Departments of Chemistry (A.G., M.Q., N.A.H.) and Pharmacology and Therapeutics (R.L.P, C.S., M.Q.), University of Florida, Gainesville, Florida; and Department of Pharmaceutical Sciences, School of Pharmacy, Bouvé College of Health Sciences, Northeastern University, Boston, Massachusetts (S.G., G.A.T.).
Departments of Chemistry (A.G., M.Q., N.A.H.) and Pharmacology and Therapeutics (R.L.P, C.S., M.Q.), University of Florida, Gainesville, Florida; and Department of Pharmaceutical Sciences, School of Pharmacy, Bouvé College of Health Sciences, Northeastern University, Boston, Massachusetts (S.G., G.A.T.)
Mol Pharmacol. 2019 Jun;95(6):606-614. doi: 10.1124/mol.119.115758. Epub 2019 Apr 3.
Nicotinic acetylcholine receptors (nAChRs) are members of the Cys-loop superfamily of ligand-gated ion channels. Typically, channel activation follows the binding of agonists to the orthosteric binding sites of the receptor. 7 nAChRs have a very low probability of channel activation, which can be reversed by the binding of 7 selective positive allosteric modulators (PAMs) to putative sites within the transmembrane domains. Although typical PAMs, like PNU-120596, require coapplication of an orthosteric agonist to produce large channel activations, some, like GAT107 and B-973B [(S)-3-(3,4-difluorophenyl)-N-(1-(6-(4-(pyridin-2-yl)piperazin-1-yl)pyrazin-2-yl)ethyl)propanamide], are characterized as allosteric activating PAMs, which also bind to an allosteric activation (AA) site in the extracellular domain and activate the 7 ion channel by themselves. We had previously characterized ,-diethyl-'-phenylpiperazine analogs with various functions. In this work, we docked members of this family to a homology model of the 7 receptor extracellular domain. The compound 1,1-diethyl-4(naphthalene-2-yl)piperazin-1-ium (2NDEP) a weak partial agonist, showed particularly favorable docking and binding energies at the putative AA site of the receptor. We hypothesized that 2NDEP could couple with PAMs through the AA site. This hypothesis was tested with the 7 mutant C190A, which is not activated by orthosteric agonists but is effectively activated by GAT107. The results showed that 2NDEP acts as an allosteric agonist of 7C190A when coapplied with the PAM PNU-120596. Also, the allosteric activity was nearly abolished upon coapplication with the AA site-selective antagonist 2,3,5,6MP-TQS (cis-trans-4-(2,3,5,6-tetramethylphenyl)-3a,4,5,9b-tetrahydro-3H-cyclopenta[c]quinoline-8-sulfonamide), consistent with AA site involvement. Overall, our findings show a novel mode of agonism through an allosteric site in the extracellular domain of 7 nAChR.
烟碱型乙酰胆碱受体（nAChRs）是配体门控离子通道 Cys 环超家族的成员。通常情况下，通道的激活是由激动剂与受体的正构结合位点结合引起的。7 个 nAChRs 的通道激活概率非常低，而这种概率可以通过 7 个选择性正变构调节剂（PAMs）与跨膜域内假定的结合位点结合来逆转。虽然像 PNU-120596 这样的典型 PAMs 需要与正构激动剂共同应用才能产生大的通道激活，但有些像 GAT107 和 B-973B [(S)-3-(3,4-二氟苯基)-N-(1-(6-(4-(吡啶-2-基)哌嗪-1-基)哒嗪-2-基)乙基)丙酰胺]，则被定义为变构激活 PAMs，它们也与细胞外域的变构激活（AA）位点结合，并通过自身激活 7 个离子通道。我们之前已经对具有各种功能的 - 二乙基 - - 苯基哌嗪类似物进行了表征。在这项工作中，我们将这个家族的成员对接至 7 个受体细胞外域的同源模型。化合物 1,1-二乙基-4（萘-2-基）哌嗪-1-鎓（2NDEP）是一种弱部分激动剂，在受体的假定 AA 位点表现出特别有利的对接和结合能。我们假设 2NDEP 可以通过 AA 位点与 PAMs 偶联。通过 7 个突变体 C190A 对该假设进行了测试，该突变体不能被正构激动剂激活，但可以被 GAT107 有效激活。结果表明，当与 PAM PNU-120596 共同应用时，2NDEP 可作为 7C190A 的变构激动剂。另外，当与 AA 位点选择性拮抗剂 2,3,5,6MP-TQS（顺式-反式-4-(2,3,5,6-四甲基苯基)-3a,4,5,9b-四氢-3H-环戊[c]喹啉-8-磺酰胺）共同应用时，变构活性几乎被完全消除，这与 AA 位点的参与一致。总的来说，我们的研究结果显示了一种通过 7 个 nAChR 细胞外域的变构位点发挥作用的新型激动机制。
