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Genetically engineered pigs are thought to be an alternative organ source for patients in end-stage heart failure unable to receive a timely allograft. However, cardiac xenografts exhibit growth and diastolic heart failure within 1 month after transplantation. Grafts function for up to 6 months, but only after administration of temsirolimus and afterload-reducing agents to reduce this growth. In this study we investigated the growth and hemodynamics of growth hormone receptor (GHR) knockout xenografts, without the use of adjuncts to prevent intrinsic graft growth after transplantation. Genetically engineered pig hearts were transplanted orthotopically into weight-matched baboons between 15 and 30 kg, using continuous perfusion preservation before implantation (n = 5). Xenografts included knockout of carbohydrate antigens and knockin of human transgenes for thromboregulation, complement regulation, and inflammation reduction (grafts with intact growth hormone, n = 2). Three grafts contained the additional knockout of GHR (GHR knockout grafts; n = 3). Transthoracic echocardiograms were obtained twice monthly and comprehensively analyzed by a blinded cardiologist. Hemodynamics were measured longitudinally after transplantation. All xenografts demonstrated life-supporting function after transplantation. There was no difference in intrinsic growth, measured using septal and posterior wall thickness and left ventricular mass, on transthoracic echocardiogram out to 1 month in either GHR knockout or GHR intact grafts. However, hypertrophy of the septal and posterior wall was markedly elevated by 2 months post transplantation. There was minimal hypertrophy out to 6 months in GHR knockout grafts. Physiologic mismatch was present in all grafts after transplantation, which is largely independent of growth. Xenografts with GHR knockout show reduced post-transplantation xenograft growth using echocardiography >6 months after transplantation, without the need for other adjuncts. Post-transplantation xenograft growth is a life-limiting phenomenon in recipients after cardiac xenotransplantation. In this study, genetically engineered hearts expressing multiple human transgenes, with or without intact growth hormone receptor, were transplanted orthotopically into weight-matched baboons after continuous perfusion preservation. Post-transplantation xenograft growth was investigated using echocardiography and physiologic mismatch was characterized. Results indicate markedly elevated heart rates and mean arterial pressures experienced by all transplanted xenografts. Growth hormone intact grafts showed marked post-transplantation xenograft growth compared with those with growth hormone receptor knocked out. This study demonstrates reduced xenograft growth after transplantation by using xenografts with multiple human transgenes and growth hormone receptor knockout. Implications include the possible elimination for the need for adjuncts to reduce post-transplantation cardiac xenograft growth. [Display omitted] [ABSTRACT FROM AUTHOR]

We have been testing genetically engineered (GE) pig hearts and optimizing immunosuppression (IS) in non‐human primates (NHPs) since 2005. We demonstrate how we translated this preclinical investigation into a US Food and Drug Administration (FDA)‐approved clinical cardiac xenotransplantation. First, genetically engineered (GE) pig hearts were transplanted into the abdomen of NHP along with IS, which included anti‐CD20 and anti‐CD40‐based co‐stimulation blockade antibodies. We reported 945 days of survival of three gene GE pig hearts in NHPs. Building on this proof‐of‐concept, we tested 3–10 gene‐modified GE pig hearts (in order to improve the immunocompatibility of the xenograft further) in a life‐supporting orthotopic model, but had limited success due to perioperative cardiac xenograft dysfunction (PCXD). With novel non‐ischemic continuous perfusion preservation (NICP), using the XVIVO Heart solution (XHS), life‐supporting survival was extended to 9 months. We approached the FDA under an application for "Expanded Access" (EA), to transplant a GE pig heart in a patient with end‐stage non‐ischemic cardiomyopathy. He was without other therapeutic options and dependent on VA‐ECMO. A team of FDA reviewers reviewed our preclinical research experience and data and allowed us to proceed. This clinical cardiac xenotransplantation was performed, and the patient survived for 60 days, demonstrating the translational preclinical investigation of cardiac xenotransplantation from bench to bedside. The ultimate etiology of graft failure is currently a topic of investigation and lessons learned will progress the field forward. [ABSTRACT FROM AUTHOR]

We report orthotopic (life‐supporting) survival of genetically engineered porcine cardiac xenografts (with six gene modifications) for almost 9 months in baboon recipients. This work builds on our previously reported heterotopic cardiac xenograft (three gene modifications) survival up to 945 days with an anti‐CD40 monoclonal antibody‐based immunosuppression. In this current study, life‐supporting xenografts containing multiple human complement regulatory, thromboregulatory, and anti‐inflammatory proteins, in addition to growth hormone receptor knockout (KO) and carbohydrate antigen KOs, were transplanted in the baboons. Selective "multi‐gene" xenografts demonstrate survival greater than 8 months without the requirement of adjunctive medications and without evidence of abnormal xenograft thickness or rejection. These data demonstrate that selective "multi‐gene" modifications improve cardiac xenograft survival significantly and may be foundational for paving the way to bridge transplantation in humans. [ABSTRACT FROM AUTHOR]

Background: Perioperative cardiac xenograft dysfunction (PCXD) describes a rapidly developing loss of cardiac function after xenotransplantation. PCXD occurs despite genetic modifications to increase compatibility of the heart. We report on the incidence of PCXD using static preservation in ice slush following crystalloid or blood-based cardioplegia versus continuous cold perfusion with XVIVO© heart solution (XHS) based cardioplegia. Methods: Baboons were weight matched to genetically engineered swine heart donors. Cardioplegia volume was 30 cc/kg by donor weight, with del Nido cardioplegia and the addition of 25% by volume of donor whole blood. Continuous perfusion was performed using an XVIVO © Perfusion system with XHS to which baboon RBCs were added. Results: PCXD was observed in 5/8 that were preserved with crystalloid cardioplegia followed by traditional cold, static storage on ice. By comparison, when blood cardioplegia was used followed by cold, static storage, PCXD occurred in 1/3 hearts and only in 1/5 hearts that were induced with XHS blood cardioplegia followed by continuous perfusion. Survival averaged 17 hours in those with traditional preservation and storage, followed by 11.47 days and 15.03 days using blood cardioplegia and XHS+continuous preservation, respectively. Traditional preservation resulted in more inotropic support and higher average peak serum lactate 14.3±1.7 mmol/L compared to blood cardioplegia 3.6±3.0 mmol/L and continuous perfusion 3.5±1.5 mmol/L. Conclusion: Blood cardioplegia induction, alone or followed by XHS perfusion storage, reduced the incidence of PCXD and improved graft function and survival, relative to traditional crystalloid cardioplegia-slush storage alone. [ABSTRACT FROM AUTHOR]

Significant progress in understanding and overcoming cardiac xenograft rejection using a clinically relevant large animal pig-to-baboon model has accelerated in recent years. This advancement is based on improved immune suppression, which attained more effective regulation of B lymphocytes and possibly newer donor genetics. These improvements have enhanced heterotopic cardiac xenograft survival from a few weeks to over 2 years, achieved intrathoracic heterotopic cardiac xenograft survival of 50 days and orthotopic survival of 57 days. This encouraging progress has rekindled interest in xenotransplantation research and refocused efforts on preclinical orthotopic cardiac xenotransplantation. [ABSTRACT FROM AUTHOR]