Optimization of allosteric modulators of nicotinic receptors

描述(申请人提供)：最近的临床和遗传学研究提供了强有力的证据，精神分裂症患者发现的自发听觉门控缺陷可能受到位于海马区CA3区的A7尼古丁-乙酰胆碱受体(NAChRs)的影响。这种听觉/感觉门控的缺陷表现为感觉过载的体验，可能会加剧精神分裂症的积极和消极症状。人类形式的听觉缺陷的特征是对间隔50ms的成对听觉诱发电位的不完全抑制加工(P50)。P50听觉缺陷在遗传上与15号染色体上的A7 nAChR基因座有关。此外，对精神分裂症患者大脑的尸检分析表明，与正常人相比，A7 nAChR显著减少。P50听觉缺陷对胆碱能药物敏感，如非选择性激动剂(如尼古丁)和选择性A7nAChRs激动剂DMXB-A(GTS-21)。在动物模型中，对非选择性或选择性A7nAChR激动剂的敏感性也可被选择性A7nAChR拮抗剂如甲基琥珀酸乌头碱(MLA)或a-银环蛇毒素所阻断，但不能被A4？2选择性拮抗剂如二氢乙烷(DHE)所阻断。啮齿动物的P20-N40听觉诱发电位(AEP)类似于人类的P50听觉诱发电位，是评估各种药物的疗效的相关模型，这些药物可能有助于治疗精神分裂症的感觉门控缺陷。最近利用啮齿动物听觉门控缺陷模型的证据表明，选择性变构调节剂的A7nAChRs可以提高A7nAChRs的敏感性，从而纠正门控缺陷。然而，这些原则证明研究中的代表性分子(PNU-120596)具有几个使其作为治疗候选药物缺乏吸引力的属性。PNU-120596是一种A7nAChRs的变构调节剂，它显著改变了对激动剂的天然动力学反应，使得在PNU-120596存在下，被高浓度尼古丁脱敏的A7nAChRs群体可以被重新敏化。这是有问题的，因为A7nAChRs调节钙离子电导。在一种治疗方案中，由于PNU-120596的慢性存在而导致的大量和持续的钙离子内流被预测是神经毒性的。相反，我们的实验室已经开发出选择性的A7 nAChR阳性变构调节剂，它增强并保留了受体激活的天然动力学，以便在长期暴露于我们的化合物脱敏的A7 nAChR时保持不变。虽然我们的药物发现工作已经解决了保留通道活动的天然动力学的问题，但我们不确定在A7 nAChR功能已经处于次优状态(例如精神分裂症)的情况下，在避免与钙++相关的神经毒性的同时，A7 nAChR的疗效水平适合于治疗效果。拟议研究的目的是确定在A7 nAChR功能固有受损的DBA/2小鼠模型中，需要A7 nAChRs的最低水平的正变构效应才能逆转P20-N40听觉门控缺陷。候选分子将被合成，并在电生理测试中测试，以增强尼古丁引起的A7nAChR电流，以确定相对于尼古丁最大功效和效力的结构、活性关系。所提出的分子将对A7nAChRs表现出正向调制，而不激活其他nAChR亚型(例如，A4？2，A3？4)或其他环状配体门控离子通道(GABAA，5HT3A)。符合我们的受体亚型选择性、效力和疗效选择标准的候选分子的药代动力学(PK)谱将被评估是否可以达到适当的脑水平(即，对应于体外观察到的正变构调节剂的最大增强)。拟议研究的完成将确定通过A7nAChRs调节胆碱能传递的最小程度，以纠正精神分裂症动物模型中感觉抑制的缺陷。这些结果将有助于指导潜在候选治疗药物的临床前开发，这些药物将在精神分裂症和其他神经疾病(例如ADHD、阿尔茨海默氏症和其他涉及认知缺陷的疾病)中进行测试，这些疾病可能会受到A7 nAChRs的选择性正向调制。

DESCRIPTION (provided by applicant): Recent clinical and genetic studies provide strong evidence that spontaneous auditory gating deficits found in patients with schizophrenia maybe influenced by a7 nicotinic-acetylcholine receptors (nAChRs) located in the CA3 region of the hippocampus. Deficits in this auditory/sensory gating manifest as a sensory overload experience that can exacerbate both the positive and negative symptoms of schizophrenia. The human form of the auditory deficit is characterized by incomplete inhibitory processing of paired auditory evoked potentials spaced 50ms apart (P50). The P50 auditory deficit is genetically linked to the a7 nAChR locus on chromosome 15. Moreover, post-mortem analyses of schizophrenic brains indicate significant reductions in a7 nAChRs compared to normal individuals. The P50 auditory deficit is sensitive to cholinergic agents such as non-selective agonists (e.g., nicotine) and the selective a7 nAChRs agonist DMXB-A (GTS-21). In animal models, sensitivity to non-selective or selective a7 nAChR agonists is also blocked by selective a7 nAChR antagonists such as methyllycaconitine (MLA) or a-bungarotoxin but not a4¿2 selective antagonists such as dihydro-¿-ethroidine (DH¿E). The rodent P20-N40 auditory evoked potential (AEP) is analogous to the human P50 AEP and is a relevant model for assessing the effects of various pharmacological agents that may have utility to treat the sensory gating deficits of schizophrenia. Recent evidence