Despite the vital role of iron and vulnerability of iron metabolism in disease states, it remains largely unknown whether chemicals interacting with cellular proteins are responsible for perturbation of iron metabolism. We previously demonstrated that cisplatin was an inhibitor of the iron regulatory system by blocking IRP2 (iron regulatory protein 2) binding to an iron-responsive element (IRE) located in the 3′- or 5′-UTR (untranslated region) of key iron metabolism genes such as transferrin receptor 1 (TfR1) and ferritin mRNAs. To guide the development of new chemical probes to modulate the IRP-IRE regulatory system, we used an artificial intelligence (AI)-based ligand design and screened a chemical library composed of cysteine-reactive warheads. Using wild type and mutant IRE-luciferase reporter cells, we identified new IRP-IRE inhibitors such as V004-0872 harboring chloroacetamide, while its analog V011-6261 with chloropropanamide completely lost the inhibitory activity. V004-0872 inhibited the human IRP2 via Cys512 and caused decreased iron levels through reciprocal TfR1 downregulation and ferritin upregulation. V004-0872 increased production of mitochondrial reactive oxygen species (ROS) and exhibited cytotoxicity that was inhibited by N-acetyl cysteine but not the ferroptosis inhibitor ferrostatin-1. Furthermore, we found that widely used haloketone protease inhibitors and acetamide herbicides inhibit the IRP-IRE system. Since IRP2 overexpression is responsible for iron excess conditions to promote growth of several cancers and exacerbation of iron-overload diseases, these results and new compounds lay the groundwork for new reagents and strategies to limit the availability of iron and oxidative stress in iron-overloaded disease conditions.

中文翻译：

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通过与铁调节系统相互作用的新化学物质调节铁代谢


尽管铁起着至关重要的作用并且铁代谢在疾病状态中的脆弱性，但与细胞蛋白相互作用的化学物质是否是导致铁代谢扰动的原因在很大程度上仍然未知。我们之前证明，顺铂是通过阻断 IRP2（铁调节蛋白 2）与位于关键铁代谢基因（如转铁蛋白受体 1 （TfR1） 和铁蛋白 mRNA）的 3′-或 5′-UTR（非翻译区）的铁反应元件 （IRE） 结合而成为铁调节系统的抑制剂。为了指导开发新的化学探针来调节 IRP-IRE 调节系统，我们使用了基于人工智能 （AI） 的配体设计，并筛选了由半胱氨酸反应性弹头组成的化学库。使用野生型和突变型 IRE-荧光素酶报告细胞，我们鉴定了新的 IRP-IRE 抑制剂，例如含有氯乙酰胺的 V004-0872，而其具有氯丙酰胺的类似物 V011-6261 完全失去了抑制活性。V004-0872 通过 Cys512 抑制人 IRP2，并通过 TfR1 上调和铁蛋白上调导致铁水平降低。V004-0872 增加了线粒体活性氧 （ROS） 的产生，并表现出被 N-乙酰半胱氨酸抑制的细胞毒性，而铁死亡抑制剂 ferrostatin-1 没有抑制。此外，我们发现广泛使用的卤酮蛋白酶抑制剂和乙酰胺除草剂抑制 IRP-IRE 系统。由于 IRP2 过表达是导致铁过量条件促进多种癌症生长和铁过载疾病恶化的原因，因此这些结果和新化合物为新试剂和策略奠定了基础，以限制铁过载疾病条件下铁的可用性和氧化应激。