Tumor invasion and metastasis remain a major cause of mortality in breast cancer patients. It was reported that BP1, a homeobox isoform of DLX4, is overexpressed in 80% of breast cancer patients and in 100% of estrogen receptor negative (ER-) tumors. The prevalence of BP1 positive cells and the intensity of BP1 immunoreactivity increased with the extent of ductal proliferation and tumorigenesis. These findings imply that BP1 may play an important role in ER- breast cancer. We sought to determine the effects and mechanisms of BP1 on cell proliferation and metastasis using ER- Hs578T cells as a model. Cells were transfected with either pcDNA3.2 plasmid containing BP1 gene, or pcDNA3.2 vector, then selected and cloned. Overexpression of BP1 increased cell proliferation rate by 2-5 fold (p<0.005), and enhanced the in vitro invasive activity by 25-65 fold (p<0.001). Microarray experiments were performed to identify differentially expressed genes when BP1 is overexpressed. The gene expression profile of the transfected cell lines were compared, resulting in 71 differentially expressed genes with a fold-change of >=2.0. Of those genes, 49 were up-regulated and 22 were down-regulated. Significant pathways were identified involving cell proliferation and metastasis. These data demonstrated that overexpression of BP1 significantly enhanced cell proliferation and metastatic potential in ER- Hs578T cells. Further analysis with more ER- cell lines and patient samples is warranted to establish BP1 as a therapeutic target for ER- breast cancer.

We found that microRNA-320a (miR-320a) was an attractive prognostic biomarker in breast cancer (BC) previously, whereas its regulatory mechanism in BC was not well understood. Our aim was to identify miR-320a target gene, examine the clinical relationship between miR-320a and its target, and further explore the functions of its target in BC. In this study, miR-320a downstream target gene was determined in HEK-293T cells by dual luciferase reporter assay. Then western blotting and immunohistochemistry were used to assess miR-320a target gene expression in fresh frozen (n=19, breast cancer and matched non-malignant adjacent tissue samples) and formalin-fixed paraffin-embedded (FFPE) (n=130, invasive BC tissues, the same panel detected for miR-320a expression previously) breast tissues, respectively. The results suggested that miR-320a could significantly suppressed Rab14 3'-untranslated region luciferase-reporter activity, and thus Rab14 was first identified as miR-320a target in BC. In 19 matched breast tissues, 12 (63%) breast cancer tissues showed high expression of Rab14 compared with the corresponding normal tissues. Rab14 immunoreactivity was mainly detected in the cytoplasm, 77/130 (59.2%) showed high expression. Furthermore, Rab14 expression was found to be inversely correlated with miR-320a expression in fresh-frozen breast tissues as well as in FFPE invasive breast cancer samples. In addition, Rab14 expression levels were positively related to tumor size (P = 0.034), lymph node metastasis (P < 0.001), distant metastasis (P = 0.001), histological grade (P = 0.035) and clinical tumor lymph-node metastasis stage (P = 0.001). Patients with higher Rab14 expression showed shorter overall survival time. Moreover, silencing of Rab14 could suppress proliferation, migration and invasion in breast cancer cell lines. Collectively, our results indicate that miR-320a could target Rab14 and that they could interact biologically in BC.

Dysregulation of TRIM32 has been implicated in several human cancers, however, its clinical significance and biological function in breast cancer have not been investigated. Using immunohistochemistry, we found that TRIM32 expression is upregulated in breast cancer tissues and that it correlates with advanced stage and poor prognosis. TRIM32 is also overexpressed in 4/7 breast cancer cell lines. CCK8 and colony formation assays showed that TRIM32 depletion inhibited proliferation and colony formation in the T47D cell line, while TRIM32 overexpression promoted MCF-7 cell growth and colony formation. Cell viability and Annexin V/PI staining demonstrated that TRIM32 maintained breast cancer cell survival and reduced apoptosis rate when cells were treated with cisplatin. Western blot analysis demonstrated that TRIM32 overexpression resulted in an upregulation of p-IκB, p-p65, cIAP1, and cIAP2 and a downregulation of p21 and p27 in MCF-7 cells. TRIM32 depletion in T47D cells demonstrated the opposite results, suggesting that TRIM32 may activate the NF-κB pathway. The NF-κB inhibitor BAY 11-7082 blocked the effects of TRIM32 on cisplatin resistance and cIAP1/2 protein regulation. Taken together, the present study demonstrates that TRIM32 downregulates p21/p27 and upregulates IAP family proteins to facilitate breast cancer cell growth and inhibit drug-induced apoptosis, possibly through the NF-κB signaling pathway.

