Optimization of KCa2 Channel Activators as Neuroscience Tools and Potential Drugs

描述（由申请人提供）：小电导钙激活钾通道由KCa2.1 - 2.3（= SK1 - 3）基因编码，最为人所知的是神经元中的apamin敏感性介质后超极化电流（mAHP）。根据神经元的类型，KCa2通道的功能从确定瞬时放电率、设置紧张性放电频率到调节突发放电和潜在的儿茶酚胺释放。因此，KCa通道的药理学调节提供了显著影响神经元兴奋性的机会。虽然KCa 2通道阻滞剂如蜂毒蜂毒肽增加放电率并诱导啮齿动物癫痫发作，但KCa 2通道激活剂减慢神经元放电，因此已提出用于治疗以过度兴奋为特征的CNS疾病，如癫痫、共济失调和神经性疼痛。然而，这一令人信服的治疗假设目前仍在很大程度上未经检验，因为现有的KCa 2通道激活剂如EBIO（EC 50 300 M）或NS 309都不适合体内使用。μ 使用神经保护药物利鲁唑作为合成模板，我们的实验室最近设计了SKA-31（EC50 2 μ M），这是第一个KCa 2通道激活剂，其效力足以在体内使用，并与NIH抗惊厥筛选计划（ASP）合作证明该化合物及其几种衍生物是有效的抗惊厥药。不幸的是，SKA-31还激活血管内皮上表达的KCa3.1通道，从而降低小鼠的血压。使用经典的药物化学和自动和手动电生理学的组合，我们打算进一步探索SKA 31和EBIO周围的结构活性关系，以提高对KCa2的选择性超过KCa3.1以及效力和脑渗透。然后将评价最佳新KCa 2激活剂对一组克隆离子通道的选择性，并使用海马切片表征其对天然KCa 2通道的活性。化合物选择性激活克隆和天然KCa2通道将进一步评估药代动力学特性和脑渗透在大鼠中使用HPLC/MS。同时，我们将提交选定的化合物的ASP，其中化合物将在急性癫痫发作模型中进行测试。有希望的化合物将在杏仁核点燃的小鼠和红藻氨酸诱导的癫痫大鼠中进行测试，这两种模型更能代表人类难治性癫痫。脑渗透剂和潜在亚型选择性KCa 2通道激活剂的设计将有助于验证KCa 2通道作为治疗癫痫的新型药理学靶点，并将进一步为科学界提供工具化合物来研究KCa 2通道在共济失调，神经性疼痛和认知中的作用。 公共卫生相关性：项目叙述KCa2钾通道在决定神经元兴奋性方面发挥重要作用。因此，这些通道的激活剂已被建议作为用于治疗以神经元过度兴奋为特征的疾病如癫痫和共济失调的新疗法。在这笔资助的帮助下，我们将尝试设计一种KCa2通道激活剂，这种激活剂具有足够的有效性和选择性，可以用作科学工具化合物，甚至可以开发成药物。

DESCRIPTION (provided by applicant): Small-conductance calcium activated potassium channels are encoded by the KCa2.1-2.3 (= SK1-3) genes and are best known for underlying the apamin-sensitive medium afterhyperpolarization current (mAHP) in neurons. Depending on the type of neuron, the function of KCa2 channels varies from determining instantaneous firing rates, over setting tonic firing frequencies, to regulating burst firing and potentially catecholamine release. Pharmacological modulation of KCa channels therefore offers the opportunity to significantly affect neuronal excitability. While KCa2 channel blockers like the bee venom apamin increase firing rates and induce seizures in rodents, KCa2 channel activators slow down neuronal firing and have therefore been proposed for the treatment of CNS disorders that are characterized by hyperexcitability such as epilepsy, ataxia, and neuropathic pain. However, this compelling therapeutic hypothesis currently remains largely untested because none of the existing KCa2 channel activators such as EBIO (EC50 300 μM) or NS309 are suitable for in vivo use. Using the neuroprotective drug riluzole as a synthetic template, our laboratory recently designed SKA-31 (EC50 2 uM), the first KCa2 channel activator, which is potent enough to be used in vivo, and demonstrated in collaboration with the NIH Anticonvulsant Screening Program (ASP) that the compound and several of its derivatives are effective anticonvulsants. Unfortunately, SKA-31 also activates KCa3.1 channels, which are expressed on vascular endothelium, and thus reduces blood pressure in mice. Using a combination of classical medicinal chemistry and automated and manual electrophysiology we intend to further explore the structure activity relationship around SKA 31 and EBIO in order to improve selectivity for KCa2 over KCa3.1 as well as potency and brain penetration. The best new KCa2 activators will then be evaluated for selectivity over a panel of cloned ion channels and characterized for activity on native KCa2 channels using hippocampal slices. Compounds selectively activating cloned and native KCa2 channels will further be evaluated for pharmacokinetic properties and brain penetration in rats using HPLC/MS. In parallel, we will submit selected compounds to the ASP, where the compounds will we tested in acute seizure models. Promising compounds will then be tested in amygdala kindled mice and rats with kainate-induced epilepsy, two models that are more representative of human refractory epilepsy. The design of brain penetrant and potentially subtype selective KCa2 channel activators would help to validate KCa2 channels as novel pharmacological targets for the treatment of epilepsy and would further provide the scientific community with tool compounds to study the role of KCa2 channels in ataxia, neuropathic pain and cognition. PUBLIC HEALTH RELEVANCE: Project Narrative KCa2 potassium channels play important roles in determining neuronal excitability. Activators of these channels have therefore been suggested as new therapeutics for the treatment of diseases that are characterized by neuronal hyperexcitability such as epilepsy and ataxia. With the help of this grant we will attempt to design a KCa2 channel activator that is potent and selective enough to be used as a scientific tool compound or even to be developed into a drug.