Promotes Breast Cancer Cell Research Articles

Potassium Channel Protein KCNK6 Promotes Breast Cancer Cell Proliferation, Invasion, and Migration.

Breast cancer is the most common malignant tumor in women, and its incidence is increasing each year. To effectively treat breast cancer, it is important to identify genes involved in its occurrence and development and to exploit them as potential drug therapy targets. Here, we found that potassium channel subfamily K member 6 (KCNK6) is significantly overexpressed in breast cancer, however, its function in tumors has not been reported. We further verified that KCNK6 expression is upregulated in breast cancer biopsies. Moreover, overexpressed KCNK6 was found to enhance the proliferation, invasion, and migration ability of breast cancer cells. These effects may occur by weakening cell adhesion and reducing cell hardness, thus affecting the malignant phenotype of breast cancer cells. Our study confirmed, for the first time, that increased KCNK6 expression in breast cancer cells may promote their proliferation, invasion, and migration. Moreover, considering that ion channels serve as therapeutic targets for many small molecular drugs in clinical treatment, targeting KCNK6 may represent a novel strategy for breast cancer therapies. Hence, the results of this study provide a theoretical basis for KCNK6 to become a potential molecular target for breast cancer treatment in the future.

MiR-934 promotes breast cancer metastasis by regulation of PTEN and epithelial–mesenchymal transition

Breast cancer (BC) is the most commonly diagnosed malignancy and the leading cause of cancer-related mortality among females. Over 90 % of the cases of death in BC patients are attributed to tumor cell metastasis. Therefore, it is urgently needed to investigate the molecular mechanisms of BC metastasis. The expression of miRNA in BC was evaluated by qRT-PCR and bioinformatics analysis. Clone formation, EdU assays, and subcutaneous xenograft model were used to test the growth of BC cells. Wound healing, transwell assays, and lung-metastasis model were used to explore the effect of miR-934 knockdown on cell metastasis. The miR-934 targets in BC were identified through bioinformatics analysis and luciferase reporter assays. The expression of protein was tested by western blot. The binding of mRNA and RNA-binding-protein was verified using RIP assays. miR-934 expression was significantly elevated in BC tissues, especially in those with lymph node metastasis and associated with poor patient prognosis. Experiments in vitro and in vivo showed that that upregulated miR-934 was not necessarily required for the growth of BC cells. However, miR-934 knockdown significantly inhibited the migration and invasion abilities of BC cells. Moreover, PTEN as identified as the direct target of miR-934 in BC, and miR-934 could promote BC cell metastasis by regulation of PTEN and epithelial–mesenchymal transition (EMT). Our results suggested that targeting miR-934 may be a practical treatment for BC cell metastasis.

A Wnt-Independent LGR4-EGFR Signaling Axis in Cancer Metastasis.

Leucine-rich repeat-containing G protein-coupled receptors 4, 5, and 6 (LGR4/5/6) play critical roles in development and cancer. The widely accepted mechanism is that these proteins, together with their R-spondin ligands, stabilize Wnt receptors, thus potentiating Wnt signaling. Here we show that LGR4 enhanced breast cancer cell metastasis even when Wnt signaling was deactivated pharmacologically or genetically. Furthermore, LGR4 mutants that cannot potentiate Wnt signaling nevertheless promoted breast cancer cell migration and invasion in vitro and breast cancer metastasis in vivo. Multiomic screening identified EGFR as a crucial mediator of LGR4 activity in cancer progression. Mechanistically, LGR4 interacted with EGFR and blocked EGFR ubiquitination and degradation, resulting in persistent EGFR activation. Together, these data uncover a Wnt-independent LGR4-EGFR signaling axis with broad implications for cancer progression and targeted therapy. SIGNIFICANCE: This work demonstrates a Wnt-independent mechanism by which LGR4 promotes cancer metastasis.See related commentary by Stevens and Williams, p. 4397.

Calycosin inhibits breast cancer cell migration and invasion by suppressing EMT via BATF/TGF-β1.

In this study, we investigated the effects of calycosin on breast cancer cell progression and their underlying mechanisms. Calycosin dose- and time-dependently inhibited proliferation, migration, and invasion by T47D and MCF-7 breast cancer cells by downregulating basic leucine zipper ATF-like transcription factor (BATF) expression. Moreover, BATF promoted breast cancer cell migration and invasiveness by increasing TGFβ1 mRNA and protein levels. Bioinformatics analysis, dual luciferase reporter assays, and chromatin immunoprecipitation assays confirmed the presence of BATF-binding sites in the promoter sequence of TGFβ1 gene. Calycosin treatment inhibited epithelial-mesenchymal transition (EMT) of breast cancer cells by significantly increasing E-cadherin levels and decreasing N-cadherin, Vimentin, CD147, MMP-2, and MMP-9 levels through downregulation of BATF and TGFβ1. TGFβ1 knockdown reduced the migration and invasiveness of BATF-overexpressing breast cancer cells, whereas incubation with TGFβ1 enhanced the migration and invasiveness of calycosin-treated breast cancer cells. Our findings demonstrated that calycosin inhibited EMT and progression of breast cancer cells by suppressing BATF/TGFβ1 signaling. This suggests calycosin would be a promising therapeutic option for breast cancer patients.

