Knapp R J, Santoro G, De Leon I A, Lee K B, Edsall S A, Waite S, Malatynska E, Varga E, Calderon S N, Rice K C, Rothman R B, Porreca F, Roeske W R, Yamamura H I
Department of Pharmacology, College of Medicine, University of Arizona, Tucson, USA.
J Pharmacol Exp Ther. 1996 Jun;277(3):1284-91.
The racemic compound (+/-)-BW373U86 ¿(+/-)-4-((alpha R*)- alpha-((2S*,5R*)-4-allyl-2,5-dimethyl-1-piperazinyl)-3-hydroxy- benzyl)-N,N-diethylbenzamide dihydrochloride} is a potent delta opioid receptor agonist in the mouse vas deferens assay with little mu or kappa opioid receptor activity in the guinea pig ileum tissue preparation. In contrast, radioligand binding studies show that (+/-)-BW373U86 is only about 10-fold selective for delta over mu opioid receptors. Studies of the enantiomeric forms of (+/-)-BW373U86 and derivatives (SNC80 and related compounds) show that some of these isomers are significantly better in both receptor binding and pharmacological selectivity than (+/-)-BW373U86. In this study we have determined the binding affinities of 10 different SNC80-related compounds at cloned human delta and mu opioid receptors and measured the potency of SNC80 for the inhibition of forskolin-stimulated adenylyl cyclase. The most selective delta receptor ligand (SNC162) differed from SNC80 by the absence of the 3-methoxy substitution of the benzyl ring. The Ki for SNC162 at the delta receptor (0.625 nM) was over 8700-fold lower than that at the mu receptor (5500 nM), making this the most selective delta receptor ligand available. Reduction of the allyl side chain of SNC80 to produce radiolabeled [3H]SNC121 allowed direct measurement of the association and dissociation rate constants. SNC80 was 26-fold less potent than [D-Pen2, pCI-Phe4, D-Pen5]enkephalin in the delta receptor adenylyl cyclase inhibition assay, but showed full agonist activity with an EC50 value of 9.2 nM. The regulation of SNC80 binding affinity to the delta receptor by GTP analogs is undetectable in [3H]naltrindole binding inhibition studies, but direct binding studies with [3H]SNC121 in the presence of 100 microM 5'-guanylylimidotriphosphate show a 55% reduction in maximum binding site density consistent with a lower affinity for a part of the receptor population. Addition of 120 mM sodium chloride reduces SNC80 affinity nearly 40-fold in [3H]naltrindole binding inhibition studies. The results of these studies define specific structural features of these compounds responsible for opioid receptor interactions and suggest a possibly novel mechanism for delta receptor activation.
外消旋化合物（±）-BW373U86 {（±）-4- [（αR*）-α-（（2S*，5R*）-4-烯丙基-2,5-二甲基-1-哌嗪基）-3-羟基苄基]-N，N-二乙基苯甲酰胺二盐酸盐}在小鼠输精管试验中是一种强效的δ阿片受体激动剂，而在豚鼠回肠组织制备中几乎没有μ或κ阿片受体活性。相比之下，放射性配体结合研究表明，（±）-BW373U86对δ阿片受体的选择性仅比对μ阿片受体高约10倍。对（±）-BW373U86及其衍生物（SNC80和相关化合物）的对映体形式的研究表明，其中一些异构体在受体结合和药理选择性方面均明显优于（±）-BW373U86。在本研究中，我们测定了10种不同的SNC80相关化合物对克隆的人δ和μ阿片受体的结合亲和力，并测量了SNC80抑制福斯高林刺激的腺苷酸环化酶的效力。最具选择性的δ受体配体（SNC162）与SNC80的不同之处在于苄基环没有3-甲氧基取代。SNC162在δ受体处的Ki（0.625 nM）比在μ受体处的Ki（5500 nM）低8700多倍，这使其成为现有的最具选择性的δ受体配体。将SNC80的烯丙基侧链还原以产生放射性标记的[3H]SNC121，从而可以直接测量缔合和解离速率常数。在δ受体腺苷酸环化酶抑制试验中，SNC80的效力比[D-Pen2，pCI-Phe4，D-Pen5]脑啡肽低26倍，但显示出完全激动剂活性，EC50值为9.2 nM。在[3H]纳曲吲哚结合抑制研究中，未检测到GTP类似物对SNC80与δ受体结合亲和力的调节，但在100μM 5'-鸟苷酰亚胺三磷酸存在下用[3H]SNC121进行的直接结合研究表明，最大结合位点密度降低了55％，这与对一部分受体群体的较低亲和力一致。在[3H]纳曲吲哚结合抑制研究中，添加120 mM氯化钠可使SNC80的亲和力降低近40倍。这些研究结果确定了这些化合物与阿片受体相互作用的特定结构特征，并提出了一种可能的δ受体激活新机制。
