Development of ubiquitin-specific protease 8 (USP8) inhibitors

项目摘要 泛素特异性蛋白水解酶8(USP8)是一种半胱氨酸蛋白酶，可使受体酪氨酸激酶去泛化。USP8 调节内吞作用，并通过这一活性调节自噬- 溶酶体系统。在肿瘤学中，USP8已被证明可以去泛素和稳定突变的表皮生长。 因子受体(EGFR)在非小细胞肺癌中的表达初步研究表明，USP8抑制作用 促进泛素介导的突变型EGFR在NSCLC细胞系和原发患者样本中的降解 对EGFR激酶抑制敏感且耐受。这些结果表明，抑制UPS8可以提供一种 克服非小细胞肺癌患者吉非替尼耐药的治疗策略。然而，这些中使用的USP8抑制剂 初步研究是一种早期世代抑制剂，缺乏高质量的 探测化合物。 该研究计划的目标是开发第一个具有所需效力的可逆USP8抑制剂， 以探讨USP8作为非小细胞肺癌治疗靶点的潜力。第一 目的是对从高通量筛选中鉴定的铅支架进行药物化学优化。我 将设计每个化合物的类似物，以广泛地研究分子的活性和所需的区域 可以被修改以调节诸如溶解度和稳定性等性质的位置。对于这些研究，有一项 分子中的位置将被一次修改，然后生产性变化将被结合。在第二个目标中， 这些抑制剂将根据它们的细胞靶标结合、抗增殖作用、 以及对突变型和野生型NSCLC细胞中EGFR稳定性的影响。在第三个目标中，我将使用 新开发的USP8抑制剂作为探针在表型、蛋白质组和转录组分析中广泛应用 一组癌细胞系，以公正的方式发现新的USP8生物学。因此，这项提案概述了 首个用作细胞探针的具有所需效力和选择性的USP8抑制剂的开发 研究USP8生物学，为抑制USP8作为非小细胞肺癌的治疗策略提供证据。

Project Summary Ubiquitin specific protease 8 (USP8) is a cysteine protease that can deubiquinate receptor tyrosine kinases. USP8 regulates endocytosis and through this activity modulates the degradation of diverse substrates by the autophagy- lysosome system. In oncology, USP8 has been shown to deubiquinate and stabilize mutant epidermal growth factor receptor (EGFR) in non-small cell lung cancer (NSCLC). Preliminary studies show that USP8 inhibition promotes ubiquitin-mediated degradation of mutant EGFR in NSCLC cell lines and primary patient samples both sensitive and resistant to EGFR kinase inhibition. These results suggest that UPS8 inhibition could provide a therapeutic strategy for overcoming gefitinib-resistance in NSCLC. However, the USP8 inhibitor used in these preliminary studies is an early generation inhibitor that lacks the requisite potency and selectivity of a high-quality probe compound. The goal of this research program is to develop the first reversible USP8 inhibitors with the requisite potency, selectivity, and cell permeability to interrogate the potential of USP8 as a therapeutic target in NSCLC. The first aim is to perform medicinal chemistry optimization of lead scaffolds identified from a high throughput screen. I will design analogs of each compound to broadly survey regions of the molecule that are required for activity and positions that can be modified to modulate properties such as solubility and stability. For these studies, one position in the molecule will be modified at a time then productive changes will be combined. In the second aim, these inhibitors will be characterized with respect to their cellular target engagement, anti-proliferative effects, and effects on the stability of EGFR in mutant and wild type NSCLC cell lines. In the third aim, I will use the newly developed USP8 inhibitors as probes in phenotypic, proteomic, and transcriptomic assays across a broad set of cancer cell lines to uncover new USP8 biology in an unbiased manner. This proposal therefore outlines the development of the first USP8 inhibitors with the required potency and selectivity to be used as cellular probes to study USP8 biology, and contributes to evidence for USP8 inhibition as a therapeutic strategy for NSCLC.