Study On The Mechanisms Of LZTS1 Responsible For Invasion And Metasitasis In Breast Cancer

ObjectiveThe aim of this study was to investigate the correlation of LZTS1 protein expression and clinicopathologic features and patients long-term survival in invasive carcinoma of the breast, identify the role of LZTS1 gene in the progression of breast cancer. In vitro study was to assess whether LZTS1 plays a role in metastasis of breast cancer and to explore the molecular mechanisms.MethodsWe analyzed LZTS1 protein expression in 340 cases of invasive breast cancer by immunohistochemistry using tissue microarray, and compare the association between the expression levels of LZTS 1 and clinicopathological parameters and 5-year progression-free survival rate. To investigated the molecular mechanism of LZTS 1 protein, we over-expression LZTS1 in MDA-MB-231 cell line using gene transfection technology, then the cells proliferation status in vitro of the LZTS1 re-expression cells was analyzed by SRB and cloning formation assay, the migration and invasion capacity was observed by using wound healing assay and transwell assay. To investigate the underlying molecular mechanism of LZTS 1 to inhibit the invasion of tumor cells we constructed a monoclonal MDA-MB-231 cell line stable-expressing LZTS1 and a control clone. Moreover, we observed the morphological changes, and detected the expression changes of E- cadherin, vimentin, TGF-β1, slug, sail, cytokeratinby using real-time RT-PCR, Western blotting, immunofluorescence staining Based on the results above, we examine and compare the association between LZTS1 protein and E-cadherin, B-catenin expressions by IHC to explore the mechanism of LZTS 1 suppress tumor metastasis in vivo.Results1. In the 340 invasive breast carcinoma samples examined,297(87%) showed absence or down-regulation of LZTS1 protein, absence or down-regulation of LZTS1 protein in invasive breast carcinoma was significantly correlated with tumor histological grade (P=0.000<0.01), lymph node metastasis (P=0.000<0.01), positively correlated with PR expression (P=0.011), no significant associated with HER2/neu expression and tumor size.2. Kaplan-Meier survival analyses of these carcinoma specimens revealed a correlation between absent or deduced LZTS1 expression levels and shorter progression-free survival times (P<0.001). Multivariate analysis confirmed that LZTS1 was an independent prognostic indicator of invasive breast cancer.3. In vitro, both colony-formation assay and SRB assay showed that overexpression LZTS1 can inhibited MDA-MB-231 breast cancer cell colony formation and proliferation (P<0.05), inhibited MDA-MB-231 cell migration and invasion (P= 0.005, P=0.03).4. MDA-MB-231 cells were morphologically epithelial-like changes in LZTS1 stable-expressing cells compared with the control plasmid transfected cells, and cytokeratin, E-cadherin expression increased; vimentin and transcription factor TGF-β1, slug, sail were significantly downregulated. And LZTS1 expression was positively correlated with E-cadherin,β-catenin expression in invasive breast carcinoma samples.ConclusionLZTS1 is a potential breast cancer metastasis suppressor gene. Decrease/loss of LZTS1 protein could promote breast cancer cell invasion, metastasis of breast cancer and increases the risk of recurrence, metastasis and mortality; LZTS1 protein restored can inhibit tumor cell migration and invasion, and which may be functioned with preventing the epithelial-mesenchymal transition. Therefore, our findings establish LZTS 1 as a novel biomarker predicting progression and a potential target for future therapies for breast cancer.