Characterization of host immune cell infiltrate in human CAR T cell-mediated xenogeneic graft versus host disease in NSG mice

Chimeric antigen receptor (CAR) T cells are revolutionary cancer therapies that are Food and Drug Administration-approved for hematologic malignancies and under investigation for solid tumors. The use of allogeneic over autologous CAR T cells offers advantages, including broader availability and reduced costs. However, allogeneic CAR T cells frequently trigger graft versus host disease (GvHD), a complication observed in patients and experimental models where human CAR T cells are delivered into immunocompromised mice. To understand the contribution of the mouse immune response to human CAR T cell-mediated xenogeneic GvHD, we analyzed GvHD lesions in a human xenograft tumor model in NOD.Cg- Prkdc scid Il2rg tm1Wjl /SzJ (NSG) mice. The animals were treated with second-generation CAR T cells targeting a human tumor-specific antigen without a murine homolog. Mice treated with CAR T cells had more severe GvHD lesions than control mice receiving nontransduced (NT) T cells. Also, tumor burden was negatively correlated with GvHD lesion severity. Immunohistochemical characterization of the GvHD lesions showed that approximately 45% of the immune cell infiltrate consisted of murine cells, most of which were IBA1+ histiocytes, with a small population of CD11c+ dendritic cells. The murine histiocytes expressed activation/antigen presentation markers, including high levels of the costimulatory molecule CD86. Analysis of macrophage polarization indicated an M2-like phenotype. These findings demonstrate a significant contribution of the mouse histiocytic compartment to lesions of human CAR T cell-mediated xenogeneic GvHD. Our results suggest that CD86+ murine antigen-presenting cells help trigger and sustain the xenoreactive CAR T cell response. Furthermore, xenogeneic GvHD exhibits a shift toward M2 polarization in murine macrophages.