non-TNBC Tissues Research Articles

Neuropilin1, a novel independent prognostic factor and therapeutic target in triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is the most aggressive breast cancer, and thus, has limited treatment options. Neuropilin1 (NRP1) is a multi-functional transmembrane protein that interacts with a number of signaling receptors and plays an important role in cancer progression. Previous studies demonstrated that the expression of NRP1 is activated and promotes the progression of breast cancer particularly in TNBC compared to other molecular subtypes; however, whether or not the level of NRP1 expression is related to the progression of TNBC warrants further study. In the current study, we determined the expression and function of NRP1 and evaluated the clinical significance of NRP1 in patients with TNBC. In addition, we determined whether or not an NRP1 antagonist potentiates the inhibitory effects of paclitaxel (PTX) in patients with TNBC. In our clinical study, NRP1 had higher expression in TNBC tissues than non-TNBC tissues at the same stage, and NRP1 was an independent prognostic factor. Specifically, the high expression of NRP1 was associated with shorter survival in TNBC patients. In addition, TNBC cells treated with NRP1 antagonist significantly potentiated the effect of PTX on cell proliferation, cell migration, and apoptosis. Our findings suggest that NRP1 expression can act as an independent prognostic factor for TNBC patients, and the combination of PTX and an NRP1 antagonist may be an effective treatment regimen for TNBC.

Macrophages confer resistance to BET inhibition in triple-negative breast cancer by upregulating IKBKE

BET inhibitors (BETi) exhibit a strong anti-tumor activity in triple-negative breast cancer (TNBC). However, BETi resistance has been reported in TNBC. The mechanisms of resistance have not been demonstrated. Tumor-associated macrophages (TAMs) are frequently involved in cancer cells resistance to chemotherapy, also associated with poor prognosis in TNBC. However, the role of TAMs in BETi resistance remains unknown. Here, we found that BETi JQ1 and I-BET151 exerted anti-tumor effects in TNBC by decreasing IKBKE expression to attenuate NF-κB signaling. TAMs have been reported to associate with chemoresistance in breast cancer. Here, we firstly found that TNBC-stimulated TAMs activated NF-κB signaling by upregulating IKBKE expression to enhance breast cancer cells resistance to BETi. The IKBKE levels were also proved to be higher in clinical TNBC tissues than Non-TNBC tissues, suggesting feedback induction of IKBKE expression by TNBC-stimulated TAMs in TNBC. Moreover, the induction of IKBKE by TAMs in TNBC cells was identified to be associated with STAT3 signaling, which was activated by TAM-secreted IL-6 and IL-10. Lastly, the combination of inhibitors of BET and STAT3 exerted a synergistic inhibition effects in TAM-cocultured or TAM CM-treated TNBC cells in vitro and in vivo. Altogether, our findings illustrated TNBC-activated macrophages conferred TNBC cells resistance to BETi via IL-6 or IL-10/STAT3/IKBKE/NF-κB axis. Blockade of IKBKE or double inhibition of BET and STAT3 might be a novel strategy for treatment of TNBC.

Roles of eIF3m in the tumorigenesis of triple negative breast cancer

BackgroundWithout targets, triple negative breast cancer (TNBC) has the worst prognosis in all subtypes of breast cancer (BC). Recently, eukaryotic translation initiation factor 3 m (eIF3m) has been declared to be involved in the malignant progression of various neoplasms. The aim of this study is to explore biological functions of eIF3m in TNBC.MethodsMultiple databases, including Oncomine, KM-plotter and so on, were performed to analyze prognosis and function of eIF3m in TNBC. After transfection of eIF3m-shRNA lentivirus, CCK-8, colony formation assay, cell cycle analysis, wound healing assay, transwell assays, mitochondrial membrane potential assay and cell apoptosis analysis were performed to explore the roles of eIF3m in TNBC cell bio-behaviors. In addition, western blotting was conducted to analyze the potential molecular mechanisms of eIF3m.ResultsIn multiple databases, up-regulated eIF3m had lower overall survival, relapse-free survival and post progression survival in BC. EIF3m expression in TNBC was obviously higher than in non-TNBC or normal breast tissues. Its expression in TNBC was positively related to differentiation, lymph node invasion and distant metastasis. After knockdown of eIF3m, cell proliferation, migration, invasion and levels of mitochondrial membrane potential of MDA-MB-231 and MDA-MB-436 were all significantly suppressed, while apoptosis rates of them were obviously increased. In addition, eIF3m could regulate cell-cycle, epithelial–mesenchymal transition and apoptosis-related proteins. Combined with public databases and RT-qPCR, 14 genes were identified to be modulated by eIF3m in the development of TNBC.ConclusionseIF3m is an unfavorable indicator of TNBC, and plays a vital role in the process of TNBC tumorigenesis.

