HIGH-AFFINITY BINDING OF DTZ323, A NOVEL DERIVATIVE OF DILTIAZEM, TO RABBIT SKELETAL-MUSCLE L-TYPE CA++ CHANNELS

A novel derivative of diltiazem (1,5-benzothiazepine Ca++ antagonist), DTZ323, -5-[2-[[2-(3,4-dimethoxyphenyl)ethyl]-methylamino] -(4-methox yphe-nyl)-1,5-benzothiazepine-4(5H)-one, was characterized by radiolig and binding experiments with rabbit skeletal T-tubule membranes in ter ms of the affinity and the selectivity to the binding sites for the th ree classical calcium antagonists, such as dihydropyridines, phenylalk ylamines and benzothiazepines. DTZ323, like diltiazem and clentiazem, exhibited complete and concentration-dependent inhibition of d-cis-[H- 3]diltiazem binding to the membrane with a slope factor close to unity . Ki 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 clent iazem, respectively. DTZ323 partially inhibited the specific binding o f a dihydropyridine ligand, (+)-[H-3]PN200-110, at 37 degrees C. The e quilibrium saturation study showed that DTZ323 reduces the affinity fo r the (+)-[H-3]PN200-110 binding in a concentration-dependent manner w ith a slight decrease in the density of the binding sites. DTZ323 also inhibited the specific binding of a phenylalkylamine ligand, (-)-[H-3 ]D888, completely as did diltiazem. DTZ323 (1 mu M) had no effect on t he dissociation rate of d-cis-[H-3]diltiazem at 2 degrees C, whereas 3 0 mu M verapamil increased the dissociation rate, which suggested that DTZ323 inhibits the specific binding of d-cis-[H-3]diltiazem in a man ner similar to other competitive ligands for the benzothiazepine bindi ng site. These results indicate that DTZ323 is a selective ligand for the 1,5-benzothiazepine binding site with the highest affinity among t he diltiazem derivatives.