1. Department of Interventional Radiology, Nanfang Hospital, Southern Medical University, 1838 North Guangzhou Avenue, Guangzhou, Guangdong 510515, China.
2. Department of Oncology and Interventional Radiology, Shunde Hospital, Southern Medical University, Shunde, Foshan, Guangdong 528300, China
3. Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, FL, USA.
4. Guizhou Provincial Key Laboratory for Regenerative Medicine, Stem Cell and Tissue Engineering Research Center & Sino-US Joint Laboratory for Medical Sciences, Guiyang Medical University, Guiyang 550004, China.
5. Department of Bioengineering and Biotechnology, College of Chemical Engineering, Huaqiao University, Xiamen, Fujian 361021, China.
#These two authors contribute equally to this article.
Hepatocellular carcinoma (HCC) is the sixth most common cancer worldwide, but the overall prognosis remains disappointing especially in the advanced-stage patients. Aberration expression of Aurora kinases is tumorigenic and thus it has attracted interests as therapeutic targets in cancer treatment. Here, we investigated the proteomic response of HCC Hep3B cells to danusertib (Danu), a pan-Aurora kinase inhibitor, and then validated the proteomic results based on stable-isotope labeling by amino acids in cell culture (SILAC). The proteomic data identified that Danu modulated the expression of 542 protein molecules (279 up-regulated; 260 down-regulated; 3 stable). Ingenuity pathway analysis (IPA) and KEGG pathway analysis identified 107 and 24 signaling pathways were regulated by Danu, respectively. IPA analysis showed cellular growth and proliferation, and cell death and survival were among the top five molecular and cellular functions regulated by Danu. The verification experiments showed that Danu inhibited the proliferation of Hep3B cells with a 24-hr IC50 value of 22.03 µM. Danu treatment also arrested Hep3B cells in G2/M phase via regulating the expression of key cell cycle regulators and induced apoptosis via mitochondria-dependent pathway in a dose-dependent manner. Besides, Danu induced a marked autophagy, and inhibition of autophagy enhanced the anticancer effects of Danu, indicating a cyto-protective role of Danu-induced autophagy. Our proteomic data and Western blotting assays showed the PI3K/Akt/mTOR signaling pathway was involved in the inducing effect of Danu on apoptosis and autophagy. Collectively, our findings have demonstrated that the Aurora kinases inhibition with danusertib results in global proteomic response and exerts anticancer effects in Hep3B cells involving regulation of cell cycle, apoptosis and autophagy and associated signaling pathways.
Keywords: Hepatocellular carcinoma, Aurora kinases, danusertib, cell cycle, apoptosis, autophagy, quantitative proteomics.