Neural Functions Play Different Roles in Triple Negative Breast Cancer (TNBC) and non-TNBC

Triple negative breast cancer (TNBC) represents the most malignant subtype of breast cancer, and yet our understanding about its unique biology remains elusive. We have conducted a comparative computational analysis of transcriptomic data of TNBC and non-TNBC (NTNBC) tissue samples from the TCGA database, focused on genes involved in neural functions. Our main discoveries are: (1) while both subtypes involve neural functions, TNBC has substantially more up-regulated neural genes than NTNBC, suggesting that TNBC is more complex than NTNBC; (2) non-neural functions related to cell-microenvironment interactions and intracellular damage processing are key inducers of the neural genes in both TNBC and NTNBC, but the inducer-responder relationships are different in the two cancer subtypes; (3) key neural functions such as neural crest formation are predicted to enhance adaptive immunity in TNBC while glia development, along with a few other neural functions, induce both innate and adaptive immunity in NTNBC. These results reveal key differences in the biology between the two cancer subtypes, particularly in terms of the roles that neural functions play. Our findings may open new doors for further investigation of the distinct biology of TNBC vs. NTNBC.

HIF-2α regulates CD44 to promote cancer stem cell activation in triple-negative breast cancer via PI3K/AKT/mTOR signaling.

Breast cancer is a common malignant tumor that seriously threatens women's health. Breast cancer stem cell (CSC)-like cell population may be the main factor for breast cancer metastasis. Therefore, targeted therapy for CSCs has great potential significance. Hypoxia-inducible factor is a transcription factor widely expressed in tumors. Studies have shown that down-regulation of the hypoxia signaling pathway inhibits tumor stem cell self-renewal and increases the sensitivity of stem cells to radiotherapy and chemotherapy mediated by hypoxia-inducible factor-2α (HIF-2α). However, the specific mechanism remains unclear and further research is necessary. To investigate the effect of HIF-2α down-regulation on stem cell markers, microsphere formation, and apoptosis in breast cancer cell line MDA-MB-231 under hypoxia and its possible mechanism. Immunohistochemistry was used to detect the expression of HIF-2α and CD44 in triple-negative breast cancer (TNBC) and non-TNBC tissues. Double-labeling immunofluorescence was applied to detect the co-expression of HIF-2α and CD44 in MDA-MB-231 cells and MCF-7 cells. HIF-2α was silenced by RNA interference, and the expression of CD44 and transfection efficiency were detected by real-time fluorescent quantitative PCR. Further, flow cytometry, TdT-mediated X-dUTP nick end labeling, and mammosphere formation assays were used to evaluate the effect of HIF-2α on CSCs and apoptosis. The possible mechanisms were analyzed by Western blot. The results of immunohistochemistry showed that HIF-2α was highly expressed in both TNBC and non-TNBC, while the expression of CD44 in different molecular types of breast cancer cells was different. In in vitro experiments, it was found that HIF-2α and CD44 were expressed almost in the same cell. Compared with hypoxia + negative-sequence control, HIF-2α small interfering ribonucleic acid transfection can lower the expression of HIF-2α and CD44 mRNA(P < 0.05), increase the percentage of apoptotic cells (P < 0.05), and resulted in a reduction of CD44+/CD24- population (P < 0.05) and mammosphere formation (P < 0.05) in hypoxic MDA-MB-231 cells. Western blot analysis revealed that phosphorylated protein-serine-threonine kinase (p-AKT) and phosphorylated mammalian target of rapamycin (p-mTOR) levels in MDA-MB-231 decreased significantly after HIF-2α silencing (P < 0.05). Down-regulation of HIF-2α expression can inhibit the stemness of human breast cancer MDA-MB-231 cells and promote apoptosis, and its mechanism may be related to the CD44/phosphoinosmde-3-kinase/AKT/mTOR signaling pathway.

