De novo-designed pMHC binders facilitate T cell-mediated cytotoxicity toward cancer cells

The recognition of intracellular antigens by CD8⁺ T cells through T cell receptors (TCRs) is central for adaptive immunity against infections and cancer. However, the identification of TCRs from patient material remains complex. We present a rapid de novo minibinder (miBd) design platform leveraging state-of-the-art generative models to engineer miBds targeting the cancer-associated peptide-bound major histocompatibility complex (pMHC) SLLMWITQC/HLA-A*02:01 (NY-ESO-1). Incorporating in silico cross-panning enabled computational prescreening of specificity, and molecular dynamics simulations allowed for improved predictability of in vitro success. We identified a high-affinity NY-ESO-1 binder and confirmed its structure using cryo-electron microscopy, which, when incorporated in a chimeric antigen receptor, induced killing of NY-ESO-1⁺ melanoma cells. We further designed and validated binders to a neoantigen pMHC complex, RVTDESILSY/HLA-A*01:01, with unknown structure, demonstrating the potential for precision immunotherapy.