Milestones on the path to clinical pig organ xenotransplantation

Abstract

Progress in pig organ xenotransplantation has been made largely through (1) genetic engineering of the organ-source pig to protect its tissues from the human innate immune response, and (2) development of an immunosuppressive regimen based on blockade of the CD40/CD154 costimulation pathway to prevent the adaptive immune response. In the 1980s, after transplantation into nonhuman primates (NHPs), wild-type (genetically unmodified) pig organs were rejected within minutes or hours. In the 1990s, organs from pigs expressing a human complement-regulatory protein (CD55) transplanted into NHPs receiving intensive conventional immunosuppressive therapy functioned for days or weeks. When costimulation blockade was introduced in 2000, the adaptive immune response was suppressed more readily. The identification of galactose-α1,3-galactose as the major antigen target for human and NHP anti-pig antibodies in 1991 allowed for deletion of expression of galactose-α1,3-galactose in 2003, extending pig graft survival for up to 6 months. Subsequent gene editing to overcome molecular incompatibilities between the pig and primate coagulation systems proved additionally beneficial. The identification of 2 further pig carbohydrate xenoantigens allowed the production of 'triple-knockout' pigs that are preferred for clinical organ transplantation. These combined advances enabled the first clinical pig heart transplant to be performed and opened the door to formal clinical trials.

Keywords: clinical trial; genetically engineered; heart; kidney; nonhuman primate; pig; xenotransplantation.

Figure 1.

Timeline of advances in pig genetic engineering (bottom) and in immunological aspects of pig-to-nonhuman primate organ transplantation (top).

Figure 2.

Correlation between human serum antibody binding to pig red blood cells (RBCs) by relative geometric mean and age. Human serum (A) IgM and (B) IgG antibody binding to wild-type pig RBCs (top) and to triple-knockout pig RBCs (bottom). The dotted lines indicate no IgM or IgG binding. Note the great difference in the scale on the y-axis between A and B. (Reproduced with permission from Li et al).

Figure 3.

Steps involved in somatic cell nuclear transfer. (Reprinted with permission from Eyestone et al).

Figure 4.

GTKO pig kidney survival in baboons receiving US Food and Drug Administration–approved immunosuppressive agents (Group A, in red) was much shorter than in those receiving an anti-CD40 monoclonal antibody–based regimen (Group B, in black). GTKO, α1,3-galactosyltransferase gene knockout. (Reproduced with permission from Yamamoto et al).

Figure 5.

Human (top) and Old World monkey (OWM) (bottom) IgM (left) and IgG (middle) binding and complement–dependent cytotoxicity (CDC, at 25% serum concentration) (right) to WT, GTKO, and TKO pig peripheral blood mononuclear cells (PBMCs). Results are expressed as mean ± SEM. (*P < 0.05, **P < 0.01, N.S. ¼ not significant). On the y-axis, the dotted line represents cut-off value of binding (relative geometric mean: IgM 1.2, IgG 1.1), below which there is no binding. For CDC on the y-axis, the dotted line represents cut-off value of cytotoxicity (6.4%), below which there is no cytotoxicity. Note the difference in scale on the y-axis between IgM and IgG. GTKO, α1,3-galactosyltransferase gene knockout; TKO, triple knockout; WT, wild-type.

Figure 6.

Rejection-free survival of GTKO pig kidneys in baboons (Group 1, in black) was significantly longer than that of TKO pig kidneys (Group 2, in red). GTKO, α1,3-galactosyltransferase gene knockout; TKO, triple knockout. (Reproduced with permission from Iwase et al).

Figure 7.

Correlation of human (n ¼ 9) and baboon (n ¼ 72) serum IgM (left) and IgG (right) antibody binding with serum complement–dependent cytotoxicity (CDC, at 50% serum concentration) to TKO pPBMCs. In both humans and baboons, there was a significant increase in cytotoxicity as IgM and IgG antibody binding to TKO pPBMCs increased. In baboons, however, cytotoxicity was high whether IgM binding was high (e.g., 80% cytotoxicity at a rGM of 8) or relatively lower (e.g., 75% at a rGM of 2). **P < 0.01. pPBMC, pig peripheral blood mononuclear cell; rGM, relative geometric mean; TKO, triple knockout. (Reprinted with permission from Yamamoto et al).