CRISPR/Cas9 Gene Editing of MIR155HG in Primary Human T Cells to Prevent Acute Graft-Versus-Host Disease

Abstract Introduction: Acute graft-versus-host disease (GVHD) mediated by donor alloreactive T cells is the leading cause of non-relapse mortality in patients post-allogeneic hematopoietic cell transplantation. Here we genetically engineered genomic deletion microRNA-155 host gene (MIR155HG) to prevent GVHD. Methods: Using CRISPR/Cas9 genome editing, targeted exon 3 (Ex3) MIR155HG. Genomic was evaluated PCR and RNA expression mature miR-155 qRT-PCR. In xenogeneic acute GVHD experiments, NSG mice were irradiated transplanted with Ex3 edited or control assess functionality vivo. Histological analysis target tissues (skin liver) performed at end study. Weekly flow cytometric evaluation human peripheral blood performed. MOLM-13 leukemic cultured CD8 +effector for 72 hours cytotoxic function assessed CD107a degranulation, intracellular IFN-γ production death cytometry. Results: confirmed resulting downregulation expression. Recipients deleted display lower clinical scores, average histopathological scores significantly improved survival compared receiving cells. displayed potent CTL comparable death. Conclusion: protects against development lethal while maintaining beneficial anti-leukemic effect. Our studies present editing as a novel strategy Funding this project provided through The Ohio State University Leukemia Research Program PR RG, NIH R01CA252469 PR, T32CA090223 fellowship KB.