Therapeutic Na+ channel blockers: Receptor & Drug Design

描述(申请人提供)：该项目的长期目标是开发与疼痛控制相关的长效Na+通道阻滞剂，并更好地了解这些阻滞剂的作用机制。传统的局麻药(LAS)由于其神经阻滞时间不足，对慢性或顽固性癌症疼痛往往无能为力。我们的具体目标是(1)鉴定和合成有效阻断开放和/或失活的Na+通道的化合物，(2)评估有效的Na+通道阻滞剂的使用和状态依赖的阻断，(3)测试它们在体内作为长效LAS的效力，以及(4)在Na+通道α亚基内定位它们的受体位置。最近的筛查发现了几种新的铅结构，它们以高亲和力阻断开放的Na+通道。早期筛查显示，口服治疗神经病理性疼痛的药物，如阿米替林、氟卡胺和美西律，在其治疗血浆浓度下也能有效地阻断开放的Na+通道。我们推测，这些药物由于高亲和力阻断开放的Na+通道而减轻了受损神经中的异位高频放电。阿米替林过于有效地阻断了失活的Na+通道，实际上起到了长效LA的作用。我们计划基于这些先导结构来鉴定和合成新型的开放和/或失活的通道阻滞剂。在人HEK293细胞表达的野生型和/或失活突变的Na+通道中，将确定它们在重复脉冲期间对Na+电流的使用依赖性阻断以及静息、开放和失活通道阻断的50%抑制浓度(IC50)。通过多种途径对大鼠或绵羊注射强效Na+通道阻滞剂前后神经阻滞的感觉和运动功能进行评估。最后，我们计划通过定点突变和通过计算机模拟配体-受体复合体来界定Na+通道α亚基中选定的阻滞剂的受体。这些信息将反过来促进基于受体的药物设计。总之，这些研究应该为开发选择性靶向开放和/或失活的Na+通道的长效LAS提供新的先导结构。这些药物，无论是口服还是局部注射，对患有慢性和顽固性癌症疼痛的患者都可能是有益的。

DESCRIPTION (provided by applicant): The long-term objectives of this project are to develop long-acting Na+ channel blockers pertinent for pain management and to gain a better understanding of how these blockers work mechanistically. Traditional local anesthetics (LAs) are often inept for chronic or intractable cancer pain due to their insufficient duration of nerve block. Our specific aims are (1) to identify and synthesize compounds that potently block the open and/or inactivated Na+ channels, (2) to assess the use- and state-dependent block of potent Na+ channel blockers, (3) to test their in vivo potency as long-acting LAs, and (4) to map their receptor site within the Na+ channel alpha-subunit. Recent screening has identified several new lead structures that block open Na+ channels with high affinities. Earlier screening demonstrated that drugs taken orally for neuropathic pain, such as amitriptyline, flecainide, and mexiletine, also block open Na+ channels effectively at their therapeutic plasma concentrations. We hypothesize that these drugs alleviate ectopic high-frequency discharges found in injured nerves due to their high-affinity block of open Na+ channels. Amitriptyline, which too potently blocks the inactivated Na+ channels, indeed acts as a long-acting LA. We plan to identify and synthesize novel open and/or inactivated-channel blockers based on these lead structures. Their use-dependent block of Na+ currents during repetitive pulses and the 50% inhibitory concentration (IC50) of resting-, open-, and inactivated-channel block will be determined in wild-type and/or in inactivation-deficient mutant Na+ channels expressed in human HEK293 cells. Sensory and motor functions of nerve block will be evaluated in rats or sheep before and after injection of potent Na + channel blockers via various routes. Finally, we plan to delimit the receptor for selected blockers within the Na+ channel alpha-subunit by site-directed mutagenesis and by computer simulation of the ligand-receptor complex. This information will in turn facilitate receptor-based drug design. Together, these studies should provide us new lead structures for the development of long-acting LAs that selectively target open and/or inactivated Na+ channels. Such drugs, either taken orally or injected locally, may be beneficial for patients with chronic and intractable cancer pain.