Cytotoxic T lymphocyte antigen‐4 regulates development of xenogenic graft versus host disease in mice via modulation of host immune responses induced by changes in human T cell engraftment and gene expression

Graft versus host disease (GvHD) is a major clinical problem with a significant unmet medical need. We examined the role of cytotoxic T lymphocyte antigen‐4 (CTLA‐4) in a xenogenic GvHD (xeno‐GvHD) model induced by injection of human peripheral mononuclear cells (hPBMC) into irradiated non‐obese diabetic (NOD) SCID gamma (NSG) mice. Targeting the CTLA‐4 pathway by treatment with CTLA‐4 immunoglobulin (Ig) prevented xeno‐GvHD, while anti‐CTLA‐4 antibody treatment exacerbated the lethality and morbidity associated with GvHD. Xeno‐GvHD is associated with infiltration of hPBMCs into the lungs, spleen, stomach, liver and colon and an increase in human proinflammatory cytokines, including interferon (IFN)‐γ, tumor necrosis factor (TNF)‐α and interleukin (IL)‐5. Infiltration of donor cells and increases in cytokines were attenuated by treatment with CTLA‐4 Ig, but remained either unaffected or enhanced by anti‐CTLA‐4 antibody. Further, splenic human T cell phenotyping showed that CTLA‐4 Ig treatment prevented the engraftment of human CD45 + cells, while anti‐CTLA‐4 antibody enhanced donor T cell expansion, particularly CD4 + (CD45RO + ) subsets, including T box transcription factor TBX21 (Tbet) + CXCR3 + and CD25 + forkhead box protein 3 (FoxP3) cells. Comprehensive analysis of transcriptional profiling of human cells isolated from mouse spleen identified a set of 417 differentially expressed genes (DEGs) by CTLA‐4 Ig treatment and 13 DEGs by anti‐CTLA‐4 antibody treatment. The CTLA‐4 Ig regulated DEGs mapped to down‐regulated apoptosis, inflammasome, T helper type 17 (Th17) and regulatory T cell (T reg ) pathways and enhanced Toll‐like receptor (TLR) receptor signaling, TNF family signaling, complement system and epigenetic and transcriptional regulation, whereas anti‐CTLA‐4 antibody produced minimal to no impact on these gene pathways. Our results show an important role of co‐inhibitory CTLA‐4 signaling in xeno‐GvHD and suggest the therapeutic utility of other immune checkpoint co‐inhibitory pathways in the treatment of immune‐mediated diseases driven by hyperactive T cells.