Abstract B085: Development of a Translational Manufacturing Strategy for a Novel CD4+ CD26+ mesoCAR T Cell Product for Pancreatic Ductal Adenocarcinoma Patients

Abstract

The emergence of KRAS inhibitors is reshaping the therapeutic landscape of pancreatic ductal adenocarcinoma (PDAC). Yet responses are seldom durable, and relapse is inevitable, highlighting the ongoing need for aggressive therapeutic strategies. PDAC also remains unresponsive to adoptive T cell therapies (ACT), highlighting the need for both improved cellular substrates and clinically feasible manufacturing strategies. Chimeric antigen receptor (CAR) T cell therapy engineers a patient's lymphocytes to recognize and eliminate tumors. Previous CAR T cell approaches have focused on engineering bulk CD3+ T cells. Our previous preclinical work indicates that CD4+ T cells expressing the ectoenzyme CD26 are a superior helper subset capable of driving durable antitumor immunity. This study aims to leverage this biology to develop a streamlined, good manufacturing practice (GMP)-compatible platform to generate a novel CD26+ CD4+ mesothelin-targeted CAR T cell product to treat patients with PDAC. We established a manufacturing workflow enabling parallel isolation of bulk CD8+ T cells and CD26+ CD4+ T cells from the peripheral blood of healthy versus PDAC patients, followed by engineering with a fully humanized mesothelin CAR that triggers CD3ζ and 4-1BB downstream of antigen recognition. These CAR T cell products are expanded in the presence of a PI3Kd/g inhibitor, duvelisib, which preserves a stem-like memory phenotype. This process was optimized for translational scalability using anti-CD3/CD28/CD2 T cell activation and G-Rex culture systems, substantially reducing hands-on time and complexity. Across both healthy donors (n=3) and patients with advanced PDAC (n=3), CAR T cell products could be reproducibly generated with comparable expansion kinetics, transduction efficiencies and phenotypic stability. While CAR CD8+ T cells from healthy donors exhibited modestly higher TCF1+ LEF1+ co-expression vs. patient-derived CAR T cells, CD26+ CD4+ T cells retained robust stem-associated features, regardless of donor source. Functional testing against human PDAC cell lines with graded mesothelin expression revealed equivalent cytolytic capacity between PDAC patient and donor-derived products, with enhanced and accelerated killing observed against high-mesothelin targets. Notably, CD26+ CD4+ CAR T cells mediated effective tumor cell lysis both as a standalone product and in combination with CD8+ CAR T cells. This strategy has now been transferred to the cell manufacturing stage, generating investigational new drug (IND)-enabling runs to gain regulatory supportive data needed to open a clinical study. This platform provides a foundation for first-in-human evaluation of CD26+ CD4+ CAR T cells in PDAC and informs broader efforts to rationally incorporate helper T cell biology into next-generation cell therapies for solid tumors. Importantly, we envision deploying this product in combination with or following KRAS inhibitor therapy, intercepting minimal residual disease at the point of relapse to convert transient responses into durable remissions.