Abstract 6571: Proliferative T cell infiltration is associated with colorectal cancer molecular subtypes and cancer specific survival

Abstract The immune microenvironment is a critical regulator of colorectal cancer (CRC) biology and represents a clinically targetable tumor feature in a subset of patients. T cells are one of the most common cell types in the CRC immune microenvironment and exist in a broad variety of subtypes and activation states. Proliferation status represents a general measure of T cell activity that is applicable to many subtypes. Evidence from other tumor types suggests that proliferating cytotoxic T cells may mediate adaptive immunity in cancer. However, T cell proliferation in CRC is incompletely characterized. To investigate the role of proliferating T cells in CRC, we developed a multiplexed immunofluorescence assay assessing CD3, CD4, CD8, FOXP3, PTPRC (CD45RA and CD45RO), MKI67 (Ki67), and KRT (cytokeratin) expression, and assessed 1,610 primary resected CRC specimens from four US/Canadian cohort studies. Digital imaging and pathologist-supervised machine learning were used to decompose image data into single-cell level information. The Kaplan-Meier method was used to describe CRC-specific survival (CSS) and multivariable Cox proportional hazards models were used to examine associations of proliferating and non-proliferating T cell densities with CSS. Multivariable ordinal logistic regression was used to adjust for confounding factors (age, cancer site, sex, and study batch) in analyses of T cell densities and tumor features. Across one million T cells, 4.5 % were classified as proliferating, as defined by positive MKI67 expression. While higher densities of both proliferating and non-proliferating T cells were positively associated with microsatellite instability (MSI), a CpG island methylator high phenotype, BRAF mutant status and KRAS wildtype status, these associations were stronger for proliferating T cells (e.g., adjusted odds ratio (OR) 2.09, CI 1.62-2.70, p<0.0001 for MSI and higher proliferative T cell density versus adjusted OR 1.40, CI 1.08-1.81, p=0.01 for MSI and higher non-proliferative T cell density). Higher proliferative and non-proliferative T cell densities were both positively associated with CSS (adjusted hazard ratio (HR) 0.75, CI 0.55-1.03, comparing highest versus lowest quartile of proliferative T cell densities (Ptrend=0.018) and adjusted HR 0.42, CI 0.31-0.57, comparing highest versus lowest quartile of non-proliferative T cell densities (Ptrend<0.001). Despite comprising a minor subset of total T cell infiltration in the CRC microenvironment, proliferative T cells are more strongly associated with specific molecular subtypes and are also associated with CSS. These results, across a large series of primary human tumors, suggest that proliferation marks a subset of functionally distinct subset of T cells in the CRC microenvironment. Citation Format: Yasutoshi Takashima, Claire Elizabeth Thomas, Andressa Dias Costa, Sushma Thomas, Evertine Wesselink, Conghui Qu, Steven Gallinger, Robert C. Grant, Li Hsu, Marios Giannakis, Jeroen Huyghe, Daniel D. Buchanan, Shuji Ogino, Riki (Ulrike) Peters, Amanda I. Phipps, Jonathan A. Nowak. Proliferative T cell infiltration is associated with colorectal cancer molecular subtypes and cancer specific survival [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 6571.