Molecular Mechanism of Aurora Kinase A Regulating the Meiosis of Oocyte / 中国医学科学院学报

Aurora kinase A (AURKA),a family member of aurora kinases,is involved in mitotic entry,maturation and separation of centrosome,assembly and stabilization of bipolar spindle,and condensation and separation of chromosome.Studies have demonstrated that AURKA plays a similar role in meiosis,while the specific mechanism and the similarities and differences in its role between meiosis and mitosis remain unclear.Therefore,we reviewed the studies about the localization and activation of AURKA in oocyte meiosis,and compared the role of AURKA in regulating spindle formation,activating spindle assembly checkpoint,and correcting the kinetochore-microtubule attachment between the meiosis of oocytes and the mitosis of somatic cells.This review will lay a theoretical foundation for revealing the mechanism of AURKA in the regulation of cell division and for the clinical research related to cancer and reproduction.

Identification of linear ubiquitination substrates with UBAN domain probe3 / 中国药理学与毒理学杂志

OBJECTIVE In this research, the ubiquitin binding in ABIN and NEMO proteins (UBAN) domain of NF-kB essential modulator (NEMO) was used as a protein probe to screen potent linear ubiq-uitination substrates. Adopting the proteomics strategy, the putative substrates could be enriched and identified. This work will greatly expand our knowledge on the molecular mechanism of linear ubiquitina-tion regulation in various biological events. METHODS ® The glutathione S-transferase (GST)-UBAN protein was overexpressed by Escherichia coli and connected with glutathione agarose beads by GST tag to be used as the UBAN probe. Then, the function of the probe was verified in linear ubiquitination chain binding ability test by immunoprecipitation and Western blotting in 293T cells. Using the UBAN probe.linear ubiquitination substrates were enriched and identified by mass spectrometry in 293T cell lysate and analyzed by bioinformatics. Among them, the potential linear ubiquitination substrates Aurora kinase A (Aurora-A) and linear ubiquitin chain assembly complex (LUBAC) were co-transfected in 293T cells and the linear ubiquitination chain binding ability of Aurora-A was identified by Western blot. ® LUBAC was interfered in with siRNA and then synchronized in HeLa cells. The regulation of linear ubiquitination on the biological function of potential substrate Aurora A was observe by Western blotting. RESULTS (1) Western blotting results suggested that the UBAN probe could bind to the linear ubiquitin chain on NEMO, which confirmed the success of the system. ® Including Aurora-As a total of 403 proteins were identified as potential substrates of linear ubiquitination by mass spectrometry. The ability to bind to the linear ubiquitination chain was confirmed by immunoprecipitation between Aurora-A and the linear ubiquitination chains. ® Western blotting results showed that compared with the control group, Aurora-A phosphorylation level was up-regulated, which meant the occurrence of self-activation after knockdown of LUBAC. CONCLUSION The probe constructed with the UBAN domain can efficiently capture and enrich linear ubiquitination substrates. And the self-activation of Aurora-A may be regulated by linear ubiquitination. Significantly, the strategy of identifying the linear ubiquitination substrates established in this paper provides a novel idea and method for further exploration of biological functions of linear ubiquitination.

Mechanism of Aurora-A to promote chemoresistance in nasopharyngeal carcinoma / 中国耳鼻咽喉头颈外科

OBJECTIVE To explore the mechanism of Aurora kinase A (Aurora-A) promoting cancer cell chemotherapy resistance in nasopharyngeal carcinoma. METHODS The expression of Aurora-A in nasopharyngeal carcinoma tissues and adjacent tissues were detected by Western bolt and Q-PCR. The highexpressing Aurora A cell line CNE2 was used to detected the cell apoptosis and the expression of key pathway marker protein after Aurora-A inhibitor VX680 and cisplatin treatment by using Flow cytometry and WB. RESULTS The expression of Aurora-A in nasopharyngeal carcinoma tissues was significantly higher than that in adjacent tissues. Comparing to normal nasopharyngeal cells NP69, Aurora-A was significantly highly expressed in all of nasopharyngeal carcinoma cells and was highest in CNE2. Inhibiton of Aurora-A increased the cell apoptosis and the expression of p-AKT, p21 and Cleaved-Caspase-3 after using cisplatin or the Aurora-A inhibitor VX680 treatment. CONCLUSION The results shown that Aurora-A confer chemoresistance to cisplatin treatment through p-AKT/p21/Cleaved-Caspase-3 pathway.

