Anti-C5 Antibody Tesidolumab Reduces Early Antibody-mediated Rejection and Prolongs Survival in Renal Xenotransplantation

Abstract

Objective: Pig-to-primate renal xenotransplantation is plagued by early antibody-mediated graft loss which precludes clinical application of renal xenotransplantation. We evaluated whether temporary complement inhibition with anti-C5 antibody Tesidolumab could minimize the impact of early antibody-mediated rejection in rhesus monkeys receiving pig kidneys receiving costimulatory blockade-based immunosuppression.

Methods: Double (Gal and Sda) and triple xenoantigen (Gal, Sda, and SLA I) pigs were created using CRISPR/Cas. Kidneys from DKO and TKO pigs were transplanted into rhesus monkeys that had the least reactive crossmatches. Recipients received anti-C5 antibody weekly for 70 days, and T cell depletion, anti-CD154, mycophenolic acid, and steroids as baseline immunosuppression (n = 7). Control recipients did not receive anti-C5 therapy (n = 10).

Results: Temporary anti-C5 therapy reduced early graft loss secondary to antibody-mediated rejection and improved graft survival (P < 0.01). Deleting class I MHC (SLA I) in donor pigs did not ameliorate early antibody-mediated rejection (table). Anti-C5 therapy did not allow for the use of tacrolimus instead of anti-CD154 (table), prolonging survival to a maximum of 62 days.

Conclusion: Inhibition of the C5 complement subunit prolongs renal xenotransplant survival in a pig to non-human primate model.

FIGURE 1.

Flow cytometric crossmatch and immunosuppression protocol employed in transplants. (A) IgG flow cytometric MFI and demonstrated for 14 rhesus macaque recipients. (●) represents an individual pre-transplant serum sample tested against DKO cells (GGTA1/B4GalNT2 KO) who later received a DKO porcine kidney. (☐) represents an individual pre-transplant serum sample tested against TKO cells (GGTA1/B4GalNT2/SLA-1 KO) who later received a TKO porcine kidney. All recipients were selected for low pretransplant IgG antibody binding, but none achieved negative crossmatch. (B) Immunosuppression regimen used in all of the recipients.

FIGURE 2.

Analysis of grafts surviving to 557 days. (A) Kaplan-Meier curve analysis of recipients who received genetically engineered porcine kidneys with or without Tesidolumab included in the immunosuppression regimen. (B) H&E and confocal microscope analysis of rejected xenografts. Confocal microscopy showing deposition of IgM, and C4d in the control and tesidolumab treated recipients. Tesidoluman treated recipients had minimal histological injury while control kindeys had parenchymal hemorrhage and thrombotic microangiopathy. Deposition of C5b-9 was reduced on xenografts in tesidolumab treated recipients.

FIGURE 3.

(A) Creatinine is shown to be well controlled until acutely rising at time of graft failure. (B) Serum potassium was well controlled throughout, except during late graft failure due to rejection (C) Early graft failure recipients encounter anemia as shown by decreased hemoglobin levels. Tesidolumab treated recipients-maintained hemoglobin levels at or above normal levels more frequently than control animals for the duration of graft survival.

FIGURE 4.

(A) Immunosuppression regimen including Tacrolimus and Tesidolumab without CD154 monoclonal antibody therapy. (B) Kaplan-Meier curve analysis reveals that a tacrolimus based immunosuppression regimen fails to extend xenograft survival beyond 62 days despite the presence of C5 complement inhibition by tesidolumab. (C) Biopsy taken from explanted kidney at day 62 shows parenchymal hemorrhage and thrombotic microangiopathy, prominent IgG, IgM, C4d, and minimal C5b-9. (D) Creatinine and potassium levels rose above normal values days to weeks before rejection. Hemoglobin values remained relatively normal.