CXCR4-dependent migratory differences between peripheral Tregs and Tcons preserve GvL while limiting GvHD after HSCT

Although studies on HLA-haploidentical hematopoietic stem cells transplant (HSCT) for high-risk acute myeloid leukemia (AML) showed that regulatory and conventional T-lymphocyte (Treg/Tcon) immunotherapy prevents graft-versus-host disease (GvHD) while it exerts T-cell dependent graft-versus-leukemia (GvL) effect, its mechanism is less known. The present study clarifies the mechanisms underlying this finding. In a xenogenic immunotherapy model, non-obese diabetic-scidIl2rgtm mice were engrafted with human primary AML and were treated with allogeneic human peripheral blood Tregs followed by Tcons. Treg/Tcon immunotherapy cleared leukemia without causing GvHD. Peripheral blood Tregs are largely CD45RO+ and express low levels of CXCR4 bone marrow (BM) homing receptor. They localised in peripheral tissues (i.e., liver, gut), but not in the BM. Consequently, in the BM Tcon effector function was not downregulated and indeed Tcons killed leukemia. In contrast, in peripheral tissues, they showed no alloreactivity suggesting their effector function had been downregulated by the Tregs. Thus, the GvL effect without GvHD was due to the migratory features of peripheral blood Tregs. These data were confirmed in an MHC-mismatched BM transplant model using H-2b mice as recipients and H-2d mice as donors. Remarkably, in patients receiving haploidentical T-cell depleted HSCT with Treg/Tcon immunotherapy, the infused Tregs were not found in the BM in the first month post-transplant. In contrast, donor Tcons homed to the BM where they exerted leukemia killing. In conclusion, for the first time we show the mechanism whereby immunotherapy with Tregs/Tcons protects from GvHD while it preserves GvL in murine transplantation models and in transplanted leukemia patients.