Targeting Aurora A kinase with Alisertib (ALS) Induces Autophagy and Cell Cycle Arrest and Increases Chemosensitivity in Hepatoblastoma HepG2 Cells.

Abstract To evaluate the proteomic response to alisertib (ALS) and identify the molecular targets of ALS in the treatment of hepatoblastoma, we examined the effect of ALS with a particular focus on the proliferation, cell cycle, autophagy, apoptosis, and chemosensitivity in HepG2 cells. The stable-isotope labeling by amino acids in cell culture (SILAC) based quantitative proteomic approach was performed to evaluate the proteomic response to ALS. Cell viability was determined by MTT assay. Cell cycle distribution and apoptosis were assessed using flow cytometry. Autophagy was examined using flow cytometry and confocal microscopy. Protein expression was detected by western blotting assay. Our SILAC proteomics showed that ALS modulated the expression of 914 proteins, with 407 molecules being up-regulated and 507 molecules being down-regulated. Ingenuity pathway analysis (IPA) and KEGG pathway analysis identified 146 and 32 signaling pathways were regulated by ALS, respectively, with a characteristic pathway of active nutrition-energy metabolism being notable. Subsequently, the verification experiments showed that ALS remarkably arrested HepG2 cells in G2/M phase and led to an accumulation of aneuploidy via regulating the expression of key cell cycle regulators. ALS induced a marked autophagy in concentration- and time-dependent manners via PI3K/Akt/mTOR signaling pathway. Autophagy inhibition promoted the pro-apoptotic effect of ALS, indicating a cyto-protective role of ALS-induced autophagy. ALS increased the chemo-sensitivity of HepG2 cells to cisplatin and doxorubicin. ALS exerted anticancer effects in both monotherapy and combinational therapy in HepG2 cells. Autophagy inhibition may promote the anticancer effect of ALS and sensitize the chemotherapy in HepG2 cells.