Comparative analysis of next-generation sequencing and immunohistochemistry in MSI/MMR testing

Abstract

Objective: Loss of mismatch repair (MMR) protein expression, assessed via immunohistochemistry (IHC), and microsatellite instability (MSI) status, determined through molecular methods, are two tumor-agnostic predictive biomarkers for immunotherapy eligibility. However, there remains no consensus on the preferred testing method, nor on the type and extent of molecular testing required for optimal patient selection. This study investigates the correlation between MMR protein loss detected by IHC and MSI status identified through next-generation sequencing (NGS) to evaluate the concordance and potential complementary roles of these methods. Material and Methods: A total of 139 tumor samples were analyzed for MSI using NGS. The cohort included colorectal carcinoma (n=51), pancreatic ductal adenocarcinoma (n=22), cholangiocarcinoma (n=9), non-small cell lung carcinoma (n=6), adenoid cystic carcinoma (n=6), gastric adenocarcinoma (n=6), high-grade serous ovarian carcinoma (n=5), and 34 other tumor types. IHC was performed to assess MLH1, MSH2, MSH6, and PMS2 protein expression. The correlation between MSI status and MMR protein loss was evaluated.

Results: Twelve tumors (8.6%) were classified as MSI-High (microsatellite instable). Among them, ten exhibited MMR protein loss, whereas two MSI-High tumors (a mucinous adenocarcinoma of omental origin and a mucinous colon adenocarcinoma) retained MMR protein expression. No MMR-deficient tumors were identified as MSI-Low (microsatellite stable/MSS).

Conclusion: A strong correlation exists between IHC-based MMR loss and NGS-based MSI detection. IHC remains widely used due to its accessibility and cost-effectiveness, whereas NGS offers higher accuracy and broader genomic insights. With its ability to detect multiple alterations simultaneously, NGS is particularly valuable when tissue is scarce. Combining both methods can improve diagnostic accuracy and guide optimal immunotherapy selection.