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Our primary goal has been to discover leukotriene biosynthesis inhibitors with characteristics that are appropriate for use as clinical agents. The success of the use of zileuton in the treatment of asthma led us to explore further the use of the N-hydroxyurea class of 5-lipoxygenase inhibitors as longer-acting compounds with good lung penetration. A variety of in vitro and in vivo methods were used to evaluate a large number of compounds, from which ABT-761 [(R)-N-(3-(5-(4-fluorophenylmethyl)thien-2-yl)-1-methyl-2-pr opynyl)-N-hydroxyurea] was selected for study. ABT-761 exhibited potent and selective inhibition of leukotriene formation both in vitro and in vivo. More importantly, the compound potently inhibited antigen-induced bronchospasm in guinea pigs when given either prophylactically or therapeutically. In addition, ABT-761 was a potent inhibitor of eosinophil influx into the lungs of Brown Norway rats. These data provide added support for the role of leukotrienes in both bronchospasm and eosinophilic inflammation and characterize ABT-761 as a particularly potent inhibitor of leukotrienes formed in pulmonary tissues. These data combined with the excellent pharmacokinetic characteristics of the compound indicate its potential use in the treatment of leukotriene-dependent human disease.

These studies examined the signal transduction mechanisms by which prostaglandin (PG) E2 production can occur in human amnionic WISH cells in response to the stimuli okadaic acid, interleukin (IL)-1beta, tumor necrosis factor (TNF)-alpha, phorbol-12-myristate-13-acetate (PMA) or combinations of PMA with IL-1beta or TNF-alpha. We also investigated whether WISH cells are capable of producing TNF-alpha or IL-1beta in response to stimulation, because these cytokines can be produced in an autocrine fashion to perpetuate an inflammatory response. Our data indicate that the magnitude of PGE2 production induced by a given stimulus correlated temporally with the level of PGH synthase-2 (PGHS-2) protein. PMA or IL-1beta induced PGE2 production 2 to 4 hr after treatment, whereas the combination of these agents produced the most rapid induction 2 hr after treatment. Only okadaic acid induced the production of both PGE2 and TNF-alpha, after a lag of 12 to 18 hr. PGE2 production by all stimuli was inhibited by dexamethasone, the IL-1 receptor antagonist (IL-1ra), the specific PGHS-2 inhibitor NS-398 and the protein kinase inhibitor staurosporin. In contrast, TNF-alpha production in response to okadaic acid was inhibited by the TNF-converting enzyme inhibitor GI 129471 and staurosporin but was unaffected by either IL-1ra, dexamethasone or NS-398. We conclude that WISH cells are capable of producing bioactive proinflammatory mediators such as TNF-alpha and PGE2 through separable intracellular signal transduction mechanisms. The ability of IL-1ra to reduce PGE2 production caused by all stimuli used suggests an autocrine role for IL-1 in PGHS-2 induction in these cells.