College of Pharmacy & Graduate School of Pharmaceutical Sciences, Ewha Womans University, Seoul 03760, Korea.
College of Pharmacy, CHA University, Pocheon 11160, Korea.
J Adv Res. 2023 May;47:173-187. doi: 10.1016/j.jare.2022.08.003. Epub 2022 Aug 10.
HER2 overexpression induces cancer aggression and frequent recurrences in many solid tumors. Because HER2 overproduction is generally followed by gene amplification, inhibition of protein-protein interaction (PPI) between transcriptional factor ELF3 and its coactivator MED23 has been considered an effective but challenging strategy.
This study aimed to determine the hotspot of ELF3-MED23 PPI and further specify the essential residues and their key interactions in the hotspot which are controllable by small molecules with significant anticancer activity.
Intensive biological evaluation methods including SEAP, fluorescence polarization, LC-MS/MS-based quantitative, biosensor, GST-pull down assays, and in silico structural analysis were performed to determine hotspot of ELF3-MED23 PPI and to elicit YK1, a novel small molecule PPI inhibitor. The effects of YK1 on possible PPIs of MED23 and the efficacy of trastuzumab were assessed using cell culture and tumor xenograft mouse models.
ELF3-MED23 PPI was found to be specifically dependent on H-bondings between D400, H449 of MED23 and W138, I140 of ELF3 for upregulating HER2 gene transcription. Employing YK1, we confirmed that interruption on these H-bondings significantly attenuated the HER2-mediated oncogenic signaling cascades and exhibited significant in vitro and in vivo anticancer activity against HER2-overexpressing breast and gastric cancers even in their trastuzumab refractory clones.
Our approach to develop specific ELF3-MED23 PPI inhibitor without interfering other PPIs of MED23 can finally lead to successful development of a drug resistance-free compound to interrogate HER2 biology in diverse conditions of cancers overexpressing HER2.
HER2 过表达可诱导许多实体肿瘤的癌症侵袭和频繁复发。由于 HER2 过表达通常伴随着基因扩增，因此抑制转录因子 ELF3 与其共激活因子 MED23 之间的蛋白质-蛋白质相互作用（PPI）已被认为是一种有效但具有挑战性的策略。
本研究旨在确定 ELF3-MED23 PPI 的热点，并进一步确定热点中可由具有显著抗癌活性的小分子控制的关键残基及其关键相互作用。
采用 SEAP、荧光偏振、基于 LC-MS/MS 的定量、生物传感器、GST 下拉测定和计算结构分析等强化生物学评估方法，确定 ELF3-MED23 PPI 的热点，并筛选出新型小分子 PPI 抑制剂 YK1。使用细胞培养和肿瘤异种移植小鼠模型评估 YK1 对 MED23 可能的 PPI 的影响以及曲妥珠单抗的疗效。
发现 ELF3-MED23 PPI 特异性依赖于 MED23 的 D400、H449 和 ELF3 的 W138、I140 之间的氢键，以上调 HER2 基因转录。使用 YK1，我们证实中断这些氢键可显著减弱 HER2 介导的致癌信号级联，并在体外和体内对 HER2 过表达的乳腺癌和胃癌表现出显著的抗癌活性，即使在其曲妥珠单抗耐药克隆中也是如此。
我们开发特异性 ELF3-MED23 PPI 抑制剂而不干扰 MED23 其他 PPI 的方法最终可以成功开发出一种无耐药化合物，以在过表达 HER2 的癌症的各种条件下研究 HER2 生物学。