Identification of FOXE3 transcription factor as a potent oncogenic factor in triple-negative breast cancer

Triple-negative breast cancer (TNBC) represents a unique subgroup of breast cancers (BCa) with potential to be highly proliferative and invasive. Patient with TNBC are prone to developing resistance to chemotherapy. Therefore, TNBC usually has a poor clinical outcome. The key factors driving these malignant features remain poorly understood. In this study, we report for the first time that expression levels of FOXE3, a recently identified lens-specific transcription factor, were preferentially upregulated in TNBC tissues compared to non-TNBC tissues, and this upregulation correlated well to a poor overall/recurrence-free survival in patients. Depletion of FOXE3 in TNBC cell lines promoted cell death, cell cycle arrest, and potentiated sensitivity to docetaxel (DTX), a first-line chemotherapeutic drug for TNBC treatment. These alterations in cell growth/survival properties were accompanied by induction of CDKN1B, a gene encoding the tumor suppressor p27. We further provided the molecular evidence that FOXE3 could directly bind to the CDKN1B promoter and negatively regulate its transcription in TNBC cells. Importantly, knockdown of combined p27 and FOXE3 reversed the DTX-induced cell growth inhibition observed upon FOXE3 knockdown, indicating that the FOXE3’s effects on TNBC progression were mediated mainly through transcriptional regulation of the p27 signaling. Together, our findings suggest that FOXE3 may function as a potent oncogene during the progression of TNBC, likely affecting cell proliferation, invasion and chemosensitivity, and functioning at least in part through transcriptional repression of p27 signaling.

Positive correlation between programmed death ligand-1 and p53 in triple-negative breast cancer.

PurposeTumors with high mutation load tend to have a stronger immune response in some tumors. The correlation between expression of programmed death ligand-1 (PD-L1), a biomarker of immune response in tumors, and p53, accepted as the most frequently mutated gene in many cancers, in triple-negative breast cancer (TNBC) has not been fully investigated in cancer patients.Materials and methods132 cases of TNBC and 32 cases of non-TNBC paraffin-embedded tissue sections were selected to detect the expression of PD-L1 and p53 by immunohistochemistry, and results were correlated with clinical data and survival outcomes. The staining of PD-L1 in tumor cells (TCs) and tumor-associated immune cells (TAICs) was assessed separately.ResultsStrong positive correlations were observed between expression of p53 and PD-L1 both in TCs (r=0.338, P=0.000) and TAICs (r=0.186, P=0.033). The same positive correlation was found in the expression of PD-L1 in TCs and TAICs (r=0.764, P=0.000). Like p53 (P=0.024), positive rate of PD-L1 in TCs was significantly higher in TNBC than in non-TNBC (P=0.02). PD-L1 and p53 in TCs staining were significantly associated with histological grade, tumor size and Ki67 index (P<0.05). PD-L1 in TCs staining was also associated with lymphatic metastasis status (P=0.000). However, PD-L1 in TAICs was only related to histological grade in statistically (P=0.012). Kaplan–Meier survival analysis showed that positive groups of p53, PD-L1 in TCs and TAICs had a worse overall survival and a worse progression-free survival as compared with the negative groups, but marginal significance was found only in overall survival of PD-L1 in TCs and TAICs, and progression-free survival of PD-L1 in TAICs (P=0.074, 0.097, 0.068, respectively).ConclusionOur findings suggest that positive correlation between p53 and PD-L1 in TNBC and the higher expression rates are closely correlated with some key prognostic factors and worse survival outcomes. These findings would lay the foundation for further study on the relationship of p53 and PD-L1 and the combination of mutated p53 inhibitors and PD-1/PD-L1 antibodies in TNBC.

