Liu Chou J, Priest Birgit T, Bugianesi Randal M, Dulski Paula M, Felix John P, Dick Ivy E, Brochu Richard M, Knaus Hans-Guenther, Middleton Richard E, Kaczorowski Gregory J, Slaughter Robert S, Garcia Maria L, Köhler Martin G
Department of Ion Channels, Merck Research Laboratories, Rahway, NJ 07065, USA.
Assay Drug Dev Technol. 2006 Feb;4(1):37-48. doi: 10.1089/adt.2006.4.37.
Clinical treatment of neuropathic pain can be achieved with a number of different drugs, some of which interact with all members of the voltage-gated sodium channel (NaV1) family. However, block of central nervous system and cardiac NaV1 channels can cause dose-limiting side effects, preventing many patients from achieving adequate pain relief. Expression of the tetrodotoxin-resistant NaV1.8 subtype is restricted to small-diameter sensory neurons, and several lines of evidence indicate a role for NaV1.8 in pain processing. Given these features, NaV1.8 subtype-selective blockers are predicted to be efficacious in the treatment of neuropathic pain and to be associated with fewer adverse effects than currently available therapies. To facilitate the identification of NaV1.8-specific inhibitors, we stably expressed the human NaV1.8 channel together with the auxiliary human beta1 subunit (NaV beta1) in human embryonic kidney 293 cells. Heterologously expressed human NaV1.8/NaV beta1 channels display biophysical properties that are similar to those of tetrodotoxin-resistant channels present in mouse dorsal root ganglion neurons. A membrane potential, fluorescence resonance energy transfer-based functional assay on a fluorometric imaging plate reader (FLIPR-Tetra, Molecular Devices, Sunnyvale, CA) platform has been established. This highcapacity assay is sensitive to known state-dependent NaV1 modulators and can be used to identify novel and selective NaV1.8 inhibitors.
神经性疼痛的临床治疗可以通过多种不同药物实现，其中一些药物可与电压门控钠通道（NaV1）家族的所有成员相互作用。然而，阻断中枢神经系统和心脏的NaV1通道会导致剂量限制性副作用，使许多患者无法获得足够的疼痛缓解。对河豚毒素不敏感的NaV1.8亚型的表达仅限于小直径感觉神经元，多项证据表明NaV1.8在疼痛处理中发挥作用。鉴于这些特性，预计NaV1.8亚型选择性阻滞剂在治疗神经性疼痛方面有效，且与现有疗法相比副作用更少。为便于鉴定NaV1.8特异性抑制剂，我们在人胚肾293细胞中稳定表达了人NaV1.8通道以及辅助性人β1亚基（NaVβ1）。异源表达的人NaV1.8/NaVβ1通道表现出与小鼠背根神经节神经元中存在的对河豚毒素不敏感通道相似的生物物理特性。已在荧光成像板读数器（FLIPR-Tetra，Molecular Devices，加利福尼亚州桑尼维尔）平台上建立了基于膜电位、荧光共振能量转移的功能测定法。这种高通量测定法对已知的状态依赖性NaV1调节剂敏感，可用于鉴定新型和选择性NaV1.8抑制剂。
