CRISPR-Edited HLA Donor Liver Transplant to Reduce Rejection

Organ transplant rejection is primarily driven by immune recognition of donor HLA (human leukocyte antigen) molecules as foreign. Mismatches in HLA-A and HLA-B (class I) in particular are strongly immunogenic and can provoke T-cell mediated graft rejection. HLA class II molecules (HLA-DR, DQ, DP), expressed on donor antigen-presenting cells, can also activate CD4⁺ T cells and contribute to rejection. Current therapy relies on immunosuppressive drugs, which carry significant risks. Preclinical research has shown that genetically "erasing" HLA molecules from donor cells can blunt immune responses: for example, cells with HLA-A, HLA-B, and HLA-DR knocked out via CRISPR elicited little to no T cell proliferation in vitro, indicating greatly reduced immunogenicity. Similarly, in xenotransplant models, triple knockout of genes (including class I and the class II regulator CIITA) in donor animals significantly weakened human T-cell activation and prolonged graft survival. These findings provide a strong rationale that an HLA-edited donor organ could evade the human immune system to a large extent, potentially reducing or delaying rejection. Gene-Editing of Donor Liver Ex Vivo: In this trial, deceased-donor livers will undergo ex vivo CRISPR-Cas9 genome editing prior to transplantation. The editing targets are HLA-A and HLA-B (to eliminate the major class I alloantigens) and CIITA (class II transactivator, whose knockout abolishes HLA-DR/DQ/DP expression on donor cells). By knocking out HLA-A and -B, while leaving HLA-C expression intact, the goal is to remove the most immunogenic class I molecules yet maintain some HLA presence to mitigate natural killer cell "missing-self" responses. Disabling CIITA will prevent expression of HLA class II proteins, thus reducing CD4⁺ T cell activation against the graft. The CRISPR editing is performed during machine perfusion of the donor liver (a period in which the organ is kept alive outside the body). A CRISPR-Cas9 ribonucleoprotein (Cas9 enzyme complexed with guide RNAs for HLA-A, HLA-B, and CIITA) is delivered into the liver tissue through the perfusion circuit. Editing takes place ex vivo, avoiding direct in vivo gene therapy to the recipient. Before transplantation, the graft is assessed for successful gene knockout (for example, by biopsy immunostaining or flow cytometry to confirm absence of HLA-A/B/DR on the cell surface). Only livers with confirmed high-efficiency editing (e.g. \>90% target gene disruption) are used for transplant to ensure maximal immune-evasion benefit.