A novel next generation sequencing strategy for detecting microsatellite instability in colorectal and gastric cancer

Background: Microsatellite instability (MSI) has become a critical predictive and prognostic biomarker in multiple cancers. The mainstream MSI-detecting techniques, immunohistochemistry (IHC) and polymerase chain reaction (PCR), may have trouble profiling an expanded panel if tissue samples are not available in adequate quantity, depriving patients of treatment options, especially for those with advanced disease. Next generation sequencing, in contrast, allows for massively parallel examination of numerous targets in one single test. In this work, we developed a novel NGS-based assay to detect MSI in colorectal and gastric cancer (CRC/GC) and clinically validated its performance in comparison with IHC and PCR. Methods: Surgical or biopsy specimens collected from 104 CRC and 15 GC patients were examined for MSI status using IHC, PCR and NGS employing a novel algorithm. The concordance rates between the three techniques were assessed using z-test. Results: Of the 119 patients enrolled, 53.8% had metastases. IHC tests identified deficient MMR protein expression in 40 (33.6%) cases (dMMR), seven of which were microsatellite stable (MSS) using both PCR and NGS. A discrepancy was also observed for one MMR-proficient (pMMR) case where NGS and IHC consistently produced negative results (pMMR/MSS), while PCR reported a MSI-high (MSI-H) status. The NGS method demonstrated 97.1% (33/34) sensitivity and 100% (85/85) specificity compared to PCR,and 82.5% (33/40) sensitivity and 100% (79/79) specificity compared to IHC. The concordance rate was 99.1% between NGS and PCR, and 94.1% between NGS and IHC. Of the 33 dMMR/MSI-H patients, 30.3% harbored germline defects, suggestive of Lynch Syndrome. In addition, in line with previous reports, the dMMR/MSI-H subset showed a significantly higher median tumor mutational burden (TMB) than the pMMR/MSS patients (P=0.0039). Conclusion: Our NGS-based approach is reliable and robust for MSI status determination in CRC/GC. Citation Format: Wenyun Li, Fengmei Pi, Xiaoxia Liang, Taiyuan Cao, Yiheng Lin, Xiaohui Zhai, Mengli Huang, Hao Qin, Chan Gao, Shangli Cai, Xianrui Wu, Jian Xiao, Ping Lan. A novel next generation sequencing strategy for detecting microsatellite instability in colorectal and gastric cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3147.