Small molecule inhibitors of PSD95-nNOS protein-protein interactions as novel analgesics.
Small molecule inhibitors of PSD95-nNOS protein-protein interactions as novel analgesics.
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DOI：
10.1016/j.neuropharm.2015.05.038
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发表时间：
2015-10
影响因子：
4.7
通讯作者：
Hohmann AG
中科院分区：
文献类型：
--
作者：
Lee WH;Xu Z;Ashpole NM;Hudmon A;Kulkarni PM;Thakur GA;Lai YY;Hohmann AG
NMDA 受体 (NMDAR) 信号传导的异常增加通过激活神经元一氧化氮合酶 (nNOS) 并产生一氧化氮 (NO)，导致中枢神经系统敏化和慢性疼痛。由于支架蛋白突触后密度 95kDA (PSD95) 将 nNOS 与 NMDAR 联系在一起，因此 PSD95-nNOS 复合物代表了治疗靶点。小分子抑制剂 IC87201 (EC50: 23.94 µM) 和 ZL006 (EC50: 12.88 µM) 直接抑制 AlphaScreen 中纯化的 PSD95 和 nNOS 蛋白的结合，而不改变 PSD95 与 ErbB4 的结合。两种 PSD95-nNOS 抑制剂均可抑制谷氨酸诱导的细胞死亡，其功效与 MK-801 相当。 IC87201和ZL006优先抑制福尔马林试验中的2A期疼痛行为，并抑制足底内完全弗氏佐剂给药引起的异常性疼痛。 IC87201和ZL006抑制由化疗药物紫杉醇引起的机械性和冷异常性疼痛（IC87201和ZL006的ED50分别为2.47和0.93mg/kg i.p.）。 PSD95-nNOS 干扰剂的功效与 MK-801 相似。运动共济失调效应由 MK-801 诱导，但不是由 ZL006 或 IC87201 诱导。最后，MK-801 在甩尾测试中产生痛觉过敏，而 IC87201 和 ZL006 没有改变基础伤害性阈值。我们的研究证实了使用 AlphaScreen 和纯化的蛋白质对来确定和量化蛋白质-蛋白质相互作用的破坏的效用。我们的结果证明了 ZL006 以前未被认识的抗伤害功效，并使用两种小分子建立了 PSD95-nNOS 抑制剂在治疗神经性疼痛和炎性疼痛方面的广泛应用。总的来说，我们的结果表明，破坏 PSD95-nNOS 蛋白质-蛋白质相互作用可以有效减轻病理性疼痛，而不会产生与 NMDAR 拮抗剂相关的不良副作用（即运动性共济失调）。
Aberrant increases in NMDA receptor (NMDAR) signaling contributes to central nervous system sensitization and chronic pain by activating neuronal nitric oxide synthase (nNOS) and generating nitric oxide (NO). Because the scaffolding protein postsynaptic density 95kDA (PSD95) tethers nNOS to NMDARs, the PSD95-nNOS complex represents a therapeutic target. Small molecule inhibitors IC87201 (EC5O: 23.94 µM) and ZL006 (EC50: 12.88 µM) directly inhibited binding of purified PSD95 and nNOS proteins in AlphaScreen without altering binding of PSD95 to ErbB4. Both PSD95-nNOS inhibitors suppressed glutamate-induced cell death with efficacy comparable to MK-801. IC87201 and ZL006 preferentially suppressed phase 2A pain behavior in the formalin test and suppressed allodynia induced by intraplantar complete Freund’s adjuvant administration. IC87201 and ZL006 suppressed mechanical and cold allodynia induced by the chemotherapeutic agent paclitaxel (ED50s: 2.47 and 0.93 mg/kg i.p. for IC87201 and ZL006, respectively). Efficacy of PSD95-nNOS disruptors was similar to MK-801. Motor ataxic effects were induced by MK-801 but not by ZL006 or IC87201. Finally, MK-801 produced hyperalgesia in the tail-flick test whereas IC87201 and ZL006 did not alter basal nociceptive thresholds. Our studies establish the utility of using AlphaScreen and purified protein pairs to establish and quantify disruption of protein-protein interactions. Our results demonstrate previously unrecognized antinociceptive efficacy of ZL006 and establish, using two small molecules, a broad application for PSD95-nNOS inhibitors in treating neuropathic and inflammatory pain. Collectively, our results demonstrate that disrupting PSD95-nNOS protein-protein interactions is effective in attenuating pathological pain without producing unwanted side effects (i.e. motor ataxia) associated with NMDAR antagonists.