A vector-based short hairpin RNA targeting Aurora A inhibits breast cancer growth

Abstract

Abstract. Aurora A plays an essential role in centrosome maturation, separation and in the formation of the mitotic bipolar spindle. Overexpression or amplication of Aurora A gene has been detected in many cancer cell lines and various tumor tissues, including breast cancer, suggesting that Aurora A might be drug target for breast cancer treatment. In the current study, short hairpin RNA targeting Aurora A was cloned into pGenesil-2 plasmid vector and then transfected into MDA-MB-435S and ZR-75-30 human breast cancer cells using cationic liposome. Reduced expression of Aurora A was detected by RT-PCR and Western blot. The effect of pGenesil-2-shAURKA plasmid on tumor growth in MDA-MB-435S xenogenic implantation model was studied. pGenesil-2-shAURKA plasmid inhibited tumor growth significantly by systemantic administration. To further study the underlying mechanisms, cell apoptosis and proliferation were investigated by flow cytometric analysis, propidium iodide staining, TUNEL and Ki-67 immunostaining respectively. Increased apoptosis and reduced cell proliferation were detected in vitro and in vivo studies. In summary, our results suggested that specific knockdown of Aurora A expression by vector based shRNA may be a potential therapy for human breast cancer. IntroductionBreast cancer is the most commonly diagnosed cancer in women worldwide. More than 1.15 million women were diagnosed with breast cancer in 2002 (1). Registry data showed that breast cancer incidence has been increasing since 1973, especially in low and middle-income countries.In China, urban registries showed 20-30% increase in the past decade (2). Current breast cancer incidence and mortality rates (~180,510 and 40,940 cases in 2007 in the United States, respectively) (3) highlight the need to explore alternative therapeutic strategies.The process of cell division is instrumental to the development and progression of tumors, and targeting cell division has been proved as a successful antitumor therapy. A number of trials has addressed the benefit of adding a taxane (paclitaxel or docetaxel) to an anthracycline-based adjuvant chemotherapy regimen in breast cancer (4,5), suggesting that the molecules involved in cell cycle and division may be targets for cancer treatment.Aurora A is an oncogenic serine/threonine kinase that plays an essential role in centrosome maturation, separation and in the formation of the mitotic bipolar spindle in various organisms (6-10). It has been reported that its ectopic expression in immortalized NIH/3T3 cells is sufficient to provoke their transformation, defining Aurora A as an oncogene (11). Overexpression or amplication of Aurora A gene has been detected in many cancer cell lines and various tumor tissues, including breast cancer (reviewed in ref. 12). Nadler et al reported that Aurora A expression defines a population of patients with decreased survival, whereas Aurora B expression does not, suggesting that Aurora A might be the preferred drug target in breast cancer (13).RNA interference is a specific way of g...