Background: The study of CTLA-4 inhibitors has been one of the hot spots in the field of tumor immunotherapy. As the most immunogenic subtype of breast cancer, Triple negative breast cancer (TNBC) has a great potential in the treatment strategy. The aim of this study was to explore the relevant genes and pathways of CTLA-4 in TNBC and to explore the prognostic value, so as to provide a theoretical basis for clinical studies. Materials and methods: We used the data from The Cancer Genome Atlas (TCGA) to analyze the expression of CTLA-4 in different types of breast cancer, and analyzed the TNBC data of CTLA-4 related co-expression genes by WGCNA and enrichment analysis. LncRNA-miRNA-CTLA-4 network was constructed to explore the immune infiltration and immune checkpoint associated with CTLA-4. The effect of CTLA-4 on clinical outcomes in TNBC patients was also evaluated. Finally, we used data from GEO database to verify the differences of CTLA-4 in different molecular types of breast cancer and related prognostic results. Results: CTLA-4 was significantly higher in TNBC than in Luminal subtype and Her-2 + subtype (P=0.019 and P<0.001, separately), and was significantly higher in ER and PR negative samples than in ER and PR positive samples (P<0.001). CTLA-4 related genes mainly enriched in biological process of leukocyte differentiation, regulation of leukocyte activation and T cell activation. Hsa-mir-92a was found to be a survival significance marker associated with CTLA-4 and lncRNA-miRNA-CTLA-4 network was constructed. The results of immune infiltration analysis showed that CTLA-4 was mainly related with T cell (r=0.74). For immune checkpoints analysis, CTLA-4 was mainly related to PDCD1(r=0.72) and CD28(r=0.64). In TNBC, high expression of CTLA-4 is related to good survival (P=0.0061). Results consistent with previous analysis were obtained in the GEO database, the expression of CTLA-4 in TNBC was significantly higher than that in non-TNBC (p<0.001), CTLA-4 was associated with favorable survival of TNBC (p<0.001). Conclusion: Among all types of breast cancer, the expression of CTLA-4 was the highest in TNBC.CTLA-4 in TNBC can be regulated by hsa-mir-92a to form ceRNA networks and influence the prognosis of TNBC patients through the leukocyte differentiation, regulation of leukocyte activation and T cell activation pathway.

HES1 is a transcriptional repressor involved in cell differentiation and proliferation as well as in various cancer developments, but its expression pattern and biological roles in breast cancer have not been examined. In this study, we assessed HES1 expression in breast cancer tissues using immunohistochemistry and Western blot analyses and investigated HES1 function using MTT and Matrigel invasion assays. Significant relationships were observed between HES1 upregulation and advanced TNM stage (p=0.011), node metastasis (p=0.043), negative oestrogen receptor expression (p=0.001) and triple-negative status (p=0.001). HES1 overexpression was correlated with poor prognosis in breast cancer patients (p<0.05). The MTT and Matrigel invasion assays showed that silencing HES1 in MDA-MB-231 cells decreased cell proliferation and invasion, whereas overexpression of HES1 in MCF-7 cells enhanced its proliferation and invasion. Further analyses showed that silencing HES1 downregulated p-AKT and impeded epithelial-mesenchymal transition (EMT), whereas overexpression of HES1 upregulated AKT phosphorylation and induced EMT. Our study demonstrated that HES1 upregulation is a predictor of poor prognosis in human breast cancers and might be a critical contributor to the proliferation and invasion of breast cancer cells. Moreover, the proportion of cells with overexpression of HES1 in triple-negative breast cancer (TNBC) samples was significantly higher. Thus, HES1 might be a potential therapeutic target in the treatment of TNBC.

Objective: The aim of this study was to conduct a meta-analysis of 49 relevant studies to evaluate the prognostic value of miRNA-200c in various human malignant neoplasms. Methods: All relevant studies were identified by searching PubMed, Embase and Web of Science until August 15st, 2018. Pooled hazard ratios (HRs) with 95% confidence intervals (CIs) of miRNA-200c for overall survival (OS) and progression-free survival (PFS)/recurrence-free survival (RFS)/disease-free survival (DFS) were calculated to investigate such associations. Results: Overall, 49 eligible studies were included in this meta-analysis. Our results showed that high expression of miRNA-200c was significantly correlated with a poor OS in cancer (pooled HR = 1.32, 95% CI: 1.06-1.65), but was not significantly correlated with PFS/RFS/DFS in cancer (pooled HR=1.05, 95% CI: 0.84-1.23). In our subgroup analysis, high miRNA-200c expression predicted a significantly worse OS (pooled HR = 1.50, 95% CI: 1.12-2.01) only in Caucasians. Moreover, high miRNA-200c expression even showed significantly poor OS (pooled HR = 1.88, 95% CI: 1.39-2.54) in blood samples. In addition, a significantly unfavorable OS (pooled HR = 2.69, 95% CI: 1.49-4.85) and (pooled HR = 2.66, 95% CI: 1.07-6.59) associated with up-regulated miRNA-200c expression were observed in breast cancer and endometrial cancer, respectively. Besides, high miRNA-200c expression also showed significantly poor PFS/RFS/DFS (pooled HR=1.66, 95% CI: 1.03-2.67) in breast cancer. Conclusions: Our findings indicated that high miRNA-200c expression was a promising biomarker for patient survival and disease progression in malignant tumors, especially in breast cancer and endometrial cancer. Considering the insufficient evidence, further large-scale researches and clinical studies were needed to verify these results.