Elucidating the Role of CD8+ TCR Attributes in Cancer Immunotherapy

About This Grant

PROJECT ABSTRACT Pancreatic ductal adenocarcinoma (PDA) is a highly lethal malignancy with the lowest 5-year survival rate among all solid tumors. Adoptive cell transfer (ACT) of tumor neoepitope specific CD8+ T cells has shown therapeutic promise, but current approaches rely on peptide-MHC tetramers to select high-affinity T cell receptors (TCRs), potentially overlooking functionally superior clones. Our preliminary data reveal that low- affinity TCRs—particularly those undetectable by tetramers—can mediate potent antitumor responses, suggesting that TCR bond lifetime under mechanical force, rather than affinity alone, may dictate therapeutic efficacy. To address this, I developed a novel panel of T cell receptor exchange (TRex) mice expressing PDA- specific TCRs spanning a spectrum of affinities for the same tumor neoantigen (CB101-109:H2-Db). Strikingly, the lowest-affinity TCR (TCR5.1) demonstrated enhanced tumor control compared to higher-affinity counterparts, challenging conventional TCR selection paradigms. This proposal will define how TCR biophysical properties influence CD8+ T cell fate and function in PDA through two specific aims. Aim 1, will employ a biomembrane force probe (BFP) to quantify TCR-pMHC bond lifetimes and correlate these with ACT outcomes in orthotopic PDA models, using high-resolution ultrasound and multimodal T cell profiling. In collaboration with Brian Evavold’s lab at the University of Utah, I will use BFP analysis to quantify each TCRs affinity, bond lifetime, and ideal force conditions. I will then evaluate the effector functions of each clonotype ex vivo using titrated peptide stimulation assays and assess tumor clearance in vivo through ACT in immunocompetent orthotopic PDA models. Aim 2 will dissect competitive fitness of TCRs during immunotherapy by co-transferring T cell clones into immunocompetent hosts, combining scRNAseq and spatial analyses to uncover mechanisms driving TCR-dependent persistence. By “parking” naïve TRex CD8+ T cells in mice prior to orthotopic tumor implantation, I will identify the impact of competition between TCRs of varying affinities on the anti-tumor response during immune checkpoint blockade or CD40 agonist therapies. Tracking these clonal populations through single-cell RNA sequencing and spatial analyses will allow for the identification of the molecular and cellular mechanisms that drive TCR-dependent persistence and stem-like maintenance. Together, these studies will test the central hypothesis that low-affinity, long-lived TCR interactions promote the development of durable, stem-like T cell populations that are resistant to exhaustion - a potential key to achieving sustained antitumor responses. The successful completion of this research will provide transformative insights into TCR- intrinsic determinants of therapeutic efficacy, with direct implications for the design of improved ACT protocols for PDA and other solid tumors.