Costimulatory Blockade to Overcome Humoral Immunity

Sensitization to major histocompatibility complex (MHC) antigens affects 13-30% of patients awaiting kidney transplantation. Of these, up to 20% are highly sensitized (panel-reactive antibody >80%). This condition comes with an increased likelihood of graft loss after transplantation. Strategies that decrease alloantibody have showed to improve patient survival significantly, nevertheless, the cost and risks of plasmapheresis and other current methods are considerable, and their efficacy is limited. Furthermore about 20% of renal transplant patients develop donor- specific antibodies (DSA) after transplantation. These are strongly associated with chronic rejection but also with acute and subclinical rejection, contributing to graft loss in these settings. There is currently no accepted method of treating such patients to reliably remove DSA. In numerous murine and preclinical models, the CD40/154 and the CD28 costimulatory pathways are effectively blocked for T cell based tolerance. The therapeutic potential of this therapy for humoral immunity has not been exploited. Our aim is to determine the efficacy of blocking the CD40/154 and the CD28 costimulatory pathways as desensitization strategy and to assess the impact of these therapies on protective immunity in a sensitized non- human primate (NHP) model. Briefly, the model uses monkeys sensitized by a previously rejected renal allograft, and ultimately we aim to achieve immunologic unresponsiveness to a second kidney transplanted from the same donor. We will use state of the art methods developed by us to measure not only T and B cell sensitization and desensitization, but also will measure the affect of treatment on the protective immune response that must remain intact for avoidance of infection. We will build upon lessons learned from previous work that focused on the T and B cell response in monkeys developing de novo antibody after renal transplantation. In an earlier rigorous NHP model CD28 blockade with Belatacept effectively prevented alloantibody from developing. We now propose to use a difficult but yet more clinically relevant NHP model to evaluate strategies to desensitize the host. The immunologic tools to measure allospecific B cell, plasma cell, and antibody responses in transplantation are relevant to a host of organ transplant recipients and represent a novel contribution to this area. The need for better approaches to treat alloantibody in transplant patients - awaiting transplantation or after transplantation - is well recognized and this would constitute an important advance in the field.

In this project we 1) have established a sustainable, reproducible, and measurable model for the immunological sensitization in the rhesus monkey, 2) have for the first time tested costimulatory blockers for their potential applicability to desensitization and 3) have found signs for improved success of transplants after desensitization with new costimulatory blockers" and the proteasome inhibitor Bortezomib compared to animals without desensitization. Project summary: Costimulation is a necessary step in the activation of immune cells, which leads to their effective response to foreign features. In the case of transplantation this response is directed against the new organ and damages it. To be sensitized against antigens of potential organ donors decreases both, the chance to find a suitable organ and the expected success after transplantation. Many people have developed antibodies to potential donors, be it that they had blood transfusions, pregnancies or previous transplants.Currently these patients undergo expensive and side-effect-prone physical and pharmacological desensitization therapies, but long-term outcomes are inferior to non-sensitized patients. The scope of this project was to evaluate new substances that block costimulation and their potential to reduce antibody-producing cells and thus donor-specific antibodies.To study these effects we established a rhesus monkey model that included sensitization via skin grafting from genetically disparate animals.Then the resulting antibodies were to be reduced by therapy with costimulatory blockers targeting CD28/B7- and CD40/CD154 receptor pairs and the proteasome inhibitor bortezomib, which can directly reduce antibody-forming plasma cells.We then transplanted a kidney from their respective skin donors into desensitized and not-desensitized animals and at the same time removed the animals own kidneys.After the development of a sensitization model in the rhesus monkey and identification of the most effective drug combination for desensitization we observed an improved course of renal function and histological findings in the treated animals after transplantation.