Gaglione et al. developed TCRAFT, a pooled synthesis method to rapidly assemble large TCR libraries (>104) with high accuracy (>99%) and native ɑ/β chain pairing. Integration with RAPTR, a pMHC-targeted lentiviral screening approach for detection of activated T cells, enabled one-pot library/library antigen reactivity screening. The method elucidated specific TCR/peptide pairs among 3,808 vitiligo-derived TCRs with similar fidelity to peptide/APC screening. Overlay of clonotype identity with scRNAseq data revealed a signature of melanocyte-reactive clones and enabled comparisons between identical clones involved in melanoma and vitiligo.
Contributed by Morgan Janes
ABSTRACT: T cells initiate targeted immune responses using T cell receptors (TCRs) to recognize specific antigens. Mapping TCRs to antigens at scale remains a major challenge. Here, we developed an approach to synthesize and functionally screen tens of thousands of TCRs simultaneously. TCR rapid assembly for functional testing (TCRAFT) uses a modular strategy to rapidly and inexpensively construct large pools of TCRs from sequences while maintaining TCRα/β pairing. We applied TCRAFT to reconstruct over 3,800 TCRs from vitiligo blister fluid and mapped these TCRs to specific peptide-major histocompatibility complexes using RAPTR, an activation-based library-on-library screening approach. Vitiligo antigen-specific T cells displayed pronounced clonal expansion and transcriptomic signatures similar to antigen-specific T cells in melanoma, pointing to shared features of disease-relevant T cells in autoimmunity and cancer. Demonstrating scalability, we synthesized and screened over 30,800 TCRs from donors with pancreatic ductal adenocarcinoma to capture antigen-reactive TCRs. Our approach expands the scale and accessibility of TCR-antigen screening, which is critical to understanding immunity and developing new immunotherapies.
Author Info: (1) Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA; Koch Institute for Integrative Cancer Research, Cambridge, MA, USA. (2) Koch Inst
Author Info: (1) Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA; Koch Institute for Integrative Cancer Research, Cambridge, MA, USA. (2) Koch Institute for Integrative Cancer Research, Cambridge, MA, USA; Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA. (3) Inflammation & Immunology Research Unit, Pfizer Inc., Cambridge, MA 02139, USA. (4) Koch Institute for Integrative Cancer Research, Cambridge, MA, USA; Department of Immunology, Harvard Medical School, Boston, MA, USA. (5) Inflammation & Immunology Research Unit, Pfizer Inc., Cambridge, MA 02139, USA. (6) Inflammation & Immunology Research Unit, Pfizer Inc., Cambridge, MA 02139, USA. (7) Koch Institute for Integrative Cancer Research, Cambridge, MA, USA; Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA. (8) Department of Immunology, Harvard Medical School, Boston, MA, USA; Ragon Institute of MGH, MIT, and Harvard, Cambridge, MA, USA. (9) Department of Dermatology, University of Massachusetts Chan Medical School, 364 Plantation St, Worcester, MA, USA. (10) Program in Bioinformatics and Integrative Biology, University of Massachusetts Chan Medical School, Worcester, MA, USA. (11) Department of Medicine, Division of Gastroenterology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA; Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, MA, USA. (12) Department of Immunology, Harvard Medical School, Boston, MA, USA; Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, MA, USA. (13) Fletcher Biosciences, Inc., Cambridge, MA, USA. (14) Koch Institute for Integrative Cancer Research, Cambridge, MA, USA; Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA. (15) Koch Institute for Integrative Cancer Research, Cambridge, MA, USA; Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA. (16) Koch Institute for Integrative Cancer Research, Cambridge, MA, USA; Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA. (17) Inflammation & Immunology Research Unit, Pfizer Inc., Cambridge, MA 02139, USA. (18) Inflammation & Immunology Research Unit, Pfizer Inc., Cambridge, MA 02139, USA. (19) Koch Institute for Integrative Cancer Research, Cambridge, MA, USA; Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA. (20) Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA, USA; Department of Medicine, Harvard Medical School, Boston, MA, USA. (21) Program in Bioinformatics and Integrative Biology, University of Massachusetts Chan Medical School, Worcester, MA, USA. (22) Department of Medicine, Division of Gastroenterology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA; Department of Medicine, Harvard Medical School, Boston, MA, USA. (23) Department of Immunology, Harvard Medical School, Boston, MA, USA; Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, MA, USA. (24) Department of Dermatology, University of Massachusetts Chan Medical School, 364 Plantation St, Worcester, MA, USA. (25) Inflammation & Immunology Research Unit, Pfizer Inc., Cambridge, MA 02139, USA. (26) Koch Institute for Integrative Cancer Research, Cambridge, MA, USA; Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA; Ragon Institute of MGH, MIT, and Harvard, Cambridge, MA, USA. Electronic address: mbirnb@mit.edu.
