DY-9760e, a novel calmodulin antagonist with cytoprotective action.

Abstract We report the pharmacological characterization and cytoprotective effect of DY-9760e, 3-[2-[4-(3-chloro-2-methylphenyl)-1-piperazinyl]ethyl]-5,6-dimethoxy-1-(4-imidazolylmethyl)-1H-indazole dihydrochloride 3.5 hydrate, a novel antagonist of calmodulin. DY-9760e inhibited calmodulin-dependent enzymes, including calmodulin-dependent phosphodiesterase and myosin light chain kinase with K i values of 1.4, 2.0, 3.8 and 133 μ M, respectively. These antagonistic effects of DY-9760e were more potent than those of W-7, N -(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide, another calmodulin antagonist. This compound showed little or no effect on calmodulin-independent enzymes, such as protein kinase A and C and calpain I and II. Analysis of the hydrophobic interaction of DY-9760e with calmodulin by using 2- p -toluidinyl-naphthalene-6-sulfonate and 9-anthroylcholine revealed that, like W-7, DY-9760e bound to the hydrophobic regions of calmodulin. The [ 14 C ]DY-9760e binding assay indicated that DY-9760e bound to calmodulin at one class of binding site. Finally, DY-9760e substantially protected N1E-115 neuroblastoma cells from cytotoxicity induced by the Ca 2+ ionophore, A23187. These results indicate that DY-9760e, a novel calmodulin antagonist, possesses a cytoprotective action and suggest that calmodulin plays a critical role in mediating some of the biochemical events leading to cell death following Ca 2+ overload.