Cblb Gene Editing in T Cells Sustains Expansion and Immunogenic CAR T Tumor Killing Under Chronic Antigenic Stimulation.

CBL-B is an intracellular E3 ubiquitin ligase that acts as a T cell checkpoint by raising activation thresholds and limiting effector function. Here, genetic targeting of CBL-B enhances the performance of adoptively transferred T cells and CAR T cells under tumor microenvironment-like stress. In fully immunocompetent mouse models, Cblb deficiency or transient Cblb silencing improves control of MC-38 colon carcinoma and autochthonous mammary tumors, demonstrating that CBL-B restrains anti-tumor immunity. Cblb-deficient T cells show enhanced expansion and effector/effector-memory differentiation during an in vivo mixed lymphocyte reaction, confirming a cell-intrinsic brake function of CBL-B during sustained antigenic challenge. In a syngeneic Panc02-EpCAM model, Cblb-deficient anti-EpCAM CAR T cells show superior tumor control, enhanced infiltration, prolonged survival, and preserved effector function despite chronic antigen exposure and TGF-β. Mechanistically, Cblb targeting maintains granzyme B and IFN-γ production and is associated in vitro with increased GSDME-linked pyroptotic tumor cell death, consistent with features of immunogenic cell death. These findings extend previous CBL-B CAR T work from lymphocyte-deficient to immunocompetent settings and support CBL-B inhibition as a strategy to engineer CAR T cells that resist suppressive tumor microenvironments while promoting a more inflammatory mode of tumor killing.