VEGF and KRAS are Potential Targets of miR-206 Modulation in Triple Negative Breast Cancer

Triple negative is a subtype of breast cancer characterized by lack of expression of hormone receptors (ER, PR and Her2/neu). Due to the limited treatment options, the search for novel treatment targets continues. The aim of this study was to assess the differential expression of miR-206, VEGF and KRAS in TNBC and non-TNBC tissues and cell lines and to evaluate the modulatory effect of miR-206 on the key oncogenic targets VEGF and KRAS. The expression of miR-206, VEGF and KRAS was quantified using real time PCR in both paraffin embedded breast cancer and adjacent tissues as well as in MDA-MB-231 and MCF-7 cell lines. Cell lines were transfected with different concentrations of miR-206 mimic and their viability were assessed using MTT assay. Our results indicated that miR-206 was significantly downregulated in cancerous compared to non-cancerous tissues with a more pronounced downregulation in TNBC than non-TNBC tissues. VEGF and KRAS were significantly upregulated in TNBC compared to non-TNBC and their expression was negatively correlated to miR-206 expression. Transfection of TNBC and non-TNBC cell lines with miR-206 mimic resulted in a dose dependent reduction in cell viability as well as a significant reduction in VEGF and KRAS expression. In conclusion, based on our combined human tissues and cell line-based investigations we can suggest that VEGF and KRAS may be potential targets for miR-206-mediated regulation and that their targeting by miR-206 can be a highly efficient therapeutic strategy in TNBC.

SUN-008 The Glucocorticoid Receptor Is Essential For TGFβ and p38 MAPK Mediated Cancer Phenotypes in Triple Negative Breast Cancer

Triple negative breast cancer (TNBC) is the most metastatic and deadliest breast cancer (BC) subtype, accounting for 20-30% of all BCs. There is a critical need to identify molecular targets that could be exploited as new biomarkers of TNBC prognosis and for improving therapies. Although TNBC lacks estrogen and progesterone receptors, 15-40% of TNBC patients express the glucocorticoid receptor (GR). Women with TNBC that express high levels of GR have poor outcomes. We hypothesize that GR is a key mediator of advanced cancer phenotypes in TNBC. Specifically, we propose that GR acts as a “sensor” for stress signaling pathways commonly activated by soluble factors that are abundant within the tumor microenvironment (TME). Using TNBC models, we showed previously that GR is phosphorylated on Ser134 in response to cellular stress stimuli such as hypoxia. Herein, we show that pS134-GR is elevated in TNBC tumor tissue samples relative to non-TNBC tissues. In vitro studies in TNBC models demonstrate that GR Ser134 phosphorylation is promoted by cytokines (TGFbeta), and growth factors (HGF) and occurs in the absence of GR ligands such as Dexamethasone or cortisol. In response to stress signaling inputs, studies with kinase inhibitors confirmed that p38 MAPK is required for GR Ser134 (pS134-GR) phosphorylation. To evaluate the functional significance of pS134-GR, we created CRISPR models of MDA-MB-231 TNBC cells expressing either wt GR or phospho-mutant GR that cannot be phosphorylated at Ser134 (S134A). RNAseq studies were performed to identify pS134-GR target genes in TNBC models. Our transcriptome data demonstrated a requirement for pS134 GR in the expression of gene sets associated with TGFβ and p38 MAPK signaling. Pathway analysis revealed that pS134-GR target genes primarily regulate cancer cell migration. In vitro assays revealed that pS134-GR is essential for inducing cell migration and anchorage-independent growth in TNBC cells, even in the absence of exogenous GR ligands. Furthermore, using co-IP assays, we identified that upon phosphorylation at Ser134, GR interacts with the scaffolding protein 14-3-3zeta. Like pS134-GR, 14-3-3zeta is highly expressed in TNBC when compared to non-TNBC patients. We observed co-recruitment of both pS134-GR and 14-3-3zeta to known pS134-GR target genes (i.e. PTK6) in TNBC cells. These data prompted us to test the requirement for 14-3-3zeta in GR-mediated phenotypes. Short hairpin RNA knock-down experiments demonstrated that expression of 14-3-3zeta is required for serum and TGFbeta-induced TNBC cell migration. We conclude that the pS134-GR/14-3-3zeta complex is a key “sensor” of local stress signals within the TME (TGFbeta) and a potent mediator of cell migration in TNBC models. Further studies are aimed at exploring pS134 GR as a biomarker and therapeutic target in TNBC.

