热休克蛋白90（Hsp90）及其辅助分子促进癌症发展，靶向Hsp90具有治疗癌症的潜力。大部分相关研究都集中在靶向Hsp90的N-末端ATP结合位点上。虽然新一代抑制剂在侵袭性癌症治疗方面显示出更好的疗效，但这些抑制剂诱导细胞热休克反应（HSR）的问题被认为限制了其临床疗效。因此，具有新机制且不触发HSR的Hsp90抑制剂将具有优势。在本研究中，通过突变、化学修饰和蛋白质组学研究，我们发现辣椒素抑制Hsp90的机制。我们显示辣椒素与Hsp90的N-末端结合，并抑制其ATP酶活性。因此，辣椒素及其类似物能够体外抑制Hsp90 ATP酶依赖的孕酮受体重组。辣椒素并不诱导HSR，而是通过溶酶体和自噬通路促进Hsp70的降解。值得注意的是，辣椒素并不诱导Hsc70的降解，表明它对Hsp70具有选择性。联合应用辣椒素和Hsp90抑制剂17-AAG在细胞培养和三维瘤球模型中提高了17-AAG的抗肿瘤效果，在乳腺癌和前列腺癌模型中体现出一致性。与此一致，在硅计算对接研究中揭示了辣椒素与Hsp90的ATP结合位点的结合方式与典型的N-末端Hsp90抑制剂不同，显示出一种新的作用机制。综上所述，这些发现支持使用辣椒素作为化学支架来开发新型Hsp90 N-末端抑制剂，并且辣椒素具有作为潜在抗癌辅助治疗药物的能力。© 2023. Springer Nature Limited.

Heat shock protein 90 (Hsp90) and its co-chaperones promote cancer, and targeting Hsp90 holds promise for cancer treatment. Most of the efforts to harness this potential have focused on targeting the Hsp90 N-terminus ATP binding site. Although newer-generation inhibitors have shown improved efficacy in aggressive cancers, induction of the cellular heat shock response (HSR) by these inhibitors is thought to limit their clinical efficacy. Therefore, Hsp90 inhibitors with novel mechanisms of action and that do not trigger the HSR would be advantageous. Here, we investigated the mechanism by which capsaicin inhibits Hsp90. Through mutagenesis, chemical modifications, and proteomic studies, we show that capsaicin binds to the N-terminus of Hsp90 and inhibits its ATPase activity. Consequently, capsaicin and its analogs inhibit Hsp90 ATPase-dependent progesterone receptor reconstitution in vitro. Capsaicin did not induce the HSR, instead, it promoted the degradation of Hsp70 through the lysosome-autophagy pathway. Remarkably, capsaicin did not induce degradation of the constitutively expressed cognate Hsc70, indicating selectivity for Hsp70. Combined treatments of capsaicin and the Hsp90 inhibitor 17-AAG improved the anti-tumor efficacy of 17-AAG in cell culture and tridimensional tumor spheroid growth assays using breast and prostate cancer models. Consistent with this, in silico docking studies revealed that capsaicin binding to the ATP binding site of Hsp90 was distinct from classical N-terminus Hsp90 inhibitors, indicating a novel mechanism of action. Collectively, these findings support the use of capsaicin as a chemical scaffold to develop novel Hsp90 N-terminus inhibitors as well as its ability to be a potential cancer co-therapeutic.© 2023. Springer Nature Limited.