Clinicopathological features and mutational landscape of colorectal cancer subgroups defined by MSI and EMAST status

Colorectal cancer (CRC) remains a leading cause of cancer-related morbidity and mortality. Microsatellite instability-high (MSI-H) tumors, resulting from defective DNA mismatch repair (MMR), represent a well-defined subtype with distinctive biological behavior and immunogenicity. In contrast, tumors with elevated microsatellite alterations at tetranucleotide repeats (EMAST) are less well characterized. EMAST can manifest with MSI or arise as an isolated form of instability, delineating discrete phenotypes underpinned by distinct mechanisms. This study aimed to characterize MSI and EMAST status in CRCs. By integrating instability profiles with clinicopathological features and mutational profiles of key driver genes, we aimed to refine molecular classification and advance understanding of CRC tumorigenesis. A total of 332 CRCs were analyzed for MSI and EMAST using established panels. Clinicopathological characteristics were recorded, and mutational profiling of KRAS, BRAF, CTNNB1, PIK3CA, and TP53 was performed. MLH1 expression was assessed using immunohistochemistry. MSS/EMAST-S tumors displayed profiles typical of chromosomally stable CRC, dominated by KRAS and followed by TP53 and PIK3CA mutations. MSI-H/EMAST-H tumors were characterized by frequent BRAF mutations, right-sided location, female predominance, and lower TP53 mutation rate, consistent with the classical hypermutated, immunogenic subtype. In contrast, MSS/EMAST-H tumors exhibited unique features, including enrichment for PIK3CA and CTNNB1 mutations, larger tumor size, and poorer differentiation, suggesting an intermediate phenotype between MSS and MSI-H. MSS/EMAST-L tumors aligned with chromosomally stable, KRAS/Wnt-driven CRC. In conclusion, MSS/EMAST-H tumors represent an underrecognized CRC subtype with intermediate genomic instability and a distinctive molecular profile, with potential implications for prognostic assessment and personalized therapeutic strategies.