Allograft Survival In Primates Research Articles

Immunological evaluation of combination therapy with tacrolimus and sirolimus on long-term allograft survival in nonhuman primates

We reported that a 60-day course of combination therapy with tacrolimus and sirolimus induced long-term survival of renal allograft after withdrawal of immunosuppressants in Vervet monkeys. In the present study, the mechanism of drug-induced allograft survival was evaluated via Th1/Th2 cytokines, apoptosis and MLC activity in primates. Materials and methods Cytokines were evaluated by ELISA. MLR and CTL assays were performed by incorporation of 72 hours 3H-TdR and 4 hours 51Cr release assay. Results A 60-day course of tacrolimus with sirolimus resulted in long-term survival of kidney allografts. (67% > 100 days) without intermittent acute rejection. Low sensitivity to MLR was seen in long-term renal allograft survival among monkeys treated with tacrolimus and sirolimus. Increased levels of CD3 +CD8 +, CD3 +/CD56 + NKT cells and CD86 +CD8 −CD11 + dendritic cells were observed. A population of high expression of CD4 +FasL + was detected. In addition, the concentrations of IL-2 and IFN-γ from long-term allograft surviving monkeys was not significantly increased, rather a late phase dominance of Th2, IL-4, IL-10, and TGF-β was found correlated with long-term survival of recipients. In conclusion, the mechanism of tacrolimus and sirolimus induced long-term allograft survival in primates relates to up-regulated FasL expression, NKT cells and dendritic cells, with downregulation of MLR sensitivity. It is also associated with late-dominant expression of Th2 cytokines.

Effect of Mediators of Innate Immunity and Inflammation on CD8+ Veto Cells

Methods of allotolerance induction that are successful in mice often fail in primates. Activators of innate immunity derived from microbial pathogens, which are present in primate populations but not in pathogen-free mouse colonies, have been shown to overcome tolerance induction by co-stimulatory blockade or regulatory T cells. Unlike other strategies, veto cells promote long-term allograft survival in primates. Accordingly, the authors hypothesized that veto cells are resistant to the effects of innate immune activation. Using mixed lymphocyte reactions as a model of alloimmunization, the authors assessed the effect of inflammatory mediators on veto cell activity. RESULTS.: The authors demonstrate that the activity of veto cells is unaltered by lipopolysaccharide, double-stranded RNA, or interferon-gamma. Tumor necrosis factor-alpha modestly inhibited the veto effect, but only when veto cells were limiting. These findings may help to explain why veto cell-based therapies are more effective than other approaches in primate populations.

Effects of Combined Treatment with CD25- and CD154-Specific Monoclonal Antibodies in Non-Human Primate Allotransplantation

The CD154-specific monoclonal antibody (Mab) hu5c8 greatly prolongs allograft survival in primates. The CD25-specific Mab daclizumab has not, to date, been paired with hu5c8. We evaluated the effects of hu5c8 in vitro, alone and in combination with daclizumab on rhesus-mixed lymphocyte reactions (MLRs). We then evaluated therapy with hu5c8 and daclizumab in four monkey renal allograft recipients compared with monkeys untreated or contemporaneously treated with hu5c8 alone. Lymphocyte proliferation in MLR was reduced by both daclizumab and hu5c8, and their combined effects were additive. Rejection-free allograft survival in monkeys treated with both hu5c8 and daclizumab (74–479 days) was not significantly better than animals treated with hu5c8 alone (257–587 days), and one combined therapy animal rejected while still on hu5c8 therapy, a condition not typically seen with hu5c8 monotherapy. Although daclizumab and hu5c8 are additively effective in MLR, they do not appear to be synergistic in vivo in rhesus monkeys.

Treatment with the humanized CD154-specific monoclonal antibody, hu5C8, prevents acute rejection of primary skin allografts in nonhuman primates.

