Abstract 237: Coincidence of InDels with missense mutations: 0, 1 or 2 artifacts

Abstract

Abstract The purpose of this study was to explain a serendipitously discovered next generation sequencing (NGS) artifact. While reviewing clinical cases, we found a case of a large deletion that also contained multiple missense mutations. This phenotype was unknown to several molecular pathologists in our group. We reviewed 46 clinical cases with short insertions or deletions (InDels) (1-30 bases) or FLT3 internal tandem duplications (ITDs) (6-183 bases) of solid or hematologic malignancy processed with a clinical NGS assay and identified misaligned reads, ranging from 3 to 100% of reads showing mismapped bases. Reads that straddled the InDel with sufficient numbers of bases on both sides were correctly anchored both upstream and downstream of the InDel, and were called correctly. However, reads that ended in the middle of the InDel were incorrectly mapped onto the deletion, thereby producing artifactual missense mutations. The frequency of mismapped bases increased with InDel size, and the VAF of the InDel can be markedly underestimated. The ratio of forward to reverse reads was commonly skewed for the mismapped base compared to the ratio for the wildtype base at that position. ABRA2 was able to correct 41 to 100% of the reads with mismapped bases and led to absolute increases in the VAF from 1 to 61% along with correction of all of the SBSs except for two cases. Knowledge of the various artifacts commonly seen in next generation sequencing is essential to safely sign out molecular pathology cases. Citation Format: Kelly E. Craven, Catherine G. Fischer, LiQun Jiang, Aparna Pallavajjala, Ming-Tseh Lin, James R. Eshleman. Coincidence of InDels with missense mutations: 0, 1 or 2 artifacts [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 237.