Halofuginone (HF) is a phase 2 clinical compound that inhibits the glutamyl-prolyl-tRNA synthetase (EPRS) thereby inducing the integrated stress response (ISR). Here, we report that halofuginone indeed triggers the predicted canonical ISR adaptations, consisting of attenuation of protein synthesis and gene expression reprogramming. However, the former is surprisingly atypical and occurs to a similar magnitude in wild-type cells, cells lacking GCN2 and those incapable of phosphorylating eIF2α. Proline supplementation rescues the observed HF-induced changes indicating that they result from inhibition of EPRS. The failure of the GCN2-to-eIF2α pathway to elicit a measurable protective attenuation of translation initiation allows translation elongation defects to prevail upon HF treatment. Exploiting this vulnerability of the ISR, we show that cancer cells with increased proline dependency are more sensitive to halofuginone. This work reveals that the consequences of EPRS inhibition are more complex than anticipated and provides novel insights into ISR signaling, as well as a molecular framework to guide the targeted development of halofuginone as a therapeutic.

中文翻译：

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细胞对 halofuginone 的反应揭示了综合应激反应的 GCN2 分支的脆弱性。

卤夫酮 (HF) 是一种 2 期临床化合物，可抑制谷氨酰-脯氨酰-tRNA 合成酶 (EPRS)，从而诱导综合应激反应 (ISR)。在这里，我们报告 halofuginone 确实触发了预测的典型 ISR 适应，包括蛋白质合成的衰减和基因表达重编程。然而，前者出人意料地非典型，并且在野生型细胞、缺乏 GCN2 的细胞和不能磷酸化 eIF2α 的细胞中发生的程度相似。脯氨酸补充剂挽救了观察到的 HF 诱导的变化，表明它们是由 EPRS 的抑制作用引起的。GCN2-to-eIF2α 通路未能引起可测量的翻译起始保护性减弱，这使得翻译延伸缺陷在 HF 治疗中占上风。利用ISR的这个漏洞，我们表明，脯氨酸依赖性增加的癌细胞对 halofuginone 更敏感。这项工作揭示了 EPRS 抑制的后果比预期的更复杂，并提供了对 ISR 信号传导的新见解，以及指导卤夫酮作为治疗药物的靶向开发的分子框架。