BIRM-270 - A NOVEL INHIBITOR OF ARACHIDONATE RELEASE THAT BLOCKS LEUKOTRIENE B-4 AND PLATELET-ACTIVATING-FACTOR BIOSYNTHESIS IN HUMAN NEUTROPHILS

l-1-(2-pyridinyl)ethyl]-5-methyl-2-benzoxazolamine (BIRM 270) was iden tified as a potent and enantiomerically selective inhibitor of calcium ionophore A23187-stimulated leukotriene B-4 biosynthesis in human neu trophils. The (S)- and (R)-enantiomers exhibited IC50 values of 1 nM a nd 40 nM, respectively. BIRM 270 did not inhibit 5-lipoxygenase activi ty in a cell-free assay. In addition, the compound did not interfere w ith the conversion of exogenous 5-lipoxygenase substrate (15S)- hydrop eroxyeicosatetraenoic acid to (5S,15S)-dihydroxyeicosatetraenoic acid in intact, ionophore-stimulated neutrophils. Under the same experiment al conditions, BIRM 270 inhibited the production of 5-lipoxygenase pro ducts from endogenous substrate, suggesting that the compound affected arachidonate availability rather than metabolism. Consistent with thi s concept, the inhibition of leukotriene B-4 biosynthesis by BIRM 270 was overcome by the addition of exogenous arachidonic acid to the leuk ocyte preparation. Direct measurement of free arachidonate by gas chro matography-mass spectrometry confirmed that BIRM 270 inhibited arachid onate release from ionophore-stimulated neutrophils. The compound did not affect arachidonate reacylation. The blockage of arachidonate rele ase coincided with inhibition of leukotriene B-4 biosynthesis in these cells. BIRM 270 also inhibited ionophore-stimulated platelet-activati ng factor biosynthesis by human neutrophils. Although these results su ggest that BIRM 270 inhibited phospholipase A(2)-mediated deacylation of membrane phospholipids, the compound did not directly inhibit the h igh molecular weight, cytosolic phospholipase A(2) derived from human neutrophils or U937 cells. Thus, suppression of arachidonate mobilizat ion by BIRM 270 may be due to indirect inhibition of intracellular pho spholipase A(2) or to inhibition of another acylhydrolase activity.