Low- and high-dose post-transplant cyclophosphamide attenuates graft-versus-host disease with distinct effects on...

Allogeneic haematopoietic stem cell transplantation (alloHSCT) is a curative treatment for haematological malignancies. AlloHSCT aims to generate graft-versus-leukaemia immunity, where donor T cells eliminate residual malignant cells. However, graft-versus-host disease (GVHD), where donor T cells attack recipient tissues, is a common and often fatal side effect. Post-transplant cyclophosphamide (PTCy) can reduce GVHD, but the cellular mechanisms through which this occurs are not fully understood, and high doses may be associated with toxicity. This study aimed to determine whether lower doses of PTCy can reduce GVHD and to examine the effects of PTCy doses on human (h) immune cell subsets, T cell exhaustion, and histological GVHD in a humanised mouse model. NOD-scid-IL2Rγnull mice were injected with 2 × 107 human peripheral blood mononuclear cells on day 0, cyclophosphamide (10, 25 or 33 mg/kg) or control diluent on days 3 and 4 and monitored for GVHD development at early and late time points. Low-dose PTCy (10 mg/kg) abrogated clinical signs of GVHD with comparable efficacy to high-dose PTCy (33 mg/kg), delaying GVHD onset and prolonging mouse survival. Proportions of hPD-1+ hCD4+ and hPD-1+hCD8+ T cells were increased with low-dose PTCy but not higher doses, while hPD-1+ hTreg proportions were increased by all PTCy doses. Exhausted hPD-1+hLAG3+hCD8+ T cell proportions were increased with high-dose PTCy, but not lower doses. This study indicates that low-dose PTCy reduces GVHD with similar efficacy to that of high-dose PTCy, but this appears to be associated with differing cellular mechanisms of action.