Abstract 128: Synthetic reprogramming of γδ CAR T cells with the LTBR gene optimizes function in the TME and overcomes effector limitations.

Abstract Cell therapies have transformed outcomes in hematologic cancers, yet their efficacy in solid tumors remains limited by the profoundly immunosuppressive tumor microenvironment (TME), which employs multiple, overlapping mechanisms to suppress immune activity. Current armoring strategies address these barriers individually, leaving key vulnerabilities, while autologous approaches remain costly and difficult to scale. Allogeneic γδ CAR T cells represent an attractive, off-the-shelf platform, but their persistence and potency within the solid tumor TME require enhancement. To advance scalable and effective solid tumor therapies, we pursued a data-driven strategy to optimize γδ T cell biology while increasing TME resilience. Owing to limited knowledge of γδ T cell functional states in antitumor response, we implemented an empirical approach integrating genome-scale discovery and AI-driven analytics to uncover pathways governing γδ CAR T function. We conducted a genome-wide overexpression screen to identify reprogramming genes that protect CAR T cells from adenosine, TGF-β, immunosuppressive macrophages, and regulatory T cells while improving persistence. Among 12,000 genes screened, 51 candidates conferred protection against three or more suppressive mechanisms. Lymphotoxin beta receptor (LTβR)—a TNF superfamily receptor not normally expressed in T cells—emerged as the top hit, reprogramming T cells into highly proliferative, persistent, and multifunctional states, as confirmed by bulk and single-cell RNA-seq. In high-stress assays, LTβR expression conferred broad resistance to immunosuppression, including complete abrogation of Treg-mediated inhibition. In γδ CAR T cells, LTβR induced an αβ-like effector program characterized by enhanced cytotoxicity, persistence, and cytokine modulation. scRNA-seq revealed new mechanisms of γδ CAR T exhaustion and effector differentiation distinct from αβ CAR T cells. In vivo, LTβR-armored αβ and γδ CAR T cells targeting CLDN6 mediated complete, durable regressions in an aggressive ovarian cancer model where controls were ineffective. Finally, we established a GMP-compatible manufacturing process tailored to γδ T cell biology, producing highly pure, potent cells suitable for multi-patient allogeneic use. Together, this work defines a comprehensive next-generation engineering and manufacturing framework that overcomes γδ CAR T cell limitations and enables durable activity in solid tumors. Citation Format: Mitchell S. Wang, Ivan Reyes-Torres, Kristen C. Vogt, Caroline K. Hanlon, Ashley N. Thornal, Maria Guarino, Eric M. Johnson, Mark P. Roberto, Navamallika G. Reddy, Miao Chen, Theodore Giavridis, Mateusz Legut. Synthetic reprogramming of γδ CAR T cells with the LTBR gene optimizes function in the TME and overcomes effector limitations [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 128.