Leukotrienes (LT's), the primary products of the 5-Lipoxygenase (5-LO) pathway are important mediators of inflammatory and allergic disorders. Leukotriene B4 (LTB4) is produced from numerous cell types such as neutrophils, monocytes, macrophages, keratinocytes, and Lymphocytes. LTB4 shows several inflammatory effects, such as chemotaxis for polymorphonuclear leukocyte margination and emigration. Peptido-leukotrienes LTC4, LTD4 and LTE4) are biosynthesized in inflammatory cells including monocytes, macrophages, eosinophils and tissue mast cells. The peptido-LTs cause smooth muscle contraction, increase the permeability of post-capillary venuoles and facilitate plasma leakage and edema formation, as well as cellular diapedesis. Inflammatory stimuli, such as phagocytosis, the chemotactic peptide f-met-leu-phe, and platelet activating factor (PAF) induce the synthesis of LT's. LT's are synthesized from arachidonic acid by 5-LO in various ways. An inhibitor of 5-LO may prevent the synthesis of LT's. CMI 977 is designed to inhibit 5-LO, therefore preventing the formation of LTB4 and initiation of the arachidonic acid cascade of LT synthesis. An ex vivo whole blood 5-LO assay in small rodents has provided a useful method in the evaluation of LT biosynthesis inhibitors. This ex vivo rodent model has been adapted to a nonhuman primate. In the model, rhesus monkeys are chosen for their genetic compatibility to humans and for their world-wide acceptance as a preliminary model for human testing. Briefly, rhesus monkey blood is collected into heparinized, polypropylene tubes by venipuncture into a peripheral vessel at specified times after dosing with CMI 977, orally or intravenously. The animals are anesthetized with ketamine hydrochloride at 5-10 mg/kg, i.m. prior to dosing and each blood collection. Whole blood is centrifuged at 6000 rpm-1 for 5 minutes for pharmacokinetic analysis. Plasma is retained for determination of CMI 977 concentration by HPLC analysis. The concentration [fg/ml] of drug is calculated for each time point. Pharmacodynamic data is obtained by removing a small (0.5 ml) aliquot of blood from the sample. This sample is incubated for 30 minutes at 37 C in the presence of calcium inophore (A23187) at [50fM]. After incubation, the blood is centrifuged at 37,000 rpm-1 for 2 minutes and plasma is salvaged to enzyme immunoassay (EIA) buffer for assay for LTB4 using an EIA kit from Cayman Chemical Company. The percent inhibition form each time point is calculated. An IND filed with the FDA for CMI 977 was approved in January, 1998. Clinical trials are scheduled to begin in March, 1998.
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