Human leukocyte antigen I and levels of expression of antigen-presenting element proteins plays a role in triple negative breast cancer

Purpose : To determine the protein expression levels of leukocyte antigen I and antigen-presenting element (APM) genes, and to study their relationship with triple negative breast cancer (TNBC) in patients from Uyghur and Han, China. Methods : Immunohistochemistry was used to determine the expression levels of 10 proteins (HLA-A (Human Leukocyte Antigens-A), HLA-B, HLA-C, TAP1 (Transporter associated with Antigen Processing-1), TAP2, calreticulin, calnexin, ERp57 (Endoplasmic reticulum resident protein 57), ERAP1 (Type 1 tumor necrosis factor receptor shedding aminopeptidase regulator) and tapasin) in TNBC and non-TNBC tissue specimens, and 26 benign lesions (fibrous adenoma) specimens from 120 Uygur and Han women. Results : Immunohistochemical analysis showed that the positive expressions of HLA-A, HLA-B, TAP2, Erp57, ERAP1, calnexin and calreticulin in breast cancer tissues were significantly lower than those in breast fibroma tissues (p < 0.05). Among the 86 TNBC patients, there were 35 cases of tapasin- (40.70 %), 26 cases of tapasin+ (30.30 %), and 25 cases of tapasin++ (29.10 %). Among the 34 non-TNBC patients, there were 25 cases of tapsin- (73.50 %), 7 cases of tapasin+ (20.60 %) and 2 cases of tapasin ++ (5.9 %). The positive expression of tapasin in TNBC patients was significantly higher than that in non-TNBC patients (p < 0.05). Conclusion : Down-regulation of transcriptional expression or loss of protein expression of HLA-I and APM genes is closely related to the progression of breast cancer, and is therefore, a potential molecular marker for screening tumors. Keywords : Triple negative breast cancer, Antigen presenting element, Human leukocyte antigen (HLA)

Focal adhesion kinase (FAK) activation by estrogens involves GPER in triple-negative breast cancer cells

BackgroundFocal adhesion kinase (FAK) is a cytoplasmatic protein tyrosine kinase that associates with both integrins and growth factor receptors toward the adhesion, migration and invasion of cancer cells. The G-protein coupled estrogen receptor (GPER) has been involved in the stimulatory action of estrogens in breast tumor. In this study, we have investigated the engagement of FAK by GPER signaling in triple negative breast cancer (TNBC) cells.MethodsPublicly available large-scale database and patient data sets derived from “The Cancer Genome Atlas” (TCGA; www.cbioportal.org) were used to assess FAK expression in TNBC, non-TNBC tumors and normal breast tissues. MDA-MB 231 and SUM159 TNBC cells were used as model system. The levels of phosphorylated FAK, other transduction mediators and target genes were detected by western blotting analysis. Focal adhesion assay was carried out in order to determine the focal adhesion points and the formation of focal adhesions (FAs). Luciferase assays were performed to evaluate the promoters activity of c-FOS, EGR1 and CTGF upon GPER activation. The mRNA expression of the aforementioned genes was measured by real time-PCR. Boyden chamber and wound healing assays were used in order to evaluate cell migration. The statistical analysis was performed by ANOVA.ResultsWe first determined by bioinformatic analysis that the mRNA expression levels of the gene encoding FAK, namely PTK2, is higher in TNBC respect to non-TNBC and normal breast tissues. Next, we found that estrogenic GPER signaling triggers Y397 FAK phosphorylation as well as the increase of focal adhesion points (FAs) in TNBC cells. Besides, we ascertained that GPER and FAK activation are involved in the STAT3 nuclear accumulation and gene expression changes. As biological counterpart, we show that FAK inhibition prevents the migration of TNBC cells upon GPER activation.ConclusionsThe present data provide novel insights regarding the action of FAK in TNBC. Moreover, on the basis of our findings estrogenic GPER signaling may be considered among the transduction mechanisms engaging FAK toward breast cancer progression.

HIF1A-AS2 predicts poor prognosis and regulates cell migration and invasion in triple-negative breast cancer.

