Synthesis and Target Identification of Benzoxepane Derivatives as Potential Anti-neuroinflammatory Agents for Ischemic Stroke.

Inspired by natural anti-inflammatory benzoxepanes, a series of new benzoxepane derivatives were designed and synthesized, and 10i emerged as the most effective compound in vitro with low toxicity. Further in vivo evaluation revealed that 10i could both ameliorate sickness behaviour through anti-inflammation in LPS-induced neuroinflammatory mice model and ameliorate cerebral ischemic injury through anti-neuroinflammation in rats subjected to transient middle cerebral artery occlusion. Encouraged by the promising results, target fishing of 10i was then performed by design of photo-affinity probes, followed by photo-cross-linking, click reaction, and LC-MS/MS, leading to identification of PKM2 as a key target protein responsible for anti-inflammatory effect of 10i. Furthermore, 10i exhibited anti-neuroinflammatory effect in vitro and in vivo via inhibiting PKM2-mediated glycolysis and NLRP3 activation, indicating PKM2 as a novel target for neuroinflammation and its related brain disorders. In addition, 10i encompassed much more safety profile compared to shikonin, a reported PKM2 inhibitor, suggesting that 10i could be used as a lead compound targeting PKM2 for the treatment of inflammation-related diseases such as ischemic stroke.