Search

Heart transplantation is the only cure for patients with terminal cardiac failure, but the supply of allogeneic donor organs falls far short of the clinical need1–3. Xenotransplantation of genetically modified pig hearts has been discussed as a potential alternative4. Genetically multi-modified pig hearts that lack galactose-α1,3-galactose epitopes (α1,3-galactosyltransferase knockout) and express a human membrane cofactor protein (CD46) and human thrombomodulin have survived for up to 945 days after heterotopic abdominal transplantation in baboons5. This model demonstrated long-term acceptance of discordant xenografts with safe immunosuppression but did not predict their life-supporting function. Despite 25 years of extensive research, the maximum survival of a baboon after heart replacement with a porcine xenograft was only 57 days and this was achieved, to our knowledge, only once6. Here we show that α1,3-galactosyltransferase-knockout pig hearts that express human CD46 and thrombomodulin require non-ischaemic preservation with continuous perfusion and control of post-transplantation growth to ensure long-term orthotopic function of the xenograft in baboons, the most stringent preclinical xenotransplantation model. Consistent life-supporting function of xenografted hearts for up to 195 days is a milestone on the way to clinical cardiac xenotransplantation7.

In this Letter, Mayuko Kurome and Valeri Zakhartchenko have been added to the author list (affiliated with Institute of Molecular Animal Breeding and Biotechnology, Gene Center, LMU Munich, Munich, Germany). The author list and 'Author contributions' section have been corrected online; see accompanying Amendment.

Background As a step towards clinical cardiac xenotransplantation, our experimental heterotopic intrathoracic xenotransplantation model offers a beating and ejecting donor heart while retaining the recipient′s native organ as a backup in case of graft failure. Clinically applicable immunosuppressive regimens (IS) were investigated first, then treatments known to be effective in hypersensitized patients or those with recalcitrant rejection reactions. Methods Consecutive experiments were carried out between 2009 and 2013. Twenty-one genetically modified pigs (GGTA1-knockout/hCD46/± thrombomodulin, in one case HLA-E instead) were used as donors. In all experiments, two cycles of immunoabsorption reduced preformed antibodies. Recipient baboons were divided into two groups according to IS regimen: In group one (n = 10), pre-treatment started either one (anti-CD20) or four weeks (anti-CD20 plus the proteasome inhibitor bortezomib) prior to transplantation. The extended conventional (as for allotransplantation) immunosuppressive maintenance regimen included anti-thymocyte globuline, tacrolimus, mycophenolate mofetil, methylprednisolone and weekly anti-CD20. In group two (n = 11), myeloablative pre-treatment as in multiple myeloma patients (long and short regimens) was added to extended conventional IS; postoperative total thoracic and abdominal lymphoid irradiation (TLI; single dose of 600 cGY) was used to further reduce antibody-producing cells. Results In the perioperative course, the surgical technique was safely applied: 19 baboons were weaned off extracorporeal circulation and 17 extubated. Nine animals were lost in the early postoperative course due to causes unrelated to surgical technique or IS regimen. Excluding these early failures, median graft survival times of group 1 and 2 were 18.5 (12–50) days and 16 (7–35) days. Necropsy examination of group 1 donor organs revealed hypertrophy of the left ventricular wall in the six longer-lasting grafts; myocardial histology confirmed pre-clinical suspicion of humoral rejection, which was not inhibited by the extended conventional IS including intensified treatments, and signs of thrombotic microangiopathy. Grafts of group 2 presented with only mild-to-moderate features of humoral rejection and thrombotic microangiopathy, except in one case of delayed rejection on day 17. The other experiments in this group were terminated because of untreatable pulmonary oedema, recurring ventricular fibrillation, Aspergillus sepsis, as well as a combination of a large donor organ and late toxic side effects due to TLI. Conclusions Longer-term results were difficult to achieve in this model due to the IS regimens used. However, we conclude that heterotopic intrathoracic heart transplantation may be an option for clinical xenotransplantation.

Objectives: New immunologic approaches are needed to overcome xenogeneic rejection to make clinical cardiac xenotransplantation possible. Anti-CD40-antibody reduces the adaptive immune response by inhibiting costimulation between T-cells and antigen presenting cells respectively B-cells. We investigated the effect on different peripheral cell lines during the first five days after transplantation. Methods: Eight baboons underwent heterotopic thoracic cardiac xenotransplantation; double and triple (Gal-KO/hCD46/ ± hTM) transgenic pigs were used as donors. Treatment consisted of anti-CD20-antibody, MMF, tacrolimus, cyclophosphamide and ATG induction (dosage is listed in the table below). The last two animals additionally received the new anti-CD40-antibody (2C10R4). Lymphocytes, neutrophils, monocytes and thrombocytes were measured daily. Results: The new therapeutic regimen used caused a clear decrease of neutrophils and lymphocytes compared with previous experiments. Additionally, the dose of cyclophosphamide, tacrolimus and MMF could be markedly reduced. This led to less side effects of the basic immunosuppression. No adverse effects of the new antibody, especially no thrombotic complications as described in other costimulation treatments occurred. Conclusions concerning graft survival are not possible at this point (one animal developed a pulmonary edema on day 13, one animal a sepsis on day 35; mean survival of previous animals was 18 days). Conclusions: We could demonstrate that costimulation blockade with anti-CD40-antibody was efficient and the basic immunosuppression could be reduced. As a result the new therapeutic regimen had less toxic side effects compared with our control.

Immunosuppression Using CD40 Costimulation Blockade in a Preclinical Cardiac Xenotransplantation Model J. Abicht ,1 T. Mayr,2 S. Guethoff,2 F. Werner,1 I. Lutzmann,2 M.C. Langenmayer,3 E. Wolf,4 D. Ayares,5 K. Reimann,6 B. Reichart,7 P. Brenner.2 1Department of Anaestesiology (LMU), Munich, Germany; 2Department of Cardiac Surgery (LMU), Munich, Germany; 3Department of Veterinary Pathology (LMU), Munich, Germany; 4Department of Molecular Animal Breeding and Biotechnology(LMU), Munich, Germany; 5Revivicor Inc., Blacksburg, VA; 6MassBiologics, University of Massachusetts, Boston, MA; 7Walter-Brendel-Centre (LMU), Munich, Germany.