Investigation of the effect of mir-96 expression on autophagy in breast cancer cell lines

The difference of microRNAs (miRNA) expression level in cancer cells has been an important subject of cancer researchers. As in all cancers, miRNAs promoting or suppressing tumorigeneses in breast cancer were found and additional miRNAs are still being included. miRNAs are thought to be able to use in the subjects of breast cancer diagnosis, treatment, prognosis and also response of cancer to drug. The elucidation of the role of autophagy pathway in breast cancer is greatly important for cancer treatment and drug interactions. Overexpression of miR-96 was associated with poor prognosis in breast cancer. The drug, Trastuzumab, is often used in HER2 positive breast cancer but this drug is not effective enough. Moreover, no drug which is effective in especially triple negative breast cancer has been found yet. The drug, Everolimus, suppresses mTOR pathway, thus stimulates the autophagy pathway, and phase trials have been conducted in breast cancer. In this study, the expression levels of miR-96 and its theoretically targets associated with autophagy, ATG16L1, ATG9A, ATG7, CTSB, TP73 genes were investigated in breast cell lines. In addition, the expression levels were measured in the absence drug treatment and after treatment of trastuzumab and everolimus combination. qRT-PCR for miR-96 and qRT-PCR in fluidigm device for ATG16L1, ATG9A, ATG7, CTSB, TP73 genes were used to measure the expression levels. According to the gene expression analysis, a significant increase in miR-96 expression were detected in MCF-7 and SKBR-3 breast cancer cell lines, compared to normal epithelial cell line. Among genes theoretically targeted by miR-96, significant decrease in ATG16L1, ATG9A, ATG7 genes' expressions in SKBR3, significant decrease in CTSB gene expression in both cell lines and significant increase in TP73 gene expression in only MCF-7 were detected, compared to normal epithelial cell line. When everolimus was administered, miR-96 expression was found significantly increased in all cell lines; when everolimus and trastuzumab were administered together, a significant increase in miR-96 expression was measured in only MCF-7. Key Words: Mir-96, autophagy, breast cancer cell lines, trastuzumab, everolimus.