Scalable TCR synthesis and screening enable antigen reactivity mapping in vitiligo

Stephanie A Gaglione; Rachit S Mukkamala; Chirag Krishna; Blake E Smith; Marc H Wadsworth 2nd; Scott A Jelinsky; Caleb R Perez; Laura Schmidt-Hong; Erica L Katz; Kyle J Gellatly; Lestat R Ali; Jiao Shen; Patrick V Holec; Qingyang Henry Zhao; Amanda O Chan; Ellen J K 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

doi:10.1016/j.immuni.2026.01.001

Scalable TCR synthesis and screening enable antigen reactivity mapping in vitiligo

Immunity. 2026 Feb 10;59(2):477-493.e9. doi: 10.1016/j.immuni.2026.01.001. Epub 2026 Jan 28.

Authors

Stephanie A Gaglione¹, Rachit S Mukkamala², Chirag Krishna³, Blake E Smith⁴, Marc H Wadsworth 2nd³, Scott A Jelinsky³, Caleb R Perez², Laura Schmidt-Hong⁵, Erica L Katz⁶, Kyle J Gellatly⁷, Lestat R Ali⁸, Jiao Shen⁹, Patrick V Holec¹⁰, Qingyang Henry Zhao², Amanda O Chan², Ellen J K Xu², Kellie M Kravarik³, Julia A Guzova³, Connor S Dobson², Harshabad Singh¹¹, Manuel Garber⁷, Michael Dougan¹², Stephanie K Dougan⁹, John E Harris⁶, Aaron Winkler³, Michael E Birnbaum¹³

Affiliations

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 Department of Immunology, Harvard Medical School, Boston, MA, USA; Ragon Institute of MGH, MIT, and Harvard, Cambridge, MA, USA.
⁶ Department of Dermatology, University of Massachusetts Chan Medical School, 364 Plantation St, Worcester, MA, USA.
⁷ Program in Bioinformatics and Integrative Biology, University of Massachusetts Chan Medical School, Worcester, MA, USA.
⁸ 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.
⁹ Department of Immunology, Harvard Medical School, Boston, MA, USA; Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, MA, USA.
¹⁰ Fletcher Biosciences, Inc., Cambridge, MA, USA.
¹¹ Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA, USA; Department of Medicine, Harvard Medical School, Boston, MA, USA.
¹² 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.
¹³ 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.

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.

Keywords: T cell receptor; TCR specificity; antigen recognition; autoimmunity; high-throughput functional screening; peptide MHC; viral display; vitiligo.

MeSH terms

Epitope Mapping* / methods
Humans
Receptors, Antigen, T-Cell* / genetics
Receptors, Antigen, T-Cell* / immunology
Receptors, Antigen, T-Cell* / metabolism
T-Lymphocytes* / immunology
Vitiligo* / immunology

Substances

Receptors, Antigen, T-Cell

Abstract

MeSH terms

Substances

Grants and funding