allele fraction (MAF) in each tested sample. We then analyzed baseline tDNA and cfDNA samples from 163 patients (53 with IgM-MGUS and 110 with WM, of which 54 were asymptomatic and 56 symptomatic) and also in sequential samples of 37 patients. MAF in both cfDNA and tDNA was higher among patients with symptomatic compared to asymptomatic WM and in those with asymptomatic WM compared to IgM-MGUS patients. In addition, the evaluation of sequential samples showed that MAF decreased after treatment while increased in patients who relapsed or progressed to symptomatic WM. Thus, Cast-PCR is a highly-sensitive, cost-effective diagnostic tool for MYD88L265P detection, applicable in both tumor and cell free DNA samples which also provides a quantitative evaluation of the tumor load in patients with IgM monoclonal gammopathies.