Abstract B017: Immune engineering to enhance CAR T cell trafficking in pancreatic ductal adenocarcinoma

Abstract

Pancreatic ductal adenocarcinoma (PDA) remains one of the most lethal malignancies in the United States, largely due to its resistance to conventional treatments. Although immunotherapies have revolutionized the treatment landscape for hematological malignancies, their translation to solid tumors such as PDA remains limited. A significant barrier to effective CAR T cell therapy in PDA is the desmoplastic stroma, which obstructs T cell infiltration and promotes cellular dysfunction through physical sequestration and exhaustion. To address this challenge, our research focuses on leveraging genome engineering to enhance CAR T cell trafficking and function within the fibrotic tumor microenvironment (TME). Specifically, we aim to modulate the expression of key focal adhesion-associated genes that govern T cell motility and adhesion dynamics in PDA. We hypothesize that optimizing these pathways will confer improved intratumoral CAR T cell surveillance, yielding greater immune cell accumulation and enhanced anti-tumor activity. We utilized state-of-the-art multiphoton laser scanning microscopy (MPLSM) to visualize T cell migration and immune-ECM interactions in both live tumor slices and 3D ECM-mimicking platforms. This multimodal approach enabled us to generate multidimensional (x, y, z, c, t) datasets by coupling second harmonic generation (SHG) imaging of collagen architecture with fluorescent labeling of immune and carcinoma cells. Resulting T cell trajectories were characterized using mathematical modeling approaches to study T cell dynamics under distinct focal adhesion blockade conditions. Together, these 3D in vitro platforms constitute a powerful tool for preliminary screening of candidate targets. Our findings reveal that T cell migration on ECM substrates follows a biphasic dependence on ligand density, indicating that an adhesion ligand-rich TME may paradoxically impede T cell motility. Using this platform, we identified potential targets that improve T cell speed, persistence, and sampling in the TME. We subsequently used genome engineering to encode these migratory features into CAR T cells, thereby enhancing their infiltration in orthotopic models of pancreatic cancer. Collectively, these results suggest that engineering adhesion dynamics can potentially surmount a central limitation of T cell accumulation in solid tumors such as PDA.