Down-regulation of aurora kinase A activity inhibits pro-angiogenesis, migration and invasion of Hep2 cells / 上海交通大学学报（医学版）

Objective: To investigate the effect of aurora kinase A (AURKA) activity on pro-angiogenesis, migration and invasion of Hep2 cells. Methods: Down-regulation of AURKA phospholation in Hep2 cells with 100 nmol/L VX680. Western blotting was used to detect the expression of p-AURKA, vascular endothelial growth factor A (VEGFA) and its receptor VEGFR2 in blank control Hep2 cells and Hep2 cells treated with VX680 for 24 h and 48 h. Angiogenesis experiment was applied to observe the effect of Hep2 cells treated with VX680 on angiogenesis. CCK8 cell proliferation assay, Transwell migration and invasion experiment were used to observe the ability of cell proliferation, migration and invasion of Hep2 cells. Results: Compared with control cells, the expression of p-AURKA in Hep2 cells treated with VX680 for 48 h was decreased (P=0.000). After downregulation of p-AURKA, the expressions of VEGFA and VEGFR2 were decreased (P=0.000). Meanwhile, angiogenesis was inhibited (P=0.000), and migration and invasion of Hep2 cells were reduced (P=0.000). Conclusion: AURKA regulates pro-angiogenesis, migration and invasion of Hep2 cells, which is a new strategy for the treatment of laryngeal cancer by targeting AURKA.

Clinical significance of targeting drug-based molecular biomarkers expression in ovarian clear cell carcinoma / 中华妇产科杂志

Objective To assess the expression level of targeting drug-based molecular biomarkers in ovarian clear cell carcinoma(OCCC)tissues and its clinical significance.Methods A total of 63 OCCC patients included 40 primary OCCC and 23 recurrent OCCC for secondary cytoreductive surgery(SCS),who had received primary surgeries at Fudan University Shanghai Cancer Center between January, 2008 and December, 2015 were enrolled, and immunohistochemistry SP method was used to test human epidermal growth factor receptor (EGFR), human epidermal growth factor receptor-2 (HER2), aurora kinase A (AURKA), breast cancer susceptibility gene 1 (BRCA1), BRCA2 and programmed death-ligand 1 (PD-L1) protein expression in paraffin-embedded tissues. Results The positive rates of EGFR, HER2, AURKA, BRCA1,BRCA2 and PD-L1 in primary and recurrent tumor tissues were respectively 20%(8/40)vs 30%(7/23),22%(9/40)vs 35%(8/23),38%(15/40)vs 35%(8/23),42%(17/40)vs 39%(9/23),20%(8/40)vs 22%(5/23), 25%(10/40)vs 17%(4/23), and there were no significant differences between primary and recurrent OCCC (all P>0.05). χ2-test or Fisher exact analysis revealed that HER2 expression in recurrent tumor tissues had a relationship with chemoresistance (P<0.05), while the expression of other biomarkers showed no significant relationship with chemoresistance (all P>0.05). Further, Kaplan-Meier survival analysis showed that patients with HER2 and AURKA-positive expression had a significantly shorter progression-free survival time in primary OCCC(4 months vs 10 months,log-rank test,P<0.05 for HER2;and 4 months vs 10 months,P<0.05 for AURKA);and a shorter overall survival time after SCS in recurrent OCCC (10 months vs 44 months, P<0.05 for HER2;and 13 months vs 43 months, P<0.05 for AURKA). However,multivariate Cox proportional hazards regression analysis indicated that none of these 6 biomarkers was independent risk factor of progression-free survival time of primary OCCC or overall survival time after SCS for recurrent OCCC (P>0.05). Conclusion HER2 and AURKA could serve as prognostic factors in ovarian clear cell carcinoma.