using a rodent model of auditory gating deficit indicates that selective allosteric modulators of a7 nAChRs may boost the sensitivity of a7 nAChRs so that the gating deficit is rectified. However, the representative molecule in these proof-of-principle studies (PNU-120596) suffers from several attributes that make it unattractive as a therapeutic candidate. PNU-120596 is an allosteric modulator of a7 nAChRs which significantly alters the native kinetic response to agonist such that, in the presence of PNU-120596, a population of a7 nAChRs desensitized by high concentrations of nicotine can be re-sensitized. This is problematic since a7 nAChRs regulate Ca++ conductance. A large and sustained influx of Ca++ caused by the chronic presence of PNU-120596 in a treatment regimen is predicted to be neurotoxic. In contrast, our laboratory has developed selective a7 nAChR positive allosteric modulators which potentiate and preserve the native kinetics of receptor activation such that upon extended exposure to our compound desensitized a7 nAChRs remain as such. While the issue of retaining native kinetics of channel activity has been accomplished by our drug discovery efforts, we are uncertain as to what level of a7 nAChR efficacy is appropriate for therapeutic benefit in a setting where a7 nAChR function is already suboptimal (e.g., schizophrenia) while simultaneously avoiding Ca++-related neurotoxicity. The goal of the proposed research is to determine what minimum level of positive allosteric efficacy at a7 nAChRs is required to produce a reversal of P20-N40 auditory gating deficits in the DBA/2 mouse model of schizophrenia where a7 nAChR function is intrinsically compromised. Candidate molecules will be synthesized and tested in electrophysiological assays for enhancement of a7 nAChR currents elicited by nicotine to determine the structure activity relationships for maximum efficacy and potency relative to nicotine. The proposed molecules will show positive modulation of a7 nAChRs without activation of other nAChR subtypes (e.g., a4¿2, a3¿4) or other cys-loop ligand-gated ion channels (GABAA, 5HT3A). The pharmacokinetic (PK) profile of candidate molecules that fulfill our selection criteria for receptor subtype selectivity, potency and efficacy will be evaluated for whether appropriate brain levels (i.e., corresponding to maximum enhancement by the positive allosteric modulator observed in vitro) can be achieved. Completion of the proposed studies will establish the minimum extent of modulation of cholinergic transmission via a7 nAChRs necessary to correct a deficit of sensory inhibition in an animal model of schizophrenia. These results will help guide the pre-clinical development of potential candidate therapeutic agents that will be tested in schizophrenia and other neurological disorders (e.g., ADHD, Alzheimer's and other diseases involving cognitive deficit) that may be amenable to selective positive modulation of a7 nAChRs.