Synthesis and Biochemical Testing of Serine Protease Inhibitors

Grant Winner

Dean

Abstract

The present proposal outlines the synthesis and biochemical testing of 10 novel inhibitors of serine protease of Trypsin-like specificity. Compounds synthesized under this proposal will be evaluated for their inhibitory activity toward trypsin, thrombin, and granzyme A. Optimization of the side chains required for inhibitory potency will be pursued using Modde 5.0 software, a windows program for the generation and evaluation of statistical experimental designs. Sculpt software will also be used to optimize the chemical properties needed for enzyme inhibition. The best inhibitors will be used to probe the role of Granzyme A in the apoptotic pathways of tumor cells and of myocytes after ischemic injury. Programmed cell death, apoptosis, involves very distinctive changes within the target cell nucleus, including margination of the chromatin, DNA fragmentation and breakdown of the nuclear envelope. Cytolytic granule-mediated target cell apoptosis is effected, in part, through synergistic action of the membrane-acting protein perforin and serine proteases, such as granzymes A or B. Cytotoxic Natural Killer (NK) and T lymphocytes (CTL) recognize and kill virally infected cells and tumor cells without affecting bystander cells. One important mechanism of killing, used by both T and NK cells, involves exocytosis of cytotoxic granules. Specific therapies designed to enhance or reduce the susceptibility of individual cell types to undergo apoptosis could form the basis for a new treatment modality of many human diseases. Granzyme inhibitors than can block apoptosis in myocytes have potential for development as therapeutic agents in the treatment of ischemic heart injury. Granzyme inhibitors also hold potential in the treatment of host-vs-graft disease, autoimmune diseases and neurodegenerative diseases such as Alzheimer's disease. Publishing of this work in peer-reviewed journals and further funding for continuation of work will be sought.