Abstract A033: Ex vivo Cbl-b silenced, autologous TIL therapy in glioblastoma demonstrates proof-of-concept efficacy and translational potential

Abstract Purpose: Glioblastoma (GBM) remains largely refractory to immunotherapy due to extensive intratumoral immunosuppression and functional exhaustion of tumor-infiltrating lymphocytes (TILs). This study evaluated whether ex vivo silencing of the intracellular immune checkpoint Cbl-b could enhance the antitumor efficacy of autologous TIL therapy in GBM. Experimental Design: Tumor-reactive lymphocytes were isolated from human glioblastoma specimens and expanded using a closed, feeder-free xpsTIL manufacturing process. Transient Cbl-b silencing was achieved by electroporation with the clinically validated EPiC platform. T-cell phenotype, cytokine secretion, cytotoxic activity, and T-cell receptor (TCR) repertoire diversity were assessed in vitro. Therapeutic efficacy was evaluated in immunocompetent subcutaneous and orthotopic glioma mouse models, including studies in combination with temozolomide. Results: CD8+ TILs isolated from glioblastoma exhibited high Cbl-b expression, implicating intrinsic suppression of effector function. xpsTIL expansion reproducibly achieved 104-fold proliferation while preserving a highly polyclonal, tumor-exclusive TCR repertoire. EPiC-mediated electroporation resulted in 95% downmodulation of Cbl-b expression, leading to enhanced IL-2 and IFN-γ production and increased anti-tumor cytotoxicity. In orthotopic glioma models, adoptive transfer of Cbl-b–deficient TILs significantly improved tumor control, achieving cure rates of approximately 20% as monotherapy and up to 70% when combined with temozolomide (0% cure with temozolomide alone). Treated animals demonstrated modest protection upon intracranial tumor rechallenge. Conclusions: Ex vivo Cbl-b silencing reprograms glioblastoma-resident TILs to overcome tumor-intrinsic and therapy-induced immunosuppression, enabling durable antitumor immunity without systemic checkpoint blockade. Cbl-b–silenced xpsTILs represent a promising, clinically scalable cell therapy approach for glioblastoma. A large language model (ChatGPT) was used to assist in structuring and refining the abstract text; all scientific content was generated, reviewed, and validated by the authors. Citation Format: Prafulla C. Gokhale, David A. Reardon, Friedrich Erhart, Georg Widhalm, Benjamin K. Eschle, Brittaney Leeper, Maia Lineberry, Romana Gugenberger, Alexander Dohnal, Mario Kuttke. Ex vivo Cbl-b silenced, autologous TIL therapy in glioblastoma demonstrates proof-of-concept efficacy and translational potential abstract. In: Proceedings of the AACR Special Conference in Cancer Research: Brain Cancer; 2026 Mar 23-25; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2026;86 (6Suppl): Abstract nr A033.