TRANSCELLULAR LEUKOTRIENE BIOSYNTHESIS AND INFLAMMATION

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Leukotrienes (LT) are lipid mediators of pulmonary inflammatory disorders. LTB4 stimulates adhesion of circulating neutrophils to vascular endothelium, directs their migration to sites of inflammation, and catalyzes the release of their granule constituents. It may produce hyperalgesia, myelopoiesis, and exert myotropic effects in some organs. The cysteinyl leukotrienes (LTC4, LTD4, and LTE4) constrict cardiac and bronchial smooth muscle and promote vascular permeability. Pathophysiology actions of leukotrienes are best understood in terms of their roles in immediate hypersensitivity and asthma. Roles in chronic inflammation associated with pulmonary disorders are likely, but not fully understood. Derangements in the cellular processes which govern the accumulation and disposition of LTA4, the pivotal biosynthetic intermediate in the leukotriene enzymatic pathway, are important in the pathogenesis of neutrophil-mediated lung damage. LTA4 hydrolase, a rate- limiting enzyme in the pathway is a critical determinant in the context. We have recently identified two types of LTA4 hydrolase inhibition: i) a constitutive process involving irreversible, mechanism-based inactivation which accompanies catalysis ii) a pharmacological process involving reversible inhibition by bestatin or captopril. We propose to evaluate selective modulation of LTA4 metabolism as a new pharmacological strategy for alleviating pulmonary inflammation. We also propose to determine what are the consequences of LTA4 accumulation on neutrophils when LTA4 hydrolase is inhibited. The results of our investigations will have direct implications for the therapeutic management of allergy and inflammatory pulmonary disorders. In four specific aims we plan to investigate: I. Effect of LTA4 Hydrolase Inhibition on Pulmonary Leukotriene Formation and Pulmonary Vascular Pressure in Isolated, Perfused Lungs. II. Regulation of 5-Lipoxygenase Activity by LTA4: Inactivation of 5- Lipoxygenase via Irreversible 'Feedback" Inhibition. III. Requirements for Substrate and Inhibitor Binding at the LTA4 Hydrolase/Aminopeptidase Active Site. IV. Affinity Chromatography Separation of Catalytically Active and 'Suicide' Inactivated Forms of LTA4 Hydrolase/Aminopeptidase: Development of an Assay to Detect Transcellular Leukotriene Biosynthesis.