Abstract

The efficacy of immune checkpoint blockade (ICB) therapy in colorectal cancer (CRC) remains limited. Thus, elucidating the molecular mechanisms underlying tumor immune evasion and identifying novel predictive biomarkers are critical for improving immunotherapy outcomes in CRC patients. Here, we report that ubiquitin-specific peptidase 21 (USP21) is overexpressed in tumor tissues from CRC patients who fail to respond to immunotherapy and that its expression is correlated with CD8 + T-cell exclusion and impaired antitumor immunity. To functionally validate these clinical observations, we employed syngeneic mouse models of both microsatellite instability-high (MSI-H) and microsatellite stable (MSS) CRC. Genetic ablation of Usp21 or treatment with the USP21 inhibitor BAY-805 significantly enhanced tumor-reactive CD8 + T-cell responses, suppressed tumor progression, and synergized with anti-PD-1 therapy. Mechanistically, USP21 stabilized β-catenin by removing K48-linked ubiquitin chains, enabling its nuclear translocation and binding to the ATF3 promoter to upregulate ATF3 expression. ATF3 subsequently mediated transcriptional repression of the chemokine CCL4, thereby limiting CD8⁺ T-cell trafficking and function via the CCL4-CCR5 axis. Importantly, the therapeutic potential of targeting USP21 to enhance anti-PD-1 efficacy was further validated in huCD34 + humanized mice. Collectively, our findings identify USP21 as a pivotal regulator of immune evasion, and inhibiting USP21 represents a promising adjuvant strategy to increase the efficacy of ICB in CRC. The alternative text for this image may have been generated using AI.