The translation termination factor GSPT1 is a phenotypically relevant off-target of heterobifunctional phthalimide degraders. The translation termination factor GSPT1 is a phenotypically relevant off-target of heterobifunctional phthalimide degraders

Protein degradation is an emerging therapeutic strategy with a unique molecular pharmacology that enables the disruption of all functions associated with a target. This is particularly relevant for proteins depending on molecular scaffolding, such as transcription factors or receptor tyrosine kinases (RTKs). To address tractability of multiple RTKs for chemical degradation by the E3 ligase cereblon (CRBN), we synthesized a series of phthalimide degraders based on the promiscuous kinase inhibitors sunitinib and PHA665752. While both series failed to induce degradation of their consensus targets, individual molecules displayed pronounced efficacy in leukemia cell lines. Orthogonal target identification supported by molecular docking led us to identify the translation termination factor G1 to S phase transition 1 (GSPT1) as a converging off-target resulting from inadvertent E3 ligase modulation. This research highlights the importance of monitoring degradation events that are independent of the respective targeting ligand as a unique feature of small-molecule degraders. Overall design: RNA-seq of MOLT4 cells treated with sunitinib and the phthalimide conjugate MI-389

蛋白质降解是一种新兴的治疗策略，其独特的分子药理学特性可实现对靶点相关所有生物学功能的完全阻断。这一点对于依赖分子脚手架结构发挥功能的蛋白质而言尤为关键，例如转录因子或受体酪氨酸激酶(RTKs)。为探究多种受体酪氨酸激酶(RTKs)经E3泛素连接酶cereblon (CRBN)介导化学降解的可行性，我们基于两种多靶点激酶抑制剂——舒尼替尼与PHA665752——合成了一系列邻苯二甲酰亚胺类降解剂。尽管这两类化合物均未能诱导其预设靶点的降解，但部分单体分子在白血病细胞系中展现出显著的降解活性。通过分子对接辅助的正交靶标鉴定，我们发现翻译终止因子细胞周期G1到S期转换蛋白1(GSPT1)是由意外的E3连接酶调控所产生的共同脱靶效应蛋白。本研究凸显了监测与靶向配体无关的降解事件的重要性——这正是小分子降解剂的独特特征之一。整体实验设计：将MOLT4细胞分别用舒尼替尼及其邻苯二甲酰亚胺缀合物MI-389处理后，进行RNA测序。