Analysis of DNA mismatch repair and microsatellite instability in molecular typing of endometrial carcinoma

DNA mismatch repair (MMR)/microsatellite instability (MSI) status is an indispensable biomarker for predicting immunotherapy response, screening for Lynch syndrome (LS), and molecularly classifying endometrial carcinoma (EC). To investigate the consistency and the causes of discordance among different methods for MMR/MSI, we performed MMR immunohistochemistry (MMR-IHC) and next-generation sequencing (NGS) for MSI status (MSI-NGS). Formalin-fixed and paraffin-embedded (FFPE) specimens were collected from 141 EC patients. 38 (26.95%) patients exhibited deficient MMR (dMMR), while 103 (73.05%) harboured proficient MMR (pMMR). There were no significant differences in ages or histological types between the dMMR and pMMR groups (all P > 0.05). Molecular classification based on NGS results revealed no significant differences in ages among the POLE-mutated (POLE mut), MSI-high (MSI-H), p53-abnormal (p53 abn), and no specific molecular profile (NSMP) subtypes (all P > 0.05). The proportion of aggressive histological subtypes in the MSI-H group (44.0%, 11/25) was higher than that in the NSMP group (17.4%, 15/86; P < 0.01); lower than that in the p53 abn group (80.0%, 12/15; P < 0.05); but there was no difference compared to the POLE mut group (33.3%, 5/15; P > 0.05). MSI-NGS classified 114 (80.85%) cases as microsatellite stable (MSS) phenotype and 27 (19.15%) as MSI-H phenotype. Compared to MMR-IHC, discordance was observed in 11 cases between the two methodologies, and the consistency rate was 92.2% (130/141). Among the inconsistent 11 cases, PCR-capillary electrophoresis (PCR-CE) detection showed 2 MSI-H cases, 3 MSI-low (MSI-L), and 6 MSS. Among them, 6 cases with concurrent loss of MLH1 and PMS2 exhibited MLH1 promoter hypermethylation and lacked MMR gene mutations. Among the 38 dMMR patients, 23.7% (9/38) were diagnosed as LS with the germline mutations in MMR genes, and all of their MSI-NGS results were MSI-H. Two dedifferentiated EC (DEC) cases were detected with MLH1 promoter hypermethylation, accompanied by concurrent loss of MLH1 and PMS2, with tumor mutational burden-high (TMB-H) and no MMR gene mutations. Among the 7 (4.96%) patients with isolated MSH6 deficiency, MSI-NGS analysis revealed 3 MSS cases and 4 MSI-H, while PCR-CE detection revealed 5 MSI-H cases. Among the 3 (2.13%) patients with isolated PMS2 deficiency, MSI-NGS identified 2 MSS cases and 1 MSI-H, which were consistent with the PCR-CE results. In conclusion, MMR-IHC reveals relatively high concordance with MSI-NGS. Discordance primarily occurs in the cases with MLH1 promoter hypermethylation and isolated PMS2 or MSH6 deficiency. When necessary, comprehensive analyses should be conducted combining MLH1 gene methylation, PCR-CE testing, MMR gene mutations, and TMB status, so as to provide precise molecular classification for EC patients.