Multigene-Edited Pig-to-Rhesus-Macaque Liver Xenotransplantation: Multifaceted Profiling-of-Post-transplant Phenotypes.

Abstract

The shortage of donor livers motivates the development of pig-to-non-human-primate liver xenotransplantation. While gene-editing-mediated xenoantigen knockout reliably prevents hyperacute rejection (HAR), nevertheless, the integrated in-vivo phenotype and coagulation-associated complications of multi-gene-edited porcine livers remain poorly characterized. To address this knowledge gap, we established orthotopic auxiliary partial liver xenotransplantation in non-human primates using 6-, 8-, and 10-gene-edited porcine donors, and performed multi-modal longitudinal phenotyping covering graft function assessment, immune-coagulation profiling, PERV/PCMV DNA screening, as well as mechanistic analyses of graft injury. Consistent with prior reports, all three transplantations succeeded; xenoantigen knockout prevented HAR, and grafts maintained basal function with only mild sinusoidal congestion and limited complement-antibody deposition. Strikingly, every recipient developed severe thrombocytopenia, graft thrombosis, and thrombotic microangiopathy, which ultimately culminated in graft dysfunction. Further analyses revealed that aberrant endothelial ASGR1 and vWF expression together with porcine macrophage-mediated platelet clearance constituted principal drivers of platelet depletion; meanwhile, PERV and PCMV DNA were undetectable in recipient tissues. Collectively, although xenoantigen knockout effectively averts HAR, irreversible coagulation-cascade activation, thrombosis and consumptive coagulopathy still represent the principal barriers to long-term xenograft survival even in the setting of donor-expressed human anticoagulant transgenes hTBM, hCD39, and hEPCR. These findings provide valuable insights for future donor-engineering and complication-intervention strategies.