Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College , Beijing, China.
Autophagy. 2020 Oct;16(10):1823-1837. doi: 10.1080/15548627.2019.1709762. Epub 2020 Jan 27.
Although macroautophagy/autophagy is involved in hepatocellular carcinoma (HCC) initiation and development and has been identified as a mechanism of HCC therapy resistance, the role of ULK1 (unc-51 like autophagy activating kinase 1) in HCC remains unclear. Here, we report that both knockdown and knockout of inhibited human HCC cell proliferation and invasion, and deletion abrogated tumor growth in a xenograft mouse model. Furthermore, ablation in combination with sorafenib significantly inhibited HCC progression compared with sorafenib treatment alone or vehicle control. To identify candidate ULK1 inhibitors, we used a structure-based virtual docking approach to screen 3428 compounds. Among these compounds, XST-14 showed the highest affinity for the ULK1 protein and specifically blocked ULK1 kinase activity. Moreover, the Lys46, Tyr94 and Asp165 amino acid residues of ULK1 were required for its binding to XST-14 according to molecular docking and mutagenesis experiments. Functional assays revealed that XST-14 blocked autophagy and subsequently induced apoptosis and inhibited growth in HCC cells. More importantly, XST-14 acted synergistically with sorafenib to attenuate HCC progression by inhibiting sorafenib-induced autophagy activation both and . In addition, XST-14 was well tolerated and exhibited favorable drug metabolism and pharmacokinetic properties and low toxicity in mice. In summary, our study determined that ULK1 may represent a new therapeutic target for HCC and that targeting ULK1 in combination with sorafenib treatment may serve as a promising interventional strategy for treating HCC. 3MA: 3-methyladenine; ADV: AutoDock Vina; ATP: adenosine triphosphate; EdU: 5-ethynyl-2'-deoxyuridine; ESI: electrospray ionization; HCC: hepatocellular carcinoma; IC: half maximal inhibitory concentration; KD: kinase domain; q.o.d., every other day; SDS-PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis; SPR, surface plasmon resonance.
虽然巨自噬/自噬参与肝细胞癌（HCC）的发生和发展，并被确定为 HCC 治疗耐药的机制，但 ULK1（unc-51 样自噬激活激酶 1）在 HCC 中的作用尚不清楚。在这里，我们报告说，敲低和敲除 均抑制人 HCC 细胞的增殖和侵袭，并且 缺失消除了异种移植小鼠模型中的肿瘤生长。此外，与索拉非尼单药治疗或载体对照相比， 消融联合索拉非尼显着抑制 HCC 进展。为了鉴定候选 ULK1 抑制剂，我们使用基于结构的虚拟对接方法筛选了 3428 种化合物。在这些化合物中，XST-14 与 ULK1 蛋白的亲和力最高，并特异性阻断 ULK1 激酶活性。此外，根据分子对接和突变实验，ULK1 的 Lys46、Tyr94 和 Asp165 氨基酸残基对于其与 XST-14 的结合是必需的。功能测定表明，XST-14 阻断自噬，随后诱导 HCC 细胞凋亡和抑制生长。更重要的是，XST-14 与索拉非尼协同作用，通过抑制索拉非尼诱导的自噬激活来减弱 HCC 的进展。此外，XST-14 在小鼠中具有良好的耐受性，并表现出良好的药物代谢动力学特性和低毒性。总之，我们的研究确定 ULK1 可能代表 HCC 的一个新的治疗靶点，并且靶向 ULK1 联合索拉非尼治疗可能是治疗 HCC 的一种有前途的干预策略。3MA：3-甲基腺嘌呤；ADV：AutoDock Vina；ATP：三磷酸腺苷；EdU：5-乙炔基-2'-脱氧尿苷；ESI：电喷雾电离；HCC：肝细胞癌；IC：半最大抑制浓度；KD：激酶结构域；q.o.d.，隔天一次；SDS-PAGE，十二烷基硫酸钠-聚丙烯酰胺凝胶电泳；SPR，表面等离子体共振。
