Improved Detection of the KIT D816V Mutation in Patients with Systemic Mastocytosis Using a Quantitative and Highly Sensitive Real-Time qPCR Assay

摘要

The vast majority of patients with systemic mastocytosis (SM)carry the somatic D816V mutation in the KIT gene. TheKIT D816V mutation is one of the minor criteria for a diagnosisof SM according to the 2008 World Health Organization classification of myeloproliferativeneoplasms. In the present study, we present a real-time qPCR assay that allowsquantification of as little as 0.003% KIT D816V mutation-positivecells. A total of 61 samples from 31 cases of SM were included in the study. We detectedthe mutation in skin or bone marrow in 95% of the cases of SM. We demonstrate the clinicalrelevance of the assay by identifying as little as 0.03% mutation-positive cells in bonemarrow aspirates from SM patients and calculate the analytical sensitivity of negativesamples to determine the reliability of the result. We further demonstrate that thismethod also detects the KIT D816V mutation in peripheral blood in81% of the mutation-positive cases with SM. The method also allows comparison ofmutation-positive and mast cell fractions to determine whether the mutation is present innon-mast cells, a parameter that has recently been reported to be of prognostic importancein patients with indolent SM. Finally, the assay is suitable for use in prospectivestudies of the KIT D816V allele burden as a treatment endpoint inSM.