The aberrant expression of hypoxia-inducible factor 1 alpha (HIF1A)-antisense RNA 2 (HIF1A-AS2) was found in various human cancers including breast cancer. The aim of this study was to present more evidence about the role HIF1A-AS2 on triple-negative breast cancer (TNBC). In our results, HIF1A-AS2 was also found to be upregulated in TNBC tissues compared with non-TNBC tissues or adjacent normal tissues. Besides, HIF1A-AS2 expression was also elevated in TNBC cell lines compared with the normal breast epithelial cell line. Moreover, high expression of HIF1A-AS2 was associated with lymph node metastasis, distant metastasis and unfavorable histological grade in TNBC patients. Survival analysis showed a TNBC patient with high HIF1A-AS2 expression had shorter overall survival than patients with low HIF1A-AS2 expression, and HIF1A-AS2 high expression acted as an independent poor prognostic factor for overall survival in TNBC patients. The cell migration and invasion assays suggested inhibition of HIF1A-AS2 obviously depressed TNBC cell migration and invasion. In conclusion, HIF1A-AS2 serves as a novel biomarker for predicting clinical progression and prognosis in TNBC.

The pro-metastasis effect of circANKS1B in breast cancer

BackgroundRecent studies indicate that circular RNA (circRNA) plays a pivotal role in cancer progression. Here, we sought to investigate its role in breast cancer.MethodsCircANKS1B (a circRNA originated from exons 5 to 8 of the ANKS1B gene, hsa_circ_0007294) was identified by RNA-sequencing and validated by qRT-PCR and Sanger sequencing. Clinical breast cancer samples were used to evaluate the expression of circANKS1B and its associations with clinicopathological features and prognosis. Gain- and loss-of-function experiments in cell lines and mouse xenograft models were performed to support clinical findings and elucidate the function and underlying mechanisms of circANKS1B in breast cancer.ResultsCircANKS1B was significantly up-regulated in triple-negative breast cancer (TNBC) compared with non-TNBC tissues and cell lines. Increased circANKS1B expression was closely associated with lymph node metastasis and advanced clinical stage and served as an independent risk factor for overall survival of breast cancer patients. Functional studies revealed that circANKS1B promoted breast cancer invasion and metastasis both in vitro and in vivo by inducing epithelial-to-mesenchymal transition (EMT), while had no effect on breast cancer growth. Mechanistically, circANKS1B abundantly sponged miR-148a-3p and miR-152-3p to increase the expression of transcription factor USF1, which could transcriptionally up-regulate TGF-β1 expression, resulting in activating TGF-β1/Smad signaling to promote EMT. Moreover, we found that circANKS1B biogenesis in breast cancer was promoted by splicing factor ESRP1, whose expression was also regulated by USF1.ConclusionsOur data uncover an essential role of the novel circular RNA circANKS1B in the metastasis of breast cancer, which demonstrate that therapeutic targeting of circANKS1B may better prevent breast cancer metastasis.

HDAC9 overexpression confers invasive and angiogenic potential to triple negative breast cancer cells via modulating microRNA-206

Triple negative breast cancer (TNBC) is among the most aggressive breast cancer subtypes with poor prognosis. The purpose of this study is to better understand the molecular basis of TNBC as well as develop new therapeutic strategies. Our results demonstrate that HDAC9 is overexpressed in TNBC compared to non-TNBC cell lines and tissues and is inversely proportional with miR-206 expression levels. We show that HDAC9 selective inhibition blocked the invasion of TNBC cells in vitro and repressed the angiogenesis shown via in vivo Matrigel plug assays. Subsequent HDAC9 siRNA knockdown was then shown to restore miR-206 while also decreasing VEGF and MAPK3 levels. Furthermore, the inhibition of miR-206 neutralized the action of HDAC9 siRNA on decreasing VEGF and MAPK3 levels. This study highlights HDAC9 as a mediator of cell invasion and angiogenesis in TNBC cells through VEGF and MAPK3 by modulating miR-206 expression and suggests that selective inhibition of HDAC9 may be an efficient route for TNBC therapy.

Downregulation of RIP140 in triple-negative breast cancer inhibits the growth and proliferation of cancer cells.