Allogeneic skin transplantation remains a rigorous test of any immune intervention designed to prevent allograft rejection. To date, no single, clinically available immunosuppressant has been reported to induce long-term primary skin allograft survival in primates. We have previously shown that treatment with the humanized CD154-specific monoclonal antibody, humanized 5C8 (hu5C8), induces long-term renal allograft survival in nonhuman primates. In this study, we evaluated the efficacy of hu5C8 in preventing primary skin allograft rejection in rhesus monkeys. Ten rhesus monkeys were transplanted with full-thickness skin allografts mismatched at both class I and class II major histocompatibility loci. Of these, two were given no treatment, five were treated with hu5C8 alone, and three received hu5C8 combined with whole blood donor-specific transfusion (DST). All recipients also received skin autografts for comparison. Animals were followed by inspection, serial biopsy, mixed lymphocyte culture, and alloantibody determination. Treatment with hu5C8 alone or hu5C8 plus DST greatly prolonged allograft survival. Rejection occurred in the untreated group within 7 days. Mean allograft survival in the monotherapy hu5C8 group was >236 days and in the DST group was >202 days; these differences were not significant. Rejection eventually occurred in most animals. Allograft survival was not correlated with the development of T cell hyporesponsiveness in mixed lymphocyte culture. Rejection was not predicted by the development of donor-specific alloantibody. These results show that treatment with the CD154-specific monoclonal antibody, hu5C8, greatly delays the onset of acute skin allograft rejection.

Anti-CD45RB Monoclonal Antibody-Mediated Transplantation Tolerance

Currently, lifelong immunosuppression is required for organ transplant recipients. The majority of transplant recipients will eventually develop chronic rejection with resultant graft loss, despite treatment with powerful immunosuppressive agents. These agents are also associated with numerous toxicities including reduced immunity against infection and malignancy. Therefore, the central goal in transplant science is to devise tolerance strategies in an attempt to establish a state of prolonged non-reactivity against the allograft, accompanied with preservation of an intact immune system. Although predictable tolerance induction has been elusive, we found that short course of the novel immunomodulatory agent, anti-CD45RB monoclonal antibody, leads to indefinite acceptance of renal allografts in mice, and has been shown to markedly prolong allograft survival in primates. We review the current state of development of this antibody, and the progress made in defining its mechanism of action.

Prolongation of Cardiac Graft Survival with Anti-CD4Ig Plus hCTLA4Ig in Primates

Background.The aim of this study was to determine whether the use of combined immunotherapy with a brief course of humanized anti-CD4Ig and hCTLA4Ig would prolong heterotopic cardiac allograft survival in primates (rhesus monkeys). This model was based on work in “high responder” rats where a brief course of depletive anti-CD4 mAb plus hCTLA4Ig was successful in inducing transplantation tolerance.Methods.Heterotopic cardiac transplants were performed in rhesus recipients. Donor/recipient pairs between groups were confirmed to be reactive prior to transplantation by MLR matching. Humanized anti-CD4Ig, a recently developed anti-CD4 mAb, was given at a dose of 20 mg/kg i.v. on days −3, −2, −1, and 0. hCTLA4Ig was administered at 6 mg/kg/dose i.v. on days 0 and 2 for the first recipient and days 0, 2, 4, and 6 for the second recipient. No further immunosuppression was administered. The treated (n= 2) or untreated (n= 5) recipients were followed for graft function by daily palpitation.Results.Treatment with anti-CD4Ig plus hCTLA4Ig resulted in a significant prolongation of heart graft survival (42 days for the first recipient and 52 days for the second recipient) compared to untreated recipients (7 days × 4, 11 days × 1). FACS analysis demonstrated CD4 depletion of anti-CD4 treated animals to <2% on posttransplant day 1. The CD4+T cells gradually repopulated to 50–70% pretransplant levels just prior to rejection. No adverse responses (fever, tachypnea, tachycardia, infections) were observed.Conclusions.These are the first results demonstrating that a brief course of combined specific induction immunotherapy with humanized anti-CD4Ig plus hCTLA4Ig, in the absence of adjuvant posttransplant immunosuppression, was well tolerated and resulted in marked prolongation of cardiac allograft survival in primates.

Humanized antibody directed to the IL-2 receptor beta-chain prolongs primate cardiac allograft survival.

