Nitsche Valentin, Höfner Georg, Kaiser Jesko, Gertzen Christoph G W, Seeger Thomas, Niessen Karin V, Steinritz Dirk, Worek Franz, Gohlke Holger, Paintner Franz F, Wanner Klaus T
Department of Pharmacy - Center for Drug Research, Ludwig-Maximilians-Universität München, Munich, Germany.
Institute for Pharmaceutical and Medicinal Chemistry, Heinrich Heine Universität Düsseldorf, Düsseldorf, Germany.
Toxicol Lett. 2024 Feb;392:94-106. doi: 10.1016/j.toxlet.2024.01.003. Epub 2024 Jan 10.
Intoxications with organophosphorus compounds (OPCs) based chemical warfare agents and insecticides may result in a detrimental overstimulation of muscarinic and nicotinic acetylcholine receptors evolving into a cholinergic crisis leading to death due to respiratory failure. In the case of the nicotinic acetylcholine receptor (nAChR), overstimulation leads to a desensitization of the receptor, which cannot be pharmacologically treated so far. Still, compounds interacting with the MB327 binding site of the nAChR like the bispyridinium salt MB327 have been found to re-establish the functional activity of the desensitized receptor. Only recently, a series of quinazoline derivatives with UNC0642 as one of the most prominent representatives has been identified to address the MB327 binding site of the nAChR, as well. In this study, UNC0642 has been utilized as a reporter ligand to establish new Binding Assays for this target. These assays follow the concept of MS Binding Assays for which by assessing the amount of bound reporter ligand by mass spectrometry no radiolabeled material is required. According to the results of the performed MS Binding Assays comprising saturation and competition experiments it can be concluded, that UNC0642 used as a reporter ligand addresses the MB327 binding site of the Torpedo-nAChR. This is further supported by the outcome of ex vivo studies carried out with poisoned rat diaphragm muscles as well as by in silico studies predicting the binding mode of UNC0646, an analog of UNC0642 with the highest binding affinity, in the recently proposed binding site of MB327 (MB327-PAM-1). With UNC0642 addressing the MB327 binding site of the Torpedo-nAChR, this and related quinazoline derivatives represent a promising starting point for the development of novel ligands of the nAChR as antidotes for the treatment of intoxications with organophosphorus compounds. Further, the new MS Binding Assays are a potent alternative to established assays and of particular value, as they do not require the use of radiolabeled material and are based on a commercially available compound as reporter ligand, UNC0642, exhibiting one of the highest binding affinities for the MB327 binding site known so far.
基于有机磷化合物（OPC）的化学战剂和杀虫剂中毒，可能会导致毒蕈碱型和烟碱型乙酰胆碱受体受到有害的过度刺激，进而演变成胆碱能危象，最终因呼吸衰竭而死亡。就烟碱型乙酰胆碱受体（nAChR）而言，过度刺激会导致该受体脱敏，而目前尚无有效的药理学治疗方法。不过，已发现与nAChR的MB327结合位点相互作用的化合物，如双吡啶盐MB327，能够恢复脱敏受体的功能活性。就在最近，人们还发现了一系列喹唑啉衍生物，其中最突出的代表是UNC0642，它也能作用于nAChR的MB327结合位点。在本研究中，UNC0642被用作报告配体，以建立针对该靶点的新结合测定法。这些测定法遵循质谱结合测定法的概念，即通过质谱评估结合的报告配体的量，无需使用放射性标记材料。根据所进行的包括饱和和竞争实验在内的质谱结合测定结果，可以得出结论，用作报告配体的UNC0642作用于电鳐nAChR的MB327结合位点。用中毒大鼠膈肌进行的体外研究结果，以及计算机模拟研究预测UNC0642的类似物UNC0646在最近提出的MB327结合位点（MB327-PAM-1）中的结合模式，进一步支持了这一结论。由于UNC0642作用于电鳐nAChR的MB327结合位点，因此这种及相关的喹唑啉衍生物是开发新型nAChR配体作为治疗有机磷化合物中毒解毒剂的一个有前途的起点。此外，新的质谱结合测定法是现有测定法的有力替代方法，具有特殊价值，因为它们不需要使用放射性标记材料，并且基于一种市售化合物作为报告配体UNC0642，该化合物对MB327结合位点表现出迄今已知的最高结合亲和力之一。
