What Have We Learned From In Vitro Studies About Pig-to-primate Organ Transplantation?

Abstract

Natural preformed and de novo antibodies against pig antigens are a major cause of pig xenograft rejection in nonhuman primates (NHPs). In vivo studies in pig-to-NHP models are time consuming. In vitro assays, for example, antibody binding to pig cells, complement-dependent cytotoxicity assays, provide valuable information quickly and inexpensively. Using in vitro assays for several years, it has been documented that (1) during the first year of life, humans and NHPs develop anti-wild-type pig antibodies, but humans develop no or minimal antibody to triple-knockout (TKO) pig cells. (2) Some adult humans have no or minimal antibodies to TKO pig cells and are therefore unlikely to rapidly reject a TKO organ, particularly if the organ also expresses human "protective" proteins. (3) There is good correlation between immunoglobulin (Ig)M (but not IgG) binding and complement injury. (4) All Old World NHPs develop antibodies to TKO pig cells and are not optimal recipients of TKO organs. ( 5) galactosyltransferase gene-knockout/β4GalNT2KO pigs are preferred for Old World NHPs. ( 6) Humans develop anti-pig IgE and IgA antibodies against pig cells, but their role remains uncertain. ( 7) In a small percentage of allosensitized humans, antibodies that cross-react with swine leukocyte antigens may be detrimental to a pig organ xenograft. (8) Prior sensitization to pig antigens is unlikely to be detrimental to a subsequent allograft. ( 9) Deletion of expression of Gal and Neu5Gc is associated with a reduction in the T-cell response to pig cells. All of these valuable observations have largely predicted the results of in vivo studies.

In vitro and in vivo immune assessments of genetically‐engineered pig skin grafts in New World (squirrel) monkeys

“…Thus, removing xenoantigens (such as Neu5Gc and Sda) in pig skin might also contribute to prolonged pig skin xenograft survival in squirrel monkeys due to reduced immunogenicity. Notably, it is worth mentioning that a Gal‐associated immune response is not expected in this model as Gal is expressed in squirrel monkeys (i.e., New World monkeys) 22,27 . Nonetheless, we posit that the knockout of all three pig antigens could benefit human recipients 21,22 …”

“…Squirrel monkeys, a New World monkey, were selected as recipients because many of them, like many humans, do not make natural antibodies against TKO pigs, 22,27 whereas all Old World monkeys (e.g., baboons, cynomolgus, and rhesus monkeys) do 21,27 . Selection of the monkeys for pig skin xenotransplantation was based on those with low levels of anti‐pig IgM and IgG antibodies and serum complement‐dependent‐cytotoxicity to TKO pig peripheral blood mononuclear cells (PBMCs) pre‐transplantation, although cytotoxicity varied considerably (Table 2).…”

“…Notably, it is worth mentioning that a Gal-associated immune response is not expected in this model as Gal is expressed in squirrel monkeys (i.e., New World monkeys). 22,27 Nonetheless, we posit that the knockout of all three pig antigens could benefit human recipients. 21,22 In summary, as anticipated, monkey recipients of skin autografts (Group 1) did not become sensitized to monkey or pig antigens, all grafts remaining fully or partially viable for the 28-day period of follow-up.…”

Xenografts Show Signs of Concentric Hypertrophy and Dynamic Left Ventricular Outflow Tract Obstruction After Orthotopic Pig-to-baboon Heart Transplantation

“…9 During the past decade it was possible to dramatically reduce, and even completely inhibit, organ rejection-especially after xenogeneic kidney and heart transplantation-by applying genetically engineered donor animals and refining postoperative immunosuppressive regimens. [10][11][12] However, with resulting longer survival, unexpected organ growth has been reported after kidney 13,14 as well as heart transplantation. 3,4,15 Both intrinsic (determined by donor genetics 16 ) and extrinsic (determined by the recipient) factors such as changes in nutrition, 16 growth factors (growth hormone [GH], insulinlike growth factor 1 [IGF1]), 17 hormones (thyroid hormones, vascular endothelial growth factor, insulin, catecholamines, endothelin, angiotensin), and mechanical stress/strain 18 were held responsible for this excessive growth.…”

Incidence of serum antibodies to xenoantigens on triple‐knockout pig cells in different human groups

“…However, significant differences continue to be observed in the survival times of the recipients. Although the incidence of acute cellular rejection is low (3%), antibody‐mediated rejection continues to be problematic (incidence 45% within the first post‐transplant year), 4 and is the major reason preventing consistent long‐term survival of NHP recipients 5 …”