Donor Variability and PD-1 Expression Limit BK Polyomavirus-specific T-cell Function and Therapy.

BACKGROUND: BK polyomavirus (BKPyV) nephropathy is a major cause of premature kidney transplant failure. Current management relies on reducing immunosuppression to restore BKPyV-specific immune control. Ex vivo expansion and transfer of BKPyV-specific cytotoxic T cells prepared from third-party donors may enhance virus-specific treatment, but the efficacy seems suboptimal. METHODS: To optimize BKPyV-specific T-cell expansion protocols, we compared conventional and G-Rex expansion cultures at 10 and 14 d after stimulation with BKPyV overlapping peptide pools. Cytokine and cytotoxic responses were assessed as well as programmed cell death protein 1 (PD-1) and programmed cell death ligand 1 (PD-1L) expression on effector and target cells, respectively. RESULTS: Despite all donors being BKPyV-IgG seropositive, BKPyV-specific T-cell responses were heterogeneous and varied in magnitude between individuals. Overall, we observed higher cell counts in G-Rex compared to conventional cultures. Upon restimulation with 15mer-pools or immunodominant 9mer-pools, expanded BKPyV-specific T cells expressed polyfunctional markers, for example, interferon-γ, tumor necrosis factor-α and CD107a, and were cytotoxic for 9mP-pulsed autologous phytohemagglutinin blasts or BKPyV-infected allogeneic renal proximal tubule epithelial cells (RPTECs). Compared with conventional cultures, G-Rex-expanded CD4 and CD8 T cells showed higher PD-1 expression. Pembrolizumab reduced PD-1 expression on BKPyV-specific T cells and augmented polyfunctional BKPyV-specific T-cell responses and cytotoxicity. Interferon-𝛾 increased PD-L1 expression on BKPyV-infected RPTECs and increased viability. CONCLUSIONS: Upregulated PD-1 expression of ex vivo expanded T cells contributes to third-party donor variability and potentially impairs the efficacy of adoptive T-cell therapy. Because BKPyV-infected RPTECs increase PD-L1 under inflammatory conditions, adding immune checkpoint inhibitors ex vivo before infusion could be evaluated for enhanced clinical efficacy when attempting treatment of BKPyV-associated pathologies without jeopardizing transplantation outcomes.