The inhibitory potency and selectivity of arginine substrate site nitric-oxide synthase inhibitors is solely determined by their affinity toward the different isoenzymes

作     者：Boer, R Ulrich, WR Klein, T Mirau, B Haas, S Baur, I 

作者机构：Byk Gulden Lomberg GmbH D-78467 Constance Germany 

出 版 物：《MOLECULAR PHARMACOLOGY》 (Mol. Pharmacol.)

年 卷 期：2000年第58卷第5期

页      面：1026-1034页

核心收录：

学科分类：1007[医学-药学(可授医学、理学学位)] 10[医学] 

主　　题：精氨酸/代谢 瓜氨酸/代谢 酶抑制剂/药理学 同工酶类/拮抗剂和抑制剂 同工酶类/代谢 一氧化氮合酶/拮抗剂和抑制剂 一氧化氮合酶/代谢 甲基吡啶类/药理学 放射配体测定 底物特异性 氚 人类 

摘      要：We have investigated various nitric oxide (NO) synthase inhibitors for their affinity and selectivity toward the three human isoenzymes in radioligand binding experiments. Therefore, we developed the new radioligand [H-3]2-amino-4-picoline to measure binding of these compounds to the three human NO synthase (NOS) isoenzymes. Aminopicoline is a potent and nonselective inhibitor of all three isoforms. [H-3]2-amino-4-picoline bound saturably and with high affinity to human NOSs. Affinity constants (K-D values) of 59, 111, and 136 nM were obtained for the inducible, neuronal, and endothelial NOS isoforms (iNOS, nNOS, eNOS). Binding of [H-3]2-amino-4-picoline was competitive with the substrate arginine. From all the inhibitors tested, AMT (2-amino-5,6-dihydro-6-methyl-4H-1,3-thiazine hydrochloride) showed the highest affinity and no selectivity. L-NIL [L-N-6-(1-Iminoethyl) lysine hydrochloride] and aminoguanidine were moderately iNOS-selective while L-NA (N-G-nitro-L-arginine) and L-NAME (N-G-nitro-L-arginine methyl ester hydrochloride) showed selectivity toward the constitutive isoforms. High iNOS versus eNOS selectivity was found for 1400W, whereas several isothiourea derivatives and 1400W displayed moderate n- versus eNOS selectivity. To relate the affinity of these compounds to their inhibitory potency, we measured the inhibitory potency under almost identical conditions using a new microtiter plate assay. The inhibitory potency of selective and nonselective NOS inhibitors was almost exactly mirrored by their affinity toward the different isoenzymes. Highly significant correlations were obtained between the potency of enzyme inhibition and the inhibition of [H-3]2-amino-4-picoline binding for all three isoenzymes. These data show that the potency and selectivity of NOS inhibitors are solely determined by their affinity toward the different isoforms. Furthermore, these data identify the new radioligand [3H]2-amino-4-picoline as a very useful radiolabel for the investi