Sheinberger Yoni, Wassermann Rina, Khier Jasmine, Kassa Ephrem, Vaturi Linoy, Slonim Naama, Tverskoi Artem, Mandaby Aviad, Demishtein Alik, Klepfish Mordehay, Shapira-Lots Inbal, Alroy Iris
Anima Biotech, 2 Shoham St., Ramat Gan 5251003, Israel.
Int J Mol Sci. 2025 Aug 22;26(17):8139. doi: 10.3390/ijms26178139.
is a key oncogenic driver frequently overexpressed in non-small cell lung carcinoma (NSCLC) and other cancers, where its protein levels often exceed what would be expected from mRNA levels alone, suggesting post-transcriptional regulation. Strategies to inhibit function by targeting mRNA translation hold potential for therapeutics utility in Myc-dependent cancers. We developed TranslationLight, a high-content imaging platform which detects mRNA translation in human cells. Using this system, we conducted a high-throughput screen of ~100,000 compounds to identify small molecules that selectively modulate translation. Candidate compounds were evaluated by immunofluorescence, ribosome profiling, RNA sequencing, cellular thermal shift assays (CETSA), and subcellular localization studies of mRNA and RNA-binding proteins. We identified a lead compound, CMP76, that potently reduces Myc protein without substantially decreasing its mRNA abundance. Mechanistic investigations showed that the compound induces relocalization of mRNA into stress granules, accompanied by translational silencing. CETSA identified hnRNPK as a primary protein target, and compound treatment triggered its cytoplasmic relocalization together with formation of hnRNPK-containing granules colocalizing with mRNA. Analysis across cancer cell lines revealed that sensitivity to CMP76 was significantly associated with RBM42 dependency. This work establishes a novel therapeutic strategy to inhibit translation mediated by hnRNPK, offering a translationally targeted approach to cancer therapy.
是一种关键的致癌驱动因子，在非小细胞肺癌（NSCLC）和其他癌症中经常过度表达，其蛋白质水平往往超过仅由mRNA水平所预期的水平，提示存在转录后调控。通过靶向mRNA翻译来抑制其功能的策略在Myc依赖性癌症的治疗中具有潜在应用价值。我们开发了TranslationLight，这是一种用于检测人类细胞中mRNA翻译的高内涵成像平台。利用该系统，我们对约10万种化合物进行了高通量筛选，以鉴定选择性调节其翻译的小分子。候选化合物通过免疫荧光、核糖体分析、RNA测序、细胞热位移分析（CETSA）以及mRNA和RNA结合蛋白的亚细胞定位研究进行评估。我们鉴定出一种先导化合物CMP76，它能有效降低Myc蛋白水平而不显著降低其mRNA丰度。机制研究表明，该化合物诱导Myc mRNA重新定位到应激颗粒中，同时伴有翻译沉默。CETSA确定hnRNPK是主要的蛋白质靶点，化合物处理引发其细胞质重新定位以及与Myc mRNA共定位的含hnRNPK颗粒的形成。对癌细胞系的分析表明，对CMP76的敏感性与RBM42依赖性显著相关。这项工作建立了一种抑制由hnRNPK介导的Myc翻译的新型治疗策略，为癌症治疗提供了一种翻译靶向方法。
