Xu, E. J. | bioRxiv Science

bioRxiv2025

Scalable TCR synthesis and screening enables antigen reactivity mapping in vitiligo

T cell receptors (TCRs) mediate antigen recognition in adaptive immunity, yet large-scale mapping of TCR-antigen interactions remains a major challenge. Current approaches to synthesize and functionally screen TCRs remain technically complex and limited in throughput. We introduce a modular strategy, TCRAFT, to rapidly construct tens of thousands of TCRs for <$1 each while maintaining TCR-{beta} pairing with >99% accuracy. We integrate this approach with a high-throughput antigen discovery platform to enable library-on-library TCR-antigen screening. We reconstruct and screen 3,808 TCRs from vitiligo lesions, linking TCR specificity to transcriptional phenotypes for antigen-reactive T cells. To demonstrate scalability, we synthesize and screen 30,810 TCRs from donors with pancreatic ductal adenocarcinoma to capture antigen-specific TCRs. This workflow reduces the cost and complexity of large-scale TCR screening, enabling the expansion of the known landscape of antigen-specific TCRs in vitiligo with a method that can be readily extended to other immunological applications.

immunology↗

bioRxiv2024

Peptide-MHC-targeted retroviruses enable in vivo expansion and gene delivery to tumor-specific T cells

Tumor-infiltrating-lymphocyte (TIL) therapy has demonstrated that endogenous T cells can be harnessed to initiate an effective anti-tumor response. Despite clinical promise, current TIL production protocols involve weeks-long ex vivo expansions which can affect treatment efficacy. Therefore, additional tools are needed to engineer endogenous tumor-specific T cells to have increased potency while mitigating challenges of manufacturing. Here, we present a strategy for pseudotyping retroviral vectors with peptide-major histocompatibility complexes (pMHC) for antigen-specific gene delivery to CD8 T cells and examine the efficacy of these transduced cells in immunocompetent mouse models. We demonstrate that pMHC-targeted viruses are able to specifically deliver function-enhancing cargoes while simultaneously activating and expanding anti-tumor T cells. The specificity of these viral vectors enables in vivo engineering of tumor-specific T cells, circumventing ex vivo manufacturing processes and improving overall survival in B16F10-bearing mice. Altogether, we have established that pMHC-targeted viruses are efficient vectors for reprogramming and expanding tumor-specific populations of T cells directly in vivo, with the potential to substantially streamline engineered cell therapy production for a variety of applications.

immunology↗