Clinical testing for mismatch repair (MMR) deficiency often entails serial testing of tumour and constitutional DNA using multiple assays. To minimize cost and specimen requirements of MMR testing, we developed an integrated targeted sequencing protocol (termed MultiMMR) that tests for promoter methylation, mutations, copy number alterations, copy neutral loss-of-heterozygosity, and microsatellite instability from a single aliquot of DNA. We performed hybrid capture of DNA sequencing libraries constructed with methylated adapters on 142 samples (60 tumours and 82 constitutional samples) from 82 patients with MMR-associated colorectal, endometrial, and brain cancers as well as a synthetic DNA mix with 11 known mutations. We then split the captured material to enable parallel bisulfite and conventional sequence analysis. The panel targeted microsatellite regions and 13 genes associated with MMR, hypermutation, and hereditary colorectal cancer. MultiMMR recapitulated clinical testing results in 23/24 cases, was able to explain MMR loss in an additional 29/48 patients with incomplete/inconclusive testing, and identified all 11 MMR variants within the synthetic DNA mix. Promoter methylation and microsatellite instability analysis showed 95% and 97% concordance with clinical testing, respectively. We demonstrated feasibility for amalgamation of the current stepwise and complex clinical testing workflow into an integrated test for both hereditary and somatic causes of MMR deficiency.
Copyright © 2020. Published by Elsevier Inc.