Detection of Mismatch Repair Deficiency and Microsatellite Instability in Colorectal Adenocarcinoma by Targeted Next-Generation Sequencing

• Published in: The Journal of molecular diagnostics : JMD Vol. 19; no. 1; pp. 84 - 91
• Main Authors: Nowak, Jonathan A., Yurgelun, Matthew B., Bruce, Jacqueline L., Rojas-Rudilla, Vanesa, Hall, Dimity L., Shivdasani, Priyanka, Garcia, Elizabeth P., Agoston, Agoston T., Srivastava, Amitabh, Ogino, Shuji, Kuo, Frank C., Lindeman, Neal I., Dong, Fei
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.01.2017; American Society for Investigative Pathology
• Subjects: Adenocarcinoma - diagnosis; Adenocarcinoma - genetics; Base Sequence; Colorectal Neoplasms, Hereditary Nonpolyposis - diagnosis; Colorectal Neoplasms, Hereditary Nonpolyposis - genetics; DNA Mismatch Repair; DNA Mutational Analysis; Genes, Neoplasm; High-Throughput Nucleotide Sequencing; Humans; Microsatellite Instability; Microsatellite Repeats; Mutation; Pathology; Regular
• ISSN: 1525-1578; 1943-7811
• DOI: 10.1016/j.jmoldx.2016.07.010

Mismatch repair protein deficiency (MMR-D) and high microsatellite instability (MSI-H) are features of Lynch syndrome–associated colorectal carcinomas and have implications in clinical management. We evaluate the ability of a targeted next-generation sequencing panel to detect MMR-D and MSI-H based on mutational phenotype. Using a criterion of >40 total mutations per megabase or >5 single-base insertion or deletion mutations in repeats per megabase, sequencing achieves 92% sensitivity and 100% specificity for MMR-D by immunohistochemistry in a training cohort of 149 colorectal carcinomas and 91% sensitivity and 98% specificity for MMR-D in a validation cohort of 94 additional colorectal carcinomas. False-negative samples are attributable to tumor heterogeneity, and next-generation sequencing results are concordant with analysis of microsatellite loci by PCR. In a subset of 95 carcinomas with microsatellite analysis, sequencing achieves 100% sensitivity and 99% specificity for MSI-H in the combined training and validation set. False-positive results for MMR-D and MSI-H are attributable to ultramutated cancers with POLE mutations, which are confirmed by direct sequencing of the POLE gene and are detected by mutational signature analysis. These findings provide a framework for a targeted tumor sequencing panel to accurately detect MMR-D and MSI-H in colorectal carcinomas.