A novel TWIK2 channel inhibitor binds at the bottom of the selectivity filter and protects against LPS-induced experimental endotoxemia in vivo.
A novel TWIK2 channel inhibitor binds at the bottom of the selectivity filter and protects against LPS-induced experimental endotoxemia in vivo.
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DOI：
10.1016/j.bcp.2023.115894
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发表时间：
2023-10
影响因子：
5.8
通讯作者：
Yuanxing Zhi;Xiaoyan Wu;Yanshan Chen;Xingyuan Chen;Xiangyu Chen;Hui Luo;Xin Yi;Xiuling Lin;Liang Ma;Yao Chen;Ying Cao;Fengxian Li;Pingzheng Zhou
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文献类型：
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作者：
Yuanxing Zhi;Xiaoyan Wu;Yanshan Chen;Xingyuan Chen;Xiangyu Chen;Hui Luo;Xin Yi;Xiuling Lin;Liang Ma;Yao Chen;Ying Cao;Fengxian Li;Pingzheng Zhou
TWIK2通道在NLRP3炎性小体激活中起关键作用，TWIK2通道缺陷小鼠可保护败血症和炎症性肺损伤。然而，TWIK2通道的抑制剂目前还处于早期开发阶段，TWIK2通道化学调控的分子决定因素仍未被探索。在本研究中，我们利用全细胞膜片钳技术鉴定了NPBA及其衍生物NPBA-4对TWIK2通道的有效和选择性抑制作用。此外，TWIK2通道P1和P2(即T106A、T214A)选择性过滤器最后残基的突变基本上消除了NPBA对TWIK2通道的影响。我们的数据表明，NPBA通过与选择性过滤器底部的结合来阻断TWIK2通道，这也得到了分子对接预测的支持。此外，我们还发现NPBA能显著抑制巨噬细胞内NLRP3炎性小体的激活，减轻内毒素血症和体内脏器损伤。值得注意的是，NPBA对内毒素血症的保护作用在Kcnk6-/-小鼠中被取消。综上所述，我们的研究发现了一系列新的TWIK2通道抑制剂，并揭示了它们与TWIK2通道相互作用的不同分子决定因素。这些发现对TWIK2通道的药理作用机制提供了新的见解，并为开发选择性TWIK2通道调节剂治疗相关炎症性疾病提供了机会。
TWIK2 channel plays a critical role in NLRP3 inflammasome activation and mice deficient in TWIK2 channel are protected from sepsis and inflammatory lung injury. However, inhibitors of TWIK2 channel are currently in an early stage of development, and the molecular determinants underlying the chemical modulation of TWIK2 channel remain unexplored. In this study, we identifiedNPBAand the synthesized derivativeNPBA-4potently and selectively inhibited TWIK2 channel by using whole-cell patch clamp techniques. Furthermore, the mutation of the last residues of the selectivity filter in both P1 and P2 (i.e., T106A, T214A) of TWIK2 channel substantially abolished the effect ofNPBAon TWIK2 channel. Our data suggest thatNPBAblocked TWIK2 channel through binding at the bottom of the selectivity filter, which was also supported by molecular docking prediction. Moreover, we found thatNPBAsignificantly suppressed NLRP3 inflammasome activation in macrophages and alleviated LPS-induced endotoxemia and organ injury in vivo. Notably, the protective effects ofNPBAagainst LPS-induced endotoxemia were abolished inKcnk6-/-mice. In summary, our study has uncovered a series of novel inhibitors of TWIK2 channel and revealed their distinct molecular determinants interacting TWIK2 channel. These findings provide new insights into the mechanisms of pharmacological action on TWIK2 channel and opportunities for the development of selective TWIK2 channel modulators to treat related inflammatory diseases.