Zhu Qiaohua, Yu Xinfa, Zhou Zhi-Wei, Zhou Chengyu, Chen Xiao-Wu, Zhou Shu-Feng
Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, FL. United States.
Department of Oncology and Interventional Radiology, Shunde First People`s Hospital, Southern Medical University, Shunde, Guangdong, China.
Curr Cancer Drug Targets. 2017;17(4):386-401. doi: 10.2174/1568009616666160630182344.
Aurora A kinase represent a feasible target in cancer therapy.
To evaluate the proteomic response of human liver carcinoma cells to alisertib (ALS) and identify the molecular targets of ALS, we examined the effects of ALS 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 study was performed to evaluate the proteomic response to ALS. Cell cycle distribution and apoptosis were assessed using flow cytometry and autophagy was determined using flow cytometry and confocal microscopy.
Our SILAC proteomic study showed that ALS regulated the expression of 914 proteins, with 407 molecules being up-regulated and 507 molecules being down-regulated in HepG2 cells. Ingenuity pathway analysis (IPA) and KEGG pathway analysis identified 146 and 32 signaling pathways were regulated by ALS, respectively, which were associated with cell survival, programmed cell death, and nutrition-energy metabolism. 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 a concentration- and time-dependent manner via the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathway. Autophagy inhibition promoted the pro-apoptotic effect of ALS, indicating a cyto-protective role of ALS-induced autophagy. ALS increased the chemosensitivity of HepG2 cells to cisplatin and doxorubicin.
Taken together, ALS induces autophagy and cell cycle arrest in HepG2 cells via PI3K/Akt/mTOR-mediated pathway. Autophagy inhibition may promote the anticancer effect of ALS and sensitize the chemotherapy in HepG2 cells.
极光激酶A是癌症治疗中一个可行的靶点。
为了评估人肝癌细胞对阿利西替尼（ALS）的蛋白质组学反应并鉴定ALS的分子靶点，我们检测了ALS对HepG2细胞增殖、细胞周期、自噬、凋亡及化疗敏感性的影响。
采用基于细胞培养中氨基酸稳定同位素标记（SILAC）的定量蛋白质组学研究来评估对ALS的蛋白质组学反应。使用流式细胞术评估细胞周期分布和凋亡情况，使用流式细胞术和共聚焦显微镜检测自噬。
我们的SILAC蛋白质组学研究表明，ALS调节了914种蛋白质的表达，其中HepG2细胞中有407个分子上调，507个分子下调。 Ingenuity通路分析（IPA）和KEGG通路分析分别鉴定出146条和32条受ALS调节的信号通路，这些通路与细胞存活、程序性细胞死亡和营养能量代谢相关。随后的验证实验表明，ALS通过调节关键细胞周期调节因子的表达，使HepG2细胞显著停滞于G2/M期并导致非整倍体积累。 ALS通过磷脂酰肌醇3激酶（PI3K）/蛋白激酶B（Akt）/雷帕霉素哺乳动物靶点（mTOR）信号通路以浓度和时间依赖性方式诱导明显的自噬。自噬抑制促进了ALS的促凋亡作用，表明ALS诱导的自噬具有细胞保护作用。 ALS增加了HepG2细胞对顺铂和阿霉素的化疗敏感性。
综上所述，ALS通过PI3K/Akt/mTOR介导的途径诱导HepG2细胞自噬和细胞周期停滞。自噬抑制可能促进ALS的抗癌作用并使HepG2细胞的化疗敏感。
