Two approaches to optimize cyclosporine (CsA)-based clinical immunosuppression were pursued during the initial grant period: pharmacokinetic drug profiling to predict initial and adjust subsequent CsA doses, and development of the median effect analysis method to assess immunosuppressive drug interactions. The present proposal uses a stochastic approach to refine dose prediction and examines combinations of CsA with novel immunosuppressive agents: rapamycin (RAPA); the di-hydro- orotate dehydrogenase inhibitor, quinoline carboxylic acid (QCA); murine IgG2b monoclonal antibodies directed toward the alpha/beta T-cell receptor; and genetically engineered, soluble interleukin receptors.
Aim 1 utilizes the median effect analysis to dissect solo and combined effects of these agents on in vivo allorejection of mouse skin and vascularized heart grafts; rat renal, cardiac, small bowel, pancreato- duodenal and hepatic allografts; and canine renal transplants.
Aim 2 dissects cellular and humoral mechanisms of drug-induced unresponsiveness, particularly seeking to identify and characterize the immunodeviated blocking antibody induced by RAPA.
Aim 3 dissects the mechanisms of drug effects. Interindividual variations among CsA-treated renal transplant patients' responses after 12 hrs. of in vitro PHA activation are assessed by generation of a cytoplasmic activator of DNA replication and of IL-2 mRNA. RAPA's mechanisms of action on growth factor receptor-mediated endocytosis, phosphorylation reactions, c-AMP levels, and oncogene activation events are examined using IL-2, IL-4 and IL-6-sensitive cloned lymphocytes, epidermal growth factor-sensitive Mink lung cells, and platelet-derived growth factor-sensitive Balb-c NIH/3T3 cells.
Aim 4 examines RAPA (and QCA) pharmacokinetics, including development of new drug detection assays, measurement of canine tissue distribution, and assessment of drug interactions with CsA.
Aim 5 tests the benefit of stochastic NPML computer models to predict initial, as well as adjust subsequent posttransplant, oral CsA doses compared with the present simple proportionate models. In aggregate, the studies seek to overcome interindividual pharmacokinetic differences and standardize CsA exposure, thereby permitting clinical assessment of pharmacodynamic variation and evaluation of the impact of new immunosuppressive agents using rigorous pharmacologic tools.
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