A single-nucleotide polymorphism in the human ENTPD1 gene encoding CD39 is associated with worsened graft-versus-host disease in a humanized mouse model

Regulatory T cells (Tregs) protect against graft-versus-host disease (GVHD), a life-threatening complication of allogeneic hematopoietic stem cell transplantation. The ectoenzyme CD39 is important for increasing the immunosuppressive function of Tregs. The rs10748643 (A -> G) single-nucleotide polymorphism (SNP) in intron 1 of the human ENTPD1 gene is associated with increased proportions of CD39(+) Tregs. This study aimed to determine whether the rs10748643 SNP corresponded to increased proportions of CD39(+) Tregs in an Australian donor population, and whether this SNP influences clinical GVHD in a humanized mouse model. Donors were genotyped for the rs10748643 SNP by Sanger sequencing, and the proportion of CD39(+) T cells in donor peripheral blood was determined by flow cytometry. Donors encoding the G allele (donors(AG/GG)) demonstrated higher proportions of CD39(+)CD3(+)CD4(+)CD25(+)CD127(lo) Tregs, but not CD39(+)CD3(+)CD8(+) T cells or CD39(+)CD3(+)CD4(+) conventional T cells, compared with donors homozygous for the A allele (donors(AA)). NOD-SCID-IL2R gamma(null) mice were injected with human peripheral blood mononuclear cells from either donors(AA) (hCD39(AA) mice) or donors(AG/GG) (hCD39(AG/GG) mice). hCD39(AG/GG) mice demonstrated significantly greater weight loss and GVHD clinical scores, and significantly reduced survival, compared with hCD39(AA) mice. hCD39(AG/GG) mice showed significantly higher hCD4(+):hCD8(+) T-cell ratios than hCD39(AA) mice, but displayed similar proportions of CD3(+)hCD4(+)hCD25(+)hCD127(lo) Tregs and hCD39(+) Tregs. However, the proportion of human Tregs corresponded to survival in hCD39(AA) mice, but not in hCD39(AG/GG) mice. This study demonstrates that donors encoding the G allele show higher percentages of CD39(+) Tregs, but cause worsened GVHD in humanized mice compared with donors homozygous for the A allele.