Early administration of calcineurin inhibitors limits tolerogenic effect of PTCy in allogeneic hematopoietic transplant.

We recently demonstrated early administration of calcineurin inhibitors (CNIs) after murine allogeneic hematopoietic cell transplantation (allo-HCT) inhibits terminal exhaustion of donor T cells and hampers tolerance induction. However, the role of CNIs in regulating T-cell exhaustion in clinical allo-HCT remains to be clarified. It also remains to be elucidated why posttransplant cyclophosphamide (PTCy) could reduce chronic graft-versus-host disease (cGVHD), despite the administration of CNIs. In this study, we explored the impact of CNIs on donor T-cell exhaustion after PTCy-based haploidentical HCT. In mice, early administration of cyclosporine before PTCy preserved a population of Ly6C⁺ donor T cells with less exhausted characteristics, as has been shown in non-PTCy allo-HCT. In single-cell analysis of clinical samples from patients undergoing HLA-haploidentical peripheral blood stem cell transplantation with PTCy (PTCy-haplo-PBSCT), early administration of tacrolimus prior to PTCy promoted the expansion of effector-like CD8+ intermediate exhausted T cells (Tex-int) and CD4⁺ cytotoxic T cells (CTLs), with the former enriched for both effector- and exhaustion-associated and the latter for effector- and CD4⁺ CTL-specific gene signatures. These subsets retained high responsiveness to TCR stimulation and PD-1 blockade, and their expansion on day 28 predicted subsequent cGVHD. In contrast, delaying tacrolimus administration on day 5 after PTCy-haplo-PBSCT significantly reduced the expansion of these effector-like T cells. These findings indicate early CNI exposure favors the persistence of incompletely exhausted and clonally expanded donor T cells with pathogenic potential. Tex-int and CD4⁺ CTLs may serve as prognostic biomarkers of cGVHD risk and as promising targets for preemptive therapy of cGVHD.