The aim of the present study was to investigate the expression of receptor interacting protein 140 (RIP140) in triple-negative breast cancer (TNBC), and its association with the prognosis of patients with TNBC. A total of 179 patients with breast cancer were included in this study, with 41 cases of TNBC and 138 cases of non-TNBC. Immunohistochemical staining and western blotting were used to detect the protein expression of RIP140 in the cancerous and paracancerous tissues, revealing that expression of RIP140 was increased in TNBC tissues compared with non-TNBC tissues. High expression of RIP140 in breast cancer tissue was associated with a poorer survival time than low RIP140 expression. Using lentiviral transfection to downregulate RIP140 expression in MDA-MB-231 cells, the effects of RIP140 on the growth and proliferation of breast cancer cells was analyzed using subcutaneous tumors in BALB/c nude mice. Immunohistochemical staining using a Ki-67 antibody in subcutaneous tumor tissue was used to assess the proliferation of MDA-MB-231 cells. The short hairpin RNA-mediated downregulation of RIP140 in MDA-MB-231 cells suppressed the growth and the proliferation of subcutaneous tumors in BALB/c nude mice. Downregulation of RIP140 in breast cancer cells may therefore inhibit the growth and the proliferation of these cells, and may provide a therapeutic target for TNBC.

SOCS1 is associated with clinical progression and acts as an oncogenic role in triple-negative breast cancer.

Suppressors of cytokine signaling 1 (SOCS1) is a member of SOCS family and acts as negative regulators of cytokine signaling by direct inhibition of receptor-associated janus kinases. The clinical significance and biological function of SOCS1 in variant tumor tissues and at variant tumor stages is still controversial. The aim of our study is to confirm the expression status of SOCS1 in triple-negative breast cancer (TNBC) tissues and cell lines, and explore the clinical value and biological function of SOCS1 in TNBC. In microarray data sets (GDS2250 and GDS817), we observed SOCS1 was overexpressed in TNBC tissues and cell line compared with normal mammary tissues and mammary epithelial cell line, or non-TNBC tissues and cell line. Furthermore, SOCS1 mRNA and protein overexpression were confirmed in TNBC tissues and cell lines compared with normal mammary tissues and mammary epithelial cell lines or non-TNBC tissues and cell lines. SOCS1 protein overexpression was obviously associated with advanced clinical stage, large tumor size, more lymph node metastasis, present distant metastasis, and malign histological grade. Downregulation of SOCS1 expression suppressed TNBC cells proliferation and promoted cell apoptosis. In conclusion, SOCS1 is associated with clinical progression in TNBC patients and acts as an oncogenic role in regulating TNBC cells proliferation and apoptosis. © 2018 IUBMB Life, 70(4):320-327, 2018.

MicroRNA-130a suppresses breast cancer cell migration and invasion by targeting FOSL1 and upregulating ZO-1.

FOSL1 is frequently overexpressed in multiple types of human cancers including invasive breast cancers and implicated in cancer invasion and metastasis. However, how FOSL1 is overexpressed in cancers remains to be elucidated. Several microRNAs (miRNAs) have been shown to target FOSL1 and are downregulated in human cancers. Here, we report that miR-130a is a novel FOSL1 targeting miRNA. Using gene expression microarray analysis, we found that FOSL1 is among the most up-regulated genes in cells transfected with miR-130a inhibitors. Transient transfection-immunoblot, RNA-immunoprecipitation, and luciferase reporter assays revealed that miR-130a directly targets FOSL1 mRNA at its 3'-UTR. Overexpression of miR-130a significantly reduced the levels of FOSL1 in invasive breast cancer MDA-MB-231 and Hs578T cell lines and suppresses their migration and invasion. This inhibition can be rescued by ectopic expression of miR-130a-resistant FOSL1. Interestingly, we show that overexpression of miR-130a increased the levels of tight-junction protein ZO-1 while inhibition of miR-130a reduced the levels of ZO-1. We further show that miR-130a expression is significantly reduced in cancer tissues from triple-negative breast cancer (TNBC) patients, correlating significantly with the upregulation of FOSL1 expression, compared to non-TNBC tissues. Together, our results reveal that miR-130a directly targets FOSL1 and suppresses the inhibition of ZO-1, thus inhibiting cancer cell migration and invasion, in TNBCs.

