Cell therapy with human IL-10-producing ILC2s limits xenogeneic Graft-versus-Host Disease by inhibiting pathogenic T cell responses. Cell therapy with human IL-10-producing ILC2s limits xenogeneic Graft-versus-Host Disease by inhibiting pathogenic T cell responses

IL-10-producing group 2 innate lymphoid cells (ILC210) have important functions in regulating immune responses, yet their therapeutic potential remains largely unexplored. Here, we demonstrate that cell therapy with human ILC210 inhibits pathogenic T cell responses in humanized mouse models of Graft-versus-Host Disease (GVHD). Both prophylactic or post-onset adoptive transfer of human ILC210 significantly reduced GVHD severity and improved survival in NOD-scid IL2Rγnull (NSG) mice. Notably, ILC210 provided comparable protection to regulatory T cells (Tregs) without impairing T cell engraftment, instead decreasing intestinal T cell infiltration and suppressing CD4+Th1 and CD8+Tc1. Additionally, ILC210 conferred superior protection from GVHD than conventional IL-10-/lowILC2s, and blocking IL-10 and IL-4 abrogated the protective effects of ILC210 cell therapy. In GVHD-protected HSCT recipients, ILC2s with an ILC210 phenotype were present at normal levels and inversely correlated with Th1 cells, consistent with findings in humanized mouse models. Critically, ILC210 did not impair T cell-mediated graft-versus-leukemia effects in a humanized model. CITE-seq analysis identified CD49d and CD86 as novel markers that allow for enrichment of ILC210 from conventional ILC2s, and to track ILC210 in patient studies. These results establish a platform for >2000-fold expansion of human ILC210 with a stable phenotype and underscore ILC210 potential in cell therapies for GVHD and other immune-mediated diseases. Overall design: ILC2s, ILC3, CD56bright and CD56dim NK cells were all stained and FACS sorted from a healthy donor. Cells were expanded for 28 days in distinct growth media for each cell type. Cells were stained with TotalSeqC antibodies including hashtags and then analyzed using CITE-seq. Stained ILC2s were mixed 1:1 with CD56dim NK cells and ILC3s mixed 1:1 with CD56dim NK cells. Each mixed sample was run on separate lanes and identified using hashtag antibodies. ILC2s and CD56dim NK cells were stained with Hashtag1, CD56bright NK cells with Hashtag2 and ILC3s with Hashtag3. *************************************************************** Submitter states that missing raw files are due to patient privacy concerns. ***************************************************************