Introduction: Cardiac xenotransplantation is emerging as a highly promising strategy for addressing terminal heart diseases, with the potential to provide a solution to the global shortage of donor organs. Significant progress has been made in overcoming hyperacute rejection, a major hurdle in transitioning from preclinical to clinical application, including genetic modification of porcine donors, improved preservation techniques, and novel immunosuppressive regimens. In allotransplantation, pretransplant crossmatching is essential for identifying compatible donor-recipient pairs. Applied to xenotransplantation, crossmatching enables detection of donor-specific antibodies (DSA), assessing compatibility between porcine donors and baboon or human recipients.
Methods: This retrospective study evaluates preclinical trials in which porcine hearts were transplanted into baboons. Standard crossmatching techniques - complement-dependent cytotoxicity (CDC) and fluorescence-activated cell sorting (FACS), both considered gold standard - were adapted from clinical allotransplantation protocols to assess donor-recipient compatibility in the xenogeneic setting. Peripheral blood mononuclear cells (PBMCs) from 25 triple genetically modified German Landrace hybrid pigs and sera from corresponding baboon recipients are analyzed via FACS to detect anti-pig DSAs. Preliminary optimization included testing different media for PBMC thawing, evaluating various secondary antibodies and their optimal dilutions by titration, and cytokine stimulation with porcine IL-2 to improve cell viability in cases of sample degradation due to long-term storage.
Results: FACS crossmatch results demonstrate the technical feasibility of the adapted protocol and its ability to detect DSAs in the xenotransplantation context. A systematic evaluation is currently being conducted to investigate a potential correlation between crossmatch results and post-transplantation clinical parameters such as graft survival and signs of rejection. We aim to demonstrate that histocompatibility testing can serve as a predictive tool for transplant outcomes and for follow-up monitoring to identify de novo xenospecific antibodies potentially leading to antibody mediated rejection.
Conclusions: Although our previous preclinical trials in cardiac xenotransplantation were performed without pretransplant screening for preformed DSAs, crossmatching may serve as a valuable additional tool for donor–recipient selection and further improving outcome. In cardiac allotransplantation, crossmatching is a mandatory step to assess immunologic compatibility and reduce the risk of rejection. Implementing this approach to xenotransplantation with consideration of the findings from the retrospective study could enhance the safety and efficacy of future clinical procedures. Ongoing analyses aim to determine whether crossmatch results correlate with clinical outcomes, potentially enabling more informed donor selection in xenotransplantation.