The Role Of MTA1 In Stress Induced Breast Cancer Metastasis Associated With Chemotherapy

IntroductionBreast cancer is the most prevalent malignant tumor for female. With the progress in cancer research and clinical care, the prognosis of the breast cancer patients has improved a great deal. Most of the patients can obtain obvious improvement after treatment and even in petients with stage â…¢breast cancers the 5-year survival rate can reach 70%. However, the long-term survival of patients is still hard to achieve and the leading factor that results in the deaths of breast cancer patients is metastasis.Therefore, how to inhibit the metastasis of cancer cells has become one of the most pressing problems in breast cancer research.As a conventional means of treatment of breast cancer, chemotherapy has been widely used in clinical application. Chemotherapy agents kill cancer cells through cytotoxic effects and are usually used as an adjuvant therapy for breast cancer patients before or after surgery or as a conservative therapy for those whose tumors are inoperable. But recent studies have shown that chemotherapy has a potential effect to promote tumor metastasis. We hypothesise that hostile environmental factors can cause stress response of tumor, thereby cancer cells tend to move away from the adverse stimulation. When under attack, the natural response of any organism is to defend itself or escape; cancer cells tend to escape. Accordingly, chemotherapy agents as a powerful stress signal should play a role in stimulating tumor metastasis.Metastasis associated protein 1(MTA1) is an important molecule closely associated with metastasis of breast cancer. MTA1 overexpression can be found in a variety of human malignant tumors including breast cancer, and always indicting more advanced cancer stages, increased metastasis tendency, and unfavorable outcomes. More importantly, MTA1 is a stress response protein whose expression is significantly affected by environmental factors. Stress agents such as hypoxia, ironic and ultraviolet radiation and inflammation can induce the expression of MTA1. The effects of chemotherapy on the expression level of MTA1 in breast cancer cells have not been reported yet. So we hope to observe the effects of chemotherapy on metastatic potential of breast cancer, analysis the change of MTA1 expression level in this process and find the relationship between MTA1 and the changes of biology behavior in breast cancer cells after chemotherapy. ObjectivesThis study aims at analysing the effect of chemotherapy on metastatic potential of breast cancer by observing the changes in migration and invasion ability of breast cancer cells due to the administration of epirubicin. By comparing MTA1 expression level in epirubicin treated breast cancer cells with that in untreated cells, and by observing the changes of epirubicin treatment effect on metastatic potential after inhibition of MTA1, we also hope to analysis the role MTA1 plays in the process of biological behavior changes in breast cancer cells induced by chemotherapy. Methods1. MCF-7 and MDA-MB-231 human breast cancer cells were treated with epirubicin at half maximal inhibitory concentration for 6h or 12 h. The capacity of migration and invasion were test by wound-healing essay and transwell essay respectively.2. Total RNA and protein were extracted from epirubicin treated cells and untreated cells. Real-time PCR and western blot were used to detect the change in MTA1 expression induced by epirubicin treatment.3. MCF-7 and MDA-MB-231 cells were transfected with either NC si RNA or MTA1 specific si RNA. MTA1 expression levels in the transfected cells were compared with those in the normal cells by real-time PCR and western blot.4. MCF-7 and MDA-MB-231 cells were respectively divided into four groups. Group Control was normal-cultured control group. Cells in Epi group were treated with epirubicin for 6h. Cells in Epi + NC si RNA group were first transfected with NC si RNA and then treated with epirubicin. Cells in Epi + MTA1 si RNA group were transfected with MTA1 specific si RNA and then treated with epirubicin. Wound-healing essay and transwell essay were used to examine the migration and invasion capacity of cancer cells.5. Mouse 4T1 breast tumor model was established. Animals were divided into control group and epirubicin-giving group. Chemotherapy was administered at the dose of 3mg/kg once a week and lasted for 3 weeks. Then mice were executed and the numbers of lung surface metastases in mice from different groups were observed and compared. Results1. Cells of both two cell lines treated with epirubicin for 6h showed increased wound closure than untreated cells in wound-healing essays. In the transwell assays, MDA-MB-231 cells treated with epirubicin for 6h showed significant increase in invading cells compared with the untreated ones. But these effects were not observed in the cells treated for 12 h. There was no significant difference between treated and untreated MCF-7 cells in transwell essays because they can hardly invade the Matrigel-coated membrane.2. MTA1 m RNA and protein levels were significantly increased in epirubicin treated MCF-7 and MDA-MB-231 cells compared with the untreated cells.3. MTA1 m RNA and protein levels in cells transfected with MTA1 si RNA showed obvious decline compared with the normal cells and cells transfected with NC si RNA in both two cell lines.4. In wound-healing essays, both two kinds of cells in Epi group and Epi + NC si RNA group showed greater wound closure than cells in Control group and the average wound closure in Epi + MTA1 si RNA group was significantly reduced compared with Epi group. In transwell essays,more MDA-MB-231 cells in Epi group and Epi + NC si RNA group invaded the membrane than Control group, and the invading cells in Epi + MTA1 si RNA group were significantly less than Epi group.5. In the in vivo chemotherapy expression, we observed an obvious increase of the number of surface lung metastases in the epirubicin-giving mice compared with the untreated controls. Conclusions1. Epirubicin treatment can increase migration and invasion capacity of breast cancer cells in vitro and this process can be affected by time of administration.2. Epirubicin treatment can significantly improve the expression of MTA1 in breast cancer cells and MTA1 specific si RNA can effectively inhibit the expression of MTA1.3. The knockdown of MTA1 can neutralize the enhanced metastatic potential induced by epirubicin treatment in breast cancer cells to some extent.4. Low-dose epirubicin chemotherapy in 4T1 tumor bearing mice can promote lung metastasis of 4T1 breast cancer.