Characterization of the Indirubin Derivative LDD970 as a Small Molecule Aurora Kinase A Inhibitor in Human Colorectal Cancer Cells

Aurora kinase A plays an essential role in mitosis including chromosome separation and cytokinesis. Aberrant expression and activity of Aurora kinase A is associated with numerous malignancies including colorectal cancer followed by poor prognosis. The aim of this study is to determine the inhibitory effects of LDD970, an indirubin derivative, on Aurora kinase A in HT29 colorectal cancer cells. In vitro kinase assay revealed that, LDD970 inhibited levels of activated Aurora kinase A (IC₅₀=0.37 mM). The inhibitory effects of LDD970 on Aurora kinase A, autophosphorylation and phosphorylation of histone H3 (Ser10), were confirmed by immunoblot analysis. Moreover, LDD970 inhibited migration of HT29 cells and upregulated apoptosis-related protein cleaved PARP. In cell viability assay, LDD970 was observed to suppress HT29 cell growth (GI₅₀=4.22 µM). Although further studies are required, results of the present study suggest that LDD970 provide a valuable insight into small molecule indirubin derivative for therapeutic potential in human colorectal cancer.

Aurora Kinase A Is a Prognostic Marker in Colorectal Adenocarcinoma

BACKGROUND: Aurora kinase A (AURKA), or STK15/BTAK, is a member of the serine/threonine kinase family and plays important roles in mitosis and chromosome stability. This study investigated the clinical significance of AURKA expression in colorectal cancer patients in Korea. METHODS: AURKA protein expression was evaluated by immunohistochemistry in 151 patients with colorectal adenocarcinoma using tissue microarray blocks. We analyzed the relationship between clinicopathological characteristics and AURKA expression. In addition, the prognostic significance of various clinicopathological data for progression-free survival (PFS) was assessed. Also we evaluated copy number variations by array comparative genomic hybridization and AURKA gene amplification using fluorescence in situ hybridization in colorectal carcinoma tissues. RESULTS: AURKA gene amplification was found more frequently in the 20q13.2–13.33 gain-positive group than the group with no significant gain on the AURKA-containing locus. AURKA protein expression was detected in 45% of the cases (68/151). Positive staining for AURKA was observed more often in male patients (p = .035) and distally located tumors (p = .021). PFS was shorter in patients with AURKA expression compared to those with low-level AURKA expression (p < .001). Univariate analysis revealed that AURKA expression (p = .001), age (p = .034), lymphatic invasion (p = .001), perineural invasion (p = .002), and TNM stage (p = .013) significantly affected PFS. In a multivariate analysis of PFS, a Cox proportional hazard model confirmed that AURKA expression was an independent and significant prognostic factor in colorectal adenocarcinoma (hazard ratio, 3.944; p < .001). CONCLUSIONS: AURKA could serve as an independent factor to predict a poor prognosis in Korean colorectal adenocarcinoma patients.

Study on Improvement of Chemosensitivity of MCF-7 Cells to Epirubicin and Inhibition of Aurora Kinase A in Treatment of Breast Cancer by San-Huang Decoction / 世界科学技术-中医药现代化

This article was aimed to explore the effect ofSan-Huang (SH) decoction on improving chemosensitivity of MCF-7 cells to epirubicin and inhibition of Aurora kinase A, in order to discuss its underlying mechanism. The inhibition of MCF-7 cells proliferation on breast cancer by SH decoction was determined by CCK-8 assay. RT-PCR and western blot were used to detect the Aurora A, p53 mRNA and protein expression level of MCF-7 cells by SH decoction. The siRNA silenced Aurora A of MCF-7 cells. CCK-8 assay was used to detect the inhibition of MCF-7 cells proliferation. CCK-8 assay and AnnexinV-FITC/PI staining were used to detect the inhibition rate and apoptosis rate of MCF-7 cells treated by the combination of SH decoction and epirubicin. Western blot analysis was used to detect the expression of apoptosis-related proteins. The results showed that SH decoction inhibited the proliferation of MCF-7 cells in a dose-dependent manner (P 0.05). SH decoction can regulate the Aurora A, p53 protein and mRNA expression of MCF-7 cells. siRNA silenced Aurora A, which downregulated the inhibition rate of MCF-7 cells by SH decoction for 50.0% (from 49.2% to 24.8%). The combination of SH decoction and epirubicin enhanced the effect of epirubicin on inhibiting the proliferation rate and apoptosis rate of MCF-7 cell, regulated the expression levels of apoptosis-related protein such as c-PARP, c-Caspase 3, Bcl-2, Bax, as well as the protein level of Aurora A. It was concluded that SH decoction can increase the chemosensitivity of MCF-7 cells to epirubicin, which may be related to the inhibition of Aurora Kinase A by SH decoction.