Abstract
While microsatellite-instable (MSI) colorectal cancers (CRC), reflecting mismatch repair deficiency, often respond to immune checkpoint inhibitors, microsatellite-stable (MSS) tumors remain largely resistant. This disparity is typically attributed to differences in neoantigen load. However, whether antigen-independent mechanisms contribute to immune evasion in MSS-CRC remains unclear. To address this, we engineered a model in which MSI- and MSS-CRC cells express identical levels of a defined antigen recognized by TCR-engineered T cells. Despite equivalent antigen presentation, MSS tumors exhibited impaired T-cell activation, reduced cytotoxicity, and resistance to killing. We linked this immune evasion to the MSS tumor secretome, which suppressed immune responses even in immunogenic MSI cells by impairing immune synapse formation. Surfaceome profiling by mass spectrometry identified glycosylation-dependent alterations that impair immune recognition. Our findings demonstrate that MSS-CRC evades immune attack via intrinsic secretome-driven mechanisms, independent of antigenicity. Targeting glycosylation-linked suppressive pathways may restore T-cell responsiveness and improve immunotherapy efficacy in MSS-CRC.