Clinicopathological, Molecular, and Immune Profiling in Colorectal Cancer: Associations Between KRAS/BRAF Alterations, dMMR/MSI-H Status, and Tumor-Infiltrating Lymphocytes: A Retrospective Cross-Sectional Study.

Background: To characterize the molecular and immune profiles of resected colorectal cancer (CRC), including KRAS/NRAS/BRAF mutations, deficient mismatch repair/high microsatellite instability (dMMR/MSI-H), and tumor-infiltrating lymphocyte (TIL) positivity, using routinely available pathological methods, and to identify independent clinicopathological and molecular factors associated with TIL positivity and dMMR/MSI-H status. Methods: This retrospective cross-sectional study included 272 patients with primary colorectal adenocarcinoma who underwent surgical resection between 2018 and 2024. KRAS, NRAS, and BRAF mutations were analyzed by real-time polymerase chain reaction. Mismatch repair protein expression was evaluated by immunohistochemistry (IHC), and tumors with loss of at least one mismatch repair protein were classified as dMMR using IHC as a surrogate method for MSI-H status. TIL were assessed on hematoxylin and eosin (H&E)-stained sections. Results: KRAS mutations were detected in 40.8% of tumors, while BRAF and NRAS mutations were identified in 4.0% and 2.9% of cases, respectively. dMMR/MSI-H was present in 9.6% of tumors. Positive TIL status was observed in 35.3% of cases. Univariable analysis showed that dMMR/MSI-H status and fungating tumor morphology were significantly associated with higher TIL positivity, although these associations became borderline significant after multivariable adjustment. Independent factors associated with dMMR/MSI-H status included younger age (adjusted OR = 0.29, 95% CI: 0.12-0.69), proximal tumor location (adjusted OR = 0.25, 95% CI: 0.10-0.64), and poor tumor differentiation (adjusted OR = 2.80, 95% CI: 1.16-6.76). Subgroup analysis also showed that dMMR-related groups were more likely to have tumors in the proximal colon, poor differentiation, younger age, and higher lymphocytic infiltration. Conclusions: Integrated clinicopathological and molecular analysis identified specific CRC features associated with dMMR/MSI-H status, including proximal tumor location, poor differentiation, younger patient age, and a trend toward increased lymphocytic infiltration. Borderline associations were also observed between TIL positivity, fungating tumor morphology, and dMMR/MSI-H phenotypes. These findings support the feasibility of combined molecular and immune evaluation using routinely available pathological methods, particularly in resource-limited settings.