Vol. 281, Issue 1, 173-179, 1997

High-Affinity Binding of DTZ323, a Novel Derivative of Diltiazem, to Rabbit Skeletal Muscle L-type Ca⁺⁺ Channels¹

Masafumi Hagiwara, Satomi Adachi-Akahane and Taku Nagao

Department of Toxicology and Pharmacology, Faculty of Pharmaceutical Sciences, University of Tokyo, Tokyo, Japan

A novel derivative of diltiazem (1,5-benzothiazepine Ca⁺⁺ antagonist), DTZ323, 3-(acetyloxy)-5-[2-[[2-(3,4-dimethoxyphenyl)ethyl]-methylamino]ethyl]-2,3-dihydro-2-(4-methoxyphenyl)-1,5-benzothiazepine-4(5H)-one, was characterized by radioligand binding experiments with rabbit skeletal T-tubule membranes in terms of the affinity and the selectivity to the binding sites for the three classical calcium antagonists, such as dihydropyridines, phenylalkylamines and benzothiazepines. DTZ323, like diltiazem and clentiazem, exhibited complete and concentration-dependent inhibition of d-cis-[³H]diltiazem binding to the membrane with a slope factor close to unity. K_i values indicated that DTZ323 (K_i = 6.6 ± 0.6 nM, mean ± S.E., n = 4) was 48 times and 9 times more potent than diltiazem and clentiazem, respectively. DTZ323 partially inhibited the specific binding of a dihydropyridine ligand, (+)-[³H]PN200-110, at 37°C. The equilibrium saturation study showed that DTZ323 reduces the affinity for the (+)-[³H]PN200-110 binding in a concentration-dependent manner with a slight decrease in the density of the binding sites. DTZ323 also inhibited the specific binding of a phenylalkylamine ligand, ()-[³H]D888, completely as did diltiazem. DTZ323 (1 µM) had no effect on the dissociation rate of d-cis-[³H]diltiazem at 2°C, whereas 30 µM verapamil increased the dissociation rate, which suggested that DTZ323 inhibits the specific binding of d-cis-[³H]diltiazem in a manner similar to other competitive ligands for the benzothiazepine binding site. These results indicate that DTZ323 is a selective ligand for the 1,5-benzothiazepine binding site with the highest affinity among the diltiazem derivatives.