Pulmonary toxicity is a potentially fatal untoward effect of anticancer therapy and our overall goal is to understand the biochemical factors determining drug-induced lung injury. Our central hypothesis is the therapeutic index of the anticancer drug bleomycin (BLM) Can be enhanced with new fundamental information concerning the biochemical basis of BLM- induced lung injury. The major focus, of our proposal will be on the role of bleomycin hydrolase (BH) in protecting lungs against BLM. We also intend to investigate the biological functions of mammalian BH and the factors that regulate its pulmonary expression. We believe these studies will ultimately help to improve the usefulness of BLM by providing therapeutic strategies that reduce cancer treatment morbidity. Damage to the pulmonary endothelium appears to be an early and essential event in fibrosis and we posit that BH levels in the pulmonary endothelium are a critical factor determining pulmonary sensitivity. The full length cDNA for human BH has recently been isolated and sequenced; expression in cells results in BLM resistance. The yeast BH homolog, Gal6, appears to bind DNA making it the first DNA-binding protease. Moreover, the crystal structure has been determined suggesting it is a proteosome-like multimeric enzyme. During the next funding period, we will capitalize on important reagents that have been developed by us and others during the previous four years. We propose to generate transgenic murine models in which BH is either overexpressed or deleted that should permit novel insights into the biological and metabolic role of BH. We expect to test whether predictions from yeast BH concerning structure and function are applicable to human BH by exploring for protein partners and examining protein domains. These studies should assist in understanding the biological and biochemical functions of BH. We will delimit factors that control BH expression.
The Specific Aims are to: 1) develop cells and mice deficient in BH, 2) develop cells and mice that overexpress BH, 3) isolate and characterize potential proteins that interact with BH and, 4) analyze the regulation of BH expression.
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