This project involves a comprehensive approach toward the design and implementation of a therapy that will allow for solid organ allotransplants to be performed with a reduced requirement for maintenance immunosuppression. Therapies are evaluated in non-human primates (NHPs) and translated into clinical trials. The primary pre-clinical model is the rhesus monkey kidney (or in some cases skin) transplant model. Human studies are performed in patients undergoing kidney transplantation. Study subject outcome (generally rejection or graft survival) and in vitro analysis of material derived from these subjects (allograft biopsy material, peripheral blood cell phenotype, etc) provides feedback for the effectiveness of the therapy and suggests modifications for future study. This being the first year of the laboratory as an intramural entity, we have established the NHP transplant model at the NIH and initiated trials. The humanized CD154-specific monoclonal antibody IDEC-131 has been evaluated in NHPs. We have demonstrated that this antibody can prevent skin and kidney allograft rejection. The addition of the antiproliferative drug rapamycin improves the effect and graft survival is further improved by pre-transplant donor-specific blood transfusion (DST). The therapy appears non-toxic and clinically applicable. Long-term effects are under evaluation. The combined therapy of IDEC-131, rapamycin and DST is uniformly successful. Animals are now being withdrawn from therapy to determine the duration of the anti-rejection effect with specific reference to whether donor-specific tolerance is induced with this therapy. A clinical trial is under design. Human trials with the humanized CD52-specific monoclonal antibody Campath-1H have continued. We were the first North American group to evaluate this agent. Earlier studies have demonstrated that this agent, when used at the time of kidney transplantation greatly reduces the need for subsequent maintenance immunosuppression. Evaluation of patients maintained solely with the immunosuppressive drug rapamycin is underway. Many of these patients have developed donor-specific hyporesponsiveness in vitro. Analysis of allograft biopsies from patients following Campath-1H induction have suggested that monocyte activation associated with nuclear factor kappa B (NFkB), and the cytokine tumor necrosis factor alpha (TNFa) are responsible for Campath resistant rejections. A pilot trial was performed with 4 patients combining Campath with the TNFa sequestrant infliximab. This was the first use of this combination in humans, and it showed that this combination modestly delayed the onset of rejection and was well tolerated. A trial is underway and has accrued 2 patients combining Campath with the NFkB inhibitor deoxyspergualin. This is the first use of this combination in humans. Through collaboration with John Swanson, a trial has been initiated using the anti-T cell agent rabbit anti-thymocyte globulin to determine if this agent is as effective as Campath in allowing for immunosuppression reduction. Seventeen patients where accrued onto this trial. Most patients were able to have rejection-free survival with monotherapy rapamycin, though the induction appears logistically difficult to implement. A report of this trial has been accepted for publication in the British Journal The Lancet. Studies have been completed in humans evaluating the immediate post transplant condition of renal allograft, specifically addressing the role of innate immunity following graft reperfusion. We have determined that macrophage and monocytes are the initial cells migrating into the allograft (not T cells, the target cell for most clinically used immunosuppressants) and that other factors not typically addressed with conventional immunosuppression are hyper-expressed including complement, monocyte-colony stimulating factor and adhesion molecules. These studies are being used to evaluate novel induction agents to pair with or replace conventional therapies. The study has been accepted for publication in Transplantation. Transcriptional analysis of stable human kidney allografts has demonstrated that grafts hyper-express most pro-inflammatory costimulatory molecules, most notable CD154. The finding that so-called healthy allografts treated with conventional therapy differ markedly from normal kidneys has provided considerable insight into the causes of chronic graft rejection and supported the continued pre-clinical development of CD154-specific therapy. Through a Collaborative Research and Development Agreement (CRADA) with Applied Biosystems, we have developed a polymerase chain reaction (PCR) based assay for the rapid evaluation of biopsy tissues from renal allografts. In a pilot clinical trial we have shown that this method can accurately predict rejection prior to its clinical detection. The method also is a reliable indicator of response to therapy. Transcripts associated with the costimulation pathways of T cell activation have been highlighted as critical mediators of human allograft rejection using this method. A CRADA has been established with Wyeth to study the expression of genes in allografts using several micro-array platforms. The study has provided blood, urine and biopsy material for analysis of the expression of essentially all genes. These will be correlated with allograft and patient outcome over the 3-year course of the study. Given the importance of the costimulation pathway, we have collaborated with David Harlan to test a novel CD154-specific monoclonal antibody, NC8A4. This antibody has been tested in the NHP skin transplant model and preliminarily shown to have modest efficacy in preventing skin allograft rejection. In collaboration with Mario Roderer we have applied a 12-color flow cytometry technique for the first time in humans. The method is being used to study the repopulation of human lymphocytes and monocytes following treatment with depleting antibody preparations used at the time of transplantation. Specific attention is being directed toward T cell populations with potential regulator activity. The resistance of memory T cells to depletion is also being investigated with this technique. Surgical support has been provided for the Autoimmunity Section's islet transplantation trials in NHPs directed by David Harlan.
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