Blockade of CD40-CD154 interactions during T cell priming has been shown to be a highly effective means of inducing long-term survival of allografts and transplantation tolerance in both murine and non-human primate models. However, the clinical potential of this therapy has yet to be realized, due to the thromboembolic complications seen during clinical trials of an anti-CD154 monoclonal antibody in humans. Thus, development of novel therapeutics with improved safety profiles is imperative for the field of transplantation immunosuppression. We sought to compare the efficacy and mechanism of the most commonly used clone of anti-CD154 antibody, MR-1, with both a non-agonistic anti-CD40 mAb (G2b) and an anti-CD154 domain antibody with a silent Fc region (dAb). We first compared the efficacy of these therapies in prolonging skin graft survival in a fully allogeneic Balb/c to B6 model. Mice treated with CTLA4-Ig alone quickly rejected their grafts (MST = 17 d), whereas treating with MR-1, G2b or dAb in combination with CTLA4-Ig resulted in significant prolongation of graft survival in all groups (MST = 30.5, 33, and 28, respectively). To further investigate the mechanism of action of each of these therapies, we employed a transgenic model wherein OT-I CD8 and OT-II CD4 T cells were adoptively transferred to B6 mice that then received skin grafts that ubiquitously express the OVA protein. Treatment with all three therapies led to a reduction in splenic T cell expansion compared to untreated animals (Untreated 1.64±0.55%, MR-1 0.50±0.14%, G2b 0.59±0.16%, dAb 0.62±0.23%, p<0.05 in all groups). Functionally, treatment significantly blunted the alloreactive response, as both percentages and total numbers of interferon gamma-producing CD8 T cells in the spleen were reduced in all treatment groups (Untreated 31.05±5.68%, MR-1 10.91±4.59, G2b 14.26±4.46, dAbs 12.66±7.80, p<0.05). In addition, all treatments led to a significant increase in antigen-specific iTreg conversion (Untreated 2.13±1.11, MR-1 12.16±2.33, G2b 7.79±4.32, dAb 9.00±3.43, p<0.05), which has been shown by our group and others to be a crucial step for inducing tolerance with CD40-CD154 blockade. Taken together, these data suggest the possibility that alternative means of blocking CD40-CD154 interactions may provide a safer and equally efficacious way to induce tolerance in transplantation.

Suri, A.: Employee, BMS. Nadler, S.: Employee, BMS. Ford, M.: Grant/Research Support, BMS.

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