Pharmacological characterisation of [(pX)Phe4]nociceptin(1-13)amide analogues -: 1.: In vitro studies
Pharmacological characterisation of [(pX)Phe4]nociceptin(1-13)amide analogues -: 1.: In vitro studies
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DOI：
10.1007/s00210-002-0548-8
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发表时间：
2002-06-01
影响因子：
3.6
通讯作者：
Calo', G
中科院分区：
文献类型：
--
作者：
Bigoni, R;Rizzi, D;Calo', G
痛敏肽（NC）序列中的Phe（4）被认为是受体相互作用的最关键残基。在本研究中。我们研究了一系列NC（1-13）NH_2类似物的药理活性，其中Phe（4）的帕拉氢原子被F取代。NO2。在使用表达重组人NC受体（CHOhOP 4）的CHO细胞和大鼠大脑皮层膜进行的受体结合研究中，[（pF）Phe 4]NC（1-13）NH 2、[（pNO（2））Phe（4）]NC（1 - 13）NH 2和[（pCN）Phe（4）] NC（1 -13）NH 2显示出比NC（1- 13）NH 2更高的亲和力。[（pCl）Phe（4）]NC（1-13）NH 2的亲和力与NC（1-13）NH 2的亲和力基本相同，而其余化合物显示出降低的亲和力。在一系列功能试验中（刺激CHOhOP 4细胞和大鼠大脑皮层膜中的GTP γ S结合，抑制CHOhOP 4细胞中的cAMP蓄积）。帕拉取代的类似物表现为完全激动剂（除了[（pOH）Phe（4）]NC（1-13）NH 2，其在GTP γ S结合测定中作为部分激动剂），具有以下等级顺序效力：pF = pNO（2）大于或等于pCN大于或等于NC = NC（1-13）NH 2 = pCl大于或等于pBr > pI = pCH（3）> pOH > pNH（2）[（pF）Phe（4）]NC（1-13）NH 2和[（pNO（2））Phe（4）]NC（1-13）NH 2在表达经典阿片受体的细胞上进行的cAMP积累实验中是无活性的或显示微摩尔效力。除了[（pOH）Phe（4）]NC（1-13）NH 2显示部分激动剂/弱拮抗剂活性外，所有化合物在来自各种物种（豚鼠回肠、小鼠结肠和小鼠/大鼠输精管）的分离组织中均为完全激动剂。效价的等级顺序与其他试验中发现的相似。所有类似物的作用均未被纳洛酮改变。选择性OP 4受体拮抗剂[Nphe（1）]NC（1-13）NH 2在所有制备物中针对高效衍生物[（pF）Phe（4）] NC（1-13）NH 2和[（pNO（2））Phe（4）]NC（1-13）NH 2中的一种或两种进行测试，显示出与针对NC所发现的值相似的pA（2）值，在GTP γ S结合/大鼠大脑皮层测定中的pA（2）高得多（约1.5%）。7.5)比在其它功能测定中（ca. 6）。该研究进一步支持了NC的Phe（4）是受体占据和激活的关键残基的观点。此外，作为本研究的一部分，我们已经鉴定了两种新型的、高效的和选择性的OP 4受体激动剂，[（pF）Phe（4）]NC（1-13）NH 2和[（pNO（2））Phe（4）]NC（1-13）NH 2。
Phe(4) in the nociceptin (NC) sequence has been identified as the most critical residue for receptor interaction. In the present study. we investigated the pharmacological activity of a series of NC(1-13)NH2 analogues, in which the hydrogen atom in the para position of Phe(4) was substituted with F. NO2. CN, Cl, Br, I, CH3, OH or NH2.In receptor binding studies, performed using CHO cells expressing the recombinant human NC receptor (CHOhOP4) and in rat cerebral cortex membranes, [(pF)Phe4]NC (1-13)NH2, [(pNO(2))Phe(4)]NC(1-13)NH2, and [(pCN)Phe(4)] NC(1-13)NH2 displayed higher affinity than NC(1-13)NH2. The affinity of [(pCl)Phe(4)]NC(1-13)NH2 was essentially identical to that of NC(1-13)NH2, while the remaining compounds displayed reduced affinity. In a series of functional assays (stimulation of GTPgammaS binding in CHOhOP4 cells and rat cerebral cortex membranes and inhibition of cAMP accumulation in CHOhOP4 cells). the para substituted analogues behaved as full agonists (with the exception of [(pOH)Phe(4)]NC(1-13)NH2 which acted as a partial agonist in the GTPgammaS binding assays) with the following rank order potency:pF = pNO(2) greater than or equal to pCN greater than or equal to NC = NC(1-13)NH2= pCl greater than or equal to pBr > pI = pCH(3) > pOH > pNH(2)[(pF)Phe(4)]NC(1-13)NH2 and [(pNO(2))Phe(4)]NC(1-13)NH2 were either inactive or displayed micromolar potencies in cAMP accumulation experiments performed on cells expressing classical opioid receptors. All compounds were full agonists in isolated tissues from various species (guinea pig ileum, mouse colon and mouse/rat vas deferens) with the exception of [(pOH)Phe(4)]NC(1-13)NH2 which displayed partial agonist/weak antagonist activities. The rank order of potency was simdar to that found in the other assays. The effects of all analogues were not modified by naloxone. The selective OP4 receptor antagonist [Nphe(1)]NC(1-13)NH2, tested in all preparations against one or both of the highly potent derivatives [(pF)Phe(4)] NC(1-13)NH2 and [(pNO(2))Phe(4)]NC(1-13)NH2, showed pA(2) values similar to those found against NC, the pA(2) in the GTPgammaS binding/rat cerebral cortex assay being much higher (ca. 7.5) than in the other functional assays (ca. 6). This study further supports the notion that Phe(4) of NC is the critical residue for receptor occupation and activation. Moreover, as part of this study, we have identified two novel, highly potent and selective agonists for the OP4 receptor, [(pF)Phe(4)]NC(1-13)NH2 and [(pNO(2))Phe(4)]NC (1-13)NH2.