IL-2Rs are expressed by T cells activated in response to foreign histocompatibility Ags but not by normal cells. This difference in IL-2R expression is exploited by blockade of IL-2Rs to achieve immunosuppression. High affinity IL-2Rs involve three subunits, IL-2R alpha, IL-2R beta, and IL-2R gamma. Murine Mik beta 1, a mAb that blocks IL-2 binding to IL-2R beta, was developed as an immunosuppressive agent. There was modest prolongation of cynomolgus cardiac allograft survival in animals treated with murine Mik beta 1 (mean survival 11.8 +/- 1.6 days compared with 8.2 +/- 0.4 days in untreated animals; p = 0.06). However, murine Mik beta 1 is ineffective in recruiting primate effector cells and is neutralized by monkey Abs directed toward the infused Ab. To circumvent these limitations, a humanized form of Mik beta 1, which is a largely human IgG1k Ab, except that murine hypervariable regions are retained, was developed. In vivo plasma survival of humanized Mik beta 1 was threefold longer than simultaneously administered murine Mik beta 1 (terminal t1/2, 104 +/- 10 h vs 37 +/- 2 h). Furthermore, humanized Mik beta 1 manifests Ab-dependent cellular cytotoxicity, an activity that is absent with the parental murine Mik beta 1. Graft survival was significantly prolonged by humanized Mik beta 1 treatment with survivals of 22, 22, 24, 27, 44, and > 300 days (p vs control < 0.01; p vs murine Mik beta 1 < 0.01). Survival was not prolonged further (p > 0.3) by the addition of humanized anti-Tac, which blocks interaction of IL-2 with IL-2R alpha subunits. There was no toxicity attributable to the use of Mik beta 1 Abs. Thus, humanized Mik beta 1 prolonged cardiac allograft survival in primates without toxicity and may be effective as an adjunct to standard immunosuppressive therapy.

Anti-Tac-H, a humanized antibody to the interleukin 2 receptor, prolongs primate cardiac allograft survival.

High-affinity interleukin 2 receptors (IL-2Rs) are expressed by T cells activated in response to foreign histocompatibility antigens but not by normal resting T cells. To exploit this difference in IL-2R expression, anti-Tac-M, a murine monoclonal antibody specific for the IL-2R alpha chain, was used to inhibit organ allograft rejection. However, the use of murine anti-Tac as an immunosuppressive agent was limited by neutralization by human anti-murine antibodies and by weak recruitment of effector functions. To circumvent these difficulties, a humanized antibody to the IL-2R, anti-Tac-H, was prepared. This molecule is human with the exception of the hypervariable segments, which are retained from the mouse. In vivo survival of anti-Tac-H is 2.5-fold longer than simultaneously administered anti-Tac-M (terminal t1/2, 103 hr vs. 38 hr). In addition, anti-Tac-H is less immunogenic than anti-Tac-M when administered to cynomolgus monkeys undergoing heterotopic cardiac allografting. Specifically, all monkeys treated with anti-Tac-M developed measurable anti-anti-Tac-M levels by day 15 (mean onset, 11 days). In contrast, none of the animals receiving anti-Tac-H produced measurable antibodies to this monoclonal antibody before day 33. Finally, there was a prolongation of graft survival in the cynomolgus heterotopic cardiac allograft model in animals receiving anti-Tac. In animals that received anti-Tac-M, the allograft survival was prolonged compared to that of the control group (mean survival, 14 +/- 1.98 days compared to 9.2 +/- 0.48 days; P less than 0.025). Graft survival was further prolonged by anti-Tac-H with a mean survival of 20.0 +/- 0.55 days (compared to controls, P less than 0.001; compared to anti-Tac-M, P less than 0.02). There was no toxicity attributable to the administration of either form of anti-Tac. Thus, anti-Tac-H significantly prolonged allograft survival in primates, without toxic side effects, and may be of value as an adjunct to standard immunosuppressive therapy in humans.

Segmental pancreatic allograft survival in primates treated with total-body or lymphoid irradiation and peroperative blood transfusions.

Segmental pancreatic allograft survival in primates treated with total-body or lymphoid irradiation and peroperative blood transfusions.