Protective T-cell receptor identification for orthotopic reprogramming of immunity in refractory virus infections.

Abstract

Viral infections cause life-threatening disease in immunocompromised patients and especially following transplantation. T-cell receptor (TCR) engineering redirects specificity and can bring significant progress to emerging adoptive T-cell transfer (ACT) approaches. T-cell epitopes are well described, whereas knowledge is limited which TCRs mediate protective immunity. Here, refractory adenovirus (AdV) infection after hematopoietic stem cell transplantation (HSCT) was treated with ACT of highly purified Hexon5-specific T cells using pMHC-Streptamers against the immunodominant HLA-A*0101-restricted peptide LTDLGQNLLY. AdV was successfully controlled through this oligoclonal ACT. Novel protective TCRs were isolated ex vivo and preclinically engineered into the TCR locus of allogeneic 3rd-party primary T cells by CRISPR/Cas9-mediated orthotopic TCR replacement. Both, TCR knock-out and targeted integration of the new TCR in one single engineering step led to physiological expression of the transgenic TCR. Reprogrammed TCR-edited T cells showed strong virus-specific functionality like cytokine release, effector marker upregulation and proliferation capacity, as well as cytotoxicity against LTDLGQNLLY-presenting and AdV-infected targets. In conclusion, ex vivo isolated TCRs with clinical proven protectivity through ACT could be redirected into T cells from naïve 3rd-party donors. This approach ensures that transgenic TCRs are protective with potential off-the-shelf use and widened applicability of ACT to various refractory emerging viral infections.