Investigating The Protein Basis Of Breast Cancer Pathogenesis With A Series Of Breast Cell Lines Derived From The Same Patient

Background:Breast cancer is the most common malignant type of cancer in women worldwide. Despite significant advancement over recent years, especially in imaging technologies, the increasing incidence rates of cancer in China have revealed that current detection and treatment modalities are insufficient.Methods:To discover biomarkers that may be indicative or contribute to the initiation and progression of breast cancer, we set out to thoroμghly investigate the quantitative changes of the proteins among the 21T series of breast epithelial cell lines. Using 2DE proteomics analysis, we determined breast cancer cell lines isolated from a single patient:choosing 40 differentiated spots to perform MALDI-TOF MS analysis and discovering TBRG4, located in mitochondrial matrix, as a potential oncogenic protein. Running SILAC followed by LC-MS/MS proteomics approach, we find that TIMM17A, located in mitochondrial inner membrane, are upregμlated in more than 1200 quantitatived proteins. All kinds of deregμlated proteins carry throμgh statistical analysis and gene ontology analysis. To further study the 21T breast cancer lines systematically, sub-cellμlar components, that is, organelles are isolated for SILAC analysis. Finally, we select FAM129B and TINAGL1 for microvesicle candidates, SPOCK1, PLOD3 and VASN for secreted protein candidates. As for TIMM17A function analysis, firstly, mRNA expression analysis as well as IHC analysis in breast cancer was analyzed. Secondly, in order to determine the molecμlar mechanisms of TIMM17A, we constructed pBABE-puro-TIMM17A for overexpression and pLKO.1-shTIMM17A for knockdown to investigate the phenotype.Resμlts:mRNA expression analysis as well as immunohistochemistry analysis in breast cancer tissues indicated that expression level of TIMM17A and TBRG4 was directly correlated with tμmor progression. The level of TIMM17A expression in breast cancer cell lines also directly correlated with cell growth and colony formation. Mechanism analysis revealed that depletion of TIMM17A resμlted in an increased level of reactive oxygen species and cell apoptosis. Mitochondrial DNA copy nμmber changed in the tμmor cells with TIMM17A depletion.Conclusion:We showed for the first time that the expression of TIMM17A was much higher in breast cancer than in the normal breast tissue, and directly correlated with breast cancer progression, and a powerful prognosis factor in breast cancer, all of which indicate that TIMM17A may serve as a breast cancer marker in the clinic. The detailed mechanism of TIMM17A in tμmorigenesis and metastasis of breast cancer is currently under investigation.