A Highly Verified Assay for KRAS Mutation Detection in Tissue and Plasma of Lung, Colorectal, and Pancreatic Cancer

Abstract

Context.— KRAS Mutation Test v2 is used for the qualitative detection and identification of 28 mutations in exons 2, 3, and 4 of the human KRAS gene. Objective.— To verify the performance of KRAS Mutation Test v2 and to evaluate its accuracy by correlation with a next-generation sequencing method on Illumina MiSeq. Design.— In this study, we used formalin-fixed, paraffin-embedded tissue and plasma specimens from non–small cell lung cancer, colorectal cancer, and pancreatic cancer patients. Results of specificity, precision, analytical sensitivity, and accuracy as compared with a MiSeq method are reported. Results.— The KRAS Mutation Test v2 demonstrated exquisite sensitivity and specificity and broad coverage of KRAS mutations. Precision was 100% (108 of 108) across all samples, operators, and instruments for formalin-fixed, paraffin-embedded tissue and 99.8% (615 of 616) for plasma. Analytical sensitivity was high with detection of 1% mutant sequence in formalin-fixed, paraffin-embedded tissue samples and as low as 25 mutant sequence copies/mL for plasma samples. The test also showed high overall concordance for formalin-fixed, paraffin-embedded tumor tissue as well as for plasma specimens when compared with MiSeq sequencing results. Conclusions.— The KRAS Mutation Test v2 is a highly robust, reproducible, and sensitive test for the qualitative detection of 28 mutations in exons 2, 3, and 4 of the KRAS gene in both solid (tissue) and liquid (plasma) biopsies from colorectal cancer, non–small cell lung cancer, and pancreatic cancer, and is a convenient option for KRAS mutation testing.