Galactosyl transferase knock‐out pig lungs fail rapidly in baboons. Based on previously identified lung xenograft injury mechanisms, additional expression of human complement and coagulation pathway regulatory proteins, anti‐inflammatory enzymes and self‐recognition receptors, and knock‐down of the β4Gal xenoantigen were tested in various combinations. Transient life‐supporting GalTKO.hCD46 lung function was consistently observed in association with either hEPCR (n = 15), hTBM (n = 4), or hEPCR.hTFPI (n = 11), but the loss of vascular barrier function in the xenograft and systemic inflammation in the recipient typically occurred within 24 h. Co‐expression of hEPCR and hTBM (n = 11) and additionally blocking multiple pro‐inflammatory innate and adaptive immune mechanisms was more consistently associated with survival >1 day, with one recipient surviving for 31 days. Combining targeted genetic modifications to the lung xenograft with selective innate and adaptive immune suppression enables prolonged initial life‐supporting lung function and extends lung xenograft recipient survival, and illustrates residual barriers and candidate treatment strategies that may enable the clinical application of other organ xenografts.

No abstract

The transplantation of organs across species offers the potential to solve the shortage of human organs. While activation of human platelets by human von Willebrand factor (vWF) requires vWF activation by shear stress, contact between human platelets and porcine vWF (pvWF) leads to spontaneous platelet adhesion and activation. This non‐physiologic interaction may contribute to the thrombocytopenia and coagulation pathway dysregulation often associated with xenotransplantation of pig organs in nonhuman primates. Pigs genetically modified to decrease antibody and complement‐dependent rejection (GTKO.hCD46) were engineered to express humanized pvWF (h*pvWF) by replacing a pvWF gene region that encodes the glycoprotein Ib‐binding site with human cDNA orthologs. This modification corrected for non‐physiologic human platelet aggregation on exposure to pig plasma, while preserving in vitro platelet activation by collagen. Organs from pigs with h*pvWF demonstrated reduced platelet sequestration during lung (p ≤ .01) and liver (p ≤ .038 within 4 h) perfusion ex vivo with human blood and after pig‐to‐baboon lung transplantation (p ≤ .007). Residual platelet sequestration and activation were not prevented by the blockade of canonical platelet adhesion pathways. The h*pvWF modification prevents physiologically inappropriate activation of human or baboon platelets by porcine vWF, addressing one cause of the thrombocytopenia and platelet activation observed with xenotransplantation.