Extended tumor area-based stratification score combining tumor budding and stroma identifies a high-risk, immune-depleted group in localized microsatellite-stable colon cancer patients

Microsatellite-stable colon cancer represents a heterogeneous group of tumors, where the identification of highrisk histopathological factors is particularly critical. In this study, we evaluate two morphological features associated with mesenchymal differentiation-tumor budding and tumor-associated stroma-with the aim of developing a robust recurrence risk stratification score and exploring its relationship with the tumor microenvironment composition in this clinical context. We retrospectively analyzed 254 formalin-fixed, paraffinembedded colectomy specimens from patients with mismatch repair-proficient colon cancer (stages I to III). Tumor budding and tumor-associated stroma were assessed using two protocols: one focused on a single hotspot field and another on a broader tumor area. The tumor microenvironment composition, including the presence of tertiary lymphoid structures, was characterized using immunohistochemistry. We developed a three-tiered tumor budding-stroma (TBS) stratification score based on the evaluation of an extended tumor area. This score was independently associated with the disease-free survival probability (low-TBS: HR 0.12, 95 % CI 0.04-0.33, p < 0.001; intermediate-TBS: HR 0.26, 95 % CI 0.10-0.65, p = 0.003) and allowed the identification of a highrisk group characterized by an immune-depleted tumor microenvironment. The prognostic value of this approach outperformed that of each individual parameter and was superior to the stratification score obtained using the hotspot field-based assessment. In conclusion, the combined assessment of tumor budding and tumor-associated stroma over an extended tumor area provides a more comprehensive view of tumor heterogeneity. This approach may be suitable for routine evaluation of microsatellite-stable localized colon cancer patients.