Abstract LB-C006: Scalable TCR synthesis and screening enable antigen reactivity mapping

Abstract Mapping T cells to their antigenic targets is critical to developing new targeted cancer therapies but remains highly challenging at scale. Single-cell datasets capture thousands of T cell receptors (TCRs) from tumor-infiltrating T cells but offer little data on antigen specificity. Here, we introduce an approach to synthesize and functionally screen tens of thousands of T cell receptors simultaneously. TCRAFT uses a modular strategy to reconstruct TCRs from sequences for under 1 per TCR while preserving TCRα-β pairing. We then integrate TCRAFT with RAPTR, a library-on-library approach to screen thousands of TCRs against antigens at scale. Using this workflow, we precisely reconstructed 3, 808 TCRs from vitiligo blister fluid and screen them against hundreds of antigens to identify over 160 antigen-specific TCR clonotypes. By linking TCR specificity with corresponding single-cell gene expression data, we establish transcriptomic signatures of vitiligo-associated autoreactive T cells and correlate this signature with TILs in melanoma. Demonstrating scalability, we synthesize and screen over 30, 800 TCRs at once from donors with pancreatic ductal adenocarcinoma to isolate antigen-specific TCRs. By enabling rapid and cost-effective TCR assembly and antigen mapping, our pipeline is accessible to any research group to identify clinically relevant TCRs and decode tumor-specific immune responses. Citation Format: Stephanie A. Gaglione, Rachit S. Mukkamala, Chirag Krishna, Blake E. Smith, Marc H. Wadsworth, Caleb R. Perez, Laura Schmidt-Hong, Erica L. Katz, Kyle J. Gellatly, Lester R. Ali, Jiao Shen, Scott Jelinsky, Patrick V. Holec, Qingyang H. Zhao, Amanda O. Chan, Ellen JK. Xu, Kellie M. Kravarik, Julia A. Guzova, Connor S. Dobson, Harshabad Singh, Manuel Garber, Michael Dougan, Stephanie K. Dougan, John E. Harris, Aaron Winkler, Michael E. Birnbaum. Scalable TCR synthesis and screening enable antigen reactivity mapping abstract. In: Proceedings of the AACR Immuno-Oncology Conference (AACR IO): Discovery and Innovation in Cancer Immunology: Revolutionizing Treatment through Immunotherapy; 2026 Feb 18-21; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Immunol Res 2026;14 (2 Suppl): Abstract nr LB-C006.