Dual testing with immunohistochemistry and PCR for enhanced accuracy in dMMR/MSI detection in Greek colorectal cancer population: the impact of retesting.

Abstract

Background: Deficient mismatch repair (dMMR) and microsatellite instability (MSI) are pivotal biomarkers in colorectal cancer (CRC). This study aimed to evaluate their prevalence in a Greek CRC cohort and assess the correlation of immunohistochemistry (IHC) with polymerase chain reaction (PCR) analyses, focusing on the role of retesting to resolve discrepancies.

Methods: Formalin-fixed, paraffin-embedded tumor samples from 96 patients were tested for four MMR proteins (MLH1, MSH2, MSH6, PMS2) via IHC and five mononucleotide markers (BAT25, BAT26, NR21, NR24, NR27) via PCR. Discordant cases (pMMR/MSI or dMMR/MSS) underwent retesting to refine classification.

Results: Thirteen tumors (13.54%) were dMMR/MSI. Before retesting, the correlation coefficient (r) between IHC and PCR was 0.883, with four conflicting cases. Retesting resolved three inconsistencies, improving the correlation to 0.958 and leaving a single persistent discrepancy (pMMR/MSI). PMS2 deficiency, either alone or co-deleted with MLH1, predominated among dMMR tumors. Dual testing of 5 MSI markers and 4 MMR proteins provided diagnostic clarity.

Conclusion: In this retrospective, single-center cohort, dual assessment of MMR status by IHC and PCR was associated with near-perfect inter-method agreement and reduced the likelihood of technical misclassification. These findings support the practical value of reflex retesting as a quality-assurance step within a workflow-oriented diagnostic strategy.