A Comprehensive Immunologic Portrait of Triple-Negative Breast Cancer

Triple-negative breast cancer (TNBC) is a high-risk malignancy due to its high capacity for invasion and lack of targeted therapy. Immunotherapy continues to demonstrate efficacy in a variety of cancers, and thus may be a promising strategy for TNBC given the limited therapeutic options currently available for TNBC. In this study, we performed an exhaustive analysis of immunogenic signatures in TNBC based on 2 large-scale breast cancer (BC) genomic data. We compared enrichment levels of 26 immune cell activities and pathways among TNBC, non-TNBC, and normal tissue, and within TNBCs of different genotypic or phenotypic features. We found that almost all analyzed immune activities and pathways had significantly higher enrichment levels in TNBC than non-TNBC. Elevated enrichment of these immune activities and pathways was likely to be associated with better survival prognosis in TNBC. This study demonstrated that TNBC likely exhibits the strongest immunogenicity among BC subtypes, and thus warrants the immunotherapeutic option for TNBC.

Increased expression of biological markers as potential therapeutic targets in Saudi women with triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer that lacks the expression of hormone receptors and human epidermal growth factor receptor 2 (HER2). Although TNBC represents only 15% of all types of breast cancer, it accounts for a large number of metastatic cases and deaths. Because of the high metastatic rate and both local and systemic recurrence associated with TNBC, extensive research efforts are actively looking for target therapies to effectively treat this aggressive disease. Accordingly, this study has been initiated to investigate the differential expression of biological markers in TNBC and non-TNBC Saudi women that might be utilized as potential targeted therapy and/or predict the sensitivity to currently available therapeutic regimens. Two hundred formalin-fixed, paraffin-embedded (FFPE) breast cancer tissues were selected and divided into 3 groups: benign breast tissues (20), TNBC tissues (80) and non-TNBC tissues (100). Expression of mRNA in FFPE tissues was analyzed using real-time polymerase chain reaction (RT-PCR) for the following genes: poly (ADP-ribose) polymerase 1 (PARP-1), topoisomerase 2A (TOPO-2A), vascular endothelial growth factor (VEGF), C-MYC, basic fibroblast growth factor (bFGF), matrix metalloproteinases (MMP-2 and MMP-9), human epidermal growth factor 1 (HER1) and multidrug resistance (MDR) genes. In the TNBC group, expression of PARP-1, TOPO-2A, HER1, C-MYC, VEGF, bFGF and MMP-2 showed a highly significant increase compared to the non-TNBC group. The results of this study suggest that (1) TNBC patients will benefit more from TOPO-2A inhibitors as well as antiangiogenic and antimetastatic therapies; (2) inhibition of these target genes is emerging as one of the most exciting and promising targeted therapeutic strategies to treat TNBC in which the intended targets are DNA repair, tumor angiogenesis and metastasis.

MiR-613 inhibits cell migration and invasion by downregulating Daam1 in triple-negative breast cancer

Dishevelled-associated activator of morphogenesis 1 (Daam1) is a formin protein and participates in regulating cell migration of triple-negative breast cancer (TNBC) cells. The specific miRNA targeting Daam1 and mediating cell migration and invasion remains obscure. This experiment investigated the suppressive role of miR-613 in TNBC cells. The luciferase activity of Daam1 3′-untranslated region (3′-UTR) based reporters constructed in HEK-293T and MCF-7 cells suggested that Daam1 was the target gene of miR-613. Overexpressed miR-613 reduced the protein level of Daam1, weakened RhoA activity, and retarded the cell migration, cell invasion and colony formation of TNBC cells. Overexpression of Daam1 or RhoA rescued cell migration and invasion in miR-613-overexpressed TNBC cells, but failed to reverse colony formation. MiR-613 was significantly downregulated in breast cancer tissues compared with that in adjacent normal tissues. This downregulation in TNBC tissues and lymphnode metastatic breast cancer tissues was more obvious than that in non-TNBC tissues and non-metastatic cancer tissues, respectively. MiR-613 weakens the resistance of TNBC cells against paclitaxel rather than adriamycin, cyclophosphamide, docetaxel, and kaempferol. Taken together, miR-613 is involved in cell migration and invasion of TNBC cells via targeting Daam1/RhoA signaling pathway.