Decreased microRNA-768-3p expression indicates a poor prognosis in patients with breast cancer and promotes breast cancer cell viability, migration and invasion.

Breast cancer is the most common malignancy in women and microRNA-768-3p (miR-768-3p) is abnormally expressed in hepatocellular carcinoma, non-small cell lung carcinomas and melanoma. The aim of the present study was to evaluate the prognostic value and biological function of miR-768-3p in breast cancer. The expression of miR-768-3p in tumor tissues and adjacent tissues of 116 patients with breast cancer obtained by surgery and normal breast cell lines MCF-10A and breast cancer cell lines (MCF-7, MDA-MB-231, T-47D and SK-BR-3) were detected by reverse transcription-quantitative PCR. The association between miR-768-3p expression and the clinicopathological characteristics of patients was analyzed using the χ2 test. In addition, the Kaplan-Meier method was used for survival analysis. A Cox regression model was used to examine the effect of miR-768-3p on the prognosis of patients with breast cancer. Hemocytometer cell counting and Transwell assays were used to detect the effects of miR-768-3p on the characteristics of breast cancer cells. The target genes of miR-768-3p in breast cancer were identified by bioinformatics software and detected by luciferase reporter assay. Compared with normal tissues and normal breast cancer cells, miR-768-3p was significantly decreased in breast cancer tissues and cancer cells (P<0.001). The reduction in miR-768-3p was significantly associated with lymph node metastasis (P=0.040), Tumor Node Metastasis stage (P=0.035), and cancer subtype (P=0.008). In addition, patients with low miR-768-3p expression had a shorter overall survival time (log-rank P=0.022) compared with those with high expression and miR-768-3p may be a potential prognostic marker (hazard ratio=4.637; 95% confidence interval=1.296–16.597; P=0.018). When transfected with miR-768-3p inhibitor, cell viability, migration and invasion were significantly promoted compared with the control group (P<0.05). In addition, eukaryotic translation initiation factor 4E (eIF4E) was the target gene of miR-768-3p in breast cancer. All experiments confirmed that miR-768-3p, a tumor suppressor, inhibited the viability, migration and invasion of breast cancer cells through eIF4E. miR-768-3p may be a potential prognostic marker of breast cancer and may participate in the progression of breast cancer.

Crosstalk between progesterone receptor membrane component 1 and estrogen receptor α promotes breast cancer cell proliferation

Progesterone (P4) and estradiol (E2) have been shown to stimulate and regulate breast cancer proliferation via classical nuclear receptor signaling through progesterone receptor (PR) and estrogen receptor α (ERα), respectively. However, the basis of communication between PR/ERα and membrane receptors remains largely unknown. Here, we aim to identify classical and nonclassical endocrine signaling mechanisms that can alter cell proliferation through a possible crosstalk between PR, ERα, and progesterone receptor membrane component 1 (PGRMC1), a membrane receptor frequently observed in breast cancer cells. While P4 and E2 treatment increased cell proliferation of ER+/PR+/PGRMC1 overexpressing breast cancer cells, silencing ERα and PR or treatment with selective estrogen receptor modulator (SERM) tamoxifen, or (PR-antagonist) RU-486 decreased cell proliferation. All four treatments rapidly altered PGRMC1 mRNA levels and protein expression. Furthermore, P4 and E2 treatments rapidly activated EGFR a known interacting partner of PGRMC1 and its downstream signaling. Interestingly, downregulation of ERα by tamoxifen and ERα silencing decreased the expression levels of PGRMC1 with no repercussions to PR expression. Strikingly PGRMC1 silencing decreased ERα expression irrespective of PR. METABRIC and TCGA datasets further demonstrated that PGRMC1 expression was comparable to that of ERα in Luminal A and B breast cancers. Targeting of PR, ERα, and PGRMC1 confirmed that a crosstalk between classical and nonclassical signaling mechanisms exists in ER+ breast cancer cells that could enhance the growth of ER+/PR+/PGRMC1 overexpressing tumors.

ETV4 promotes breast cancer cell stemness by activating glycolysis and CXCR4-mediated sonic Hedgehog signaling

Cancer stem cells (CSCs) are a major cause of tumor treatment resistance, relapse and metastasis. Cancer cells exhibit reprogrammed metabolism characterized by aerobic glycolysis, which is also critical for sustaining cancer stemness. However, regulation of cancer cell metabolism rewiring and stemness is not completely understood. Here, we report that ETV4 is a key transcription factor in regulating glycolytic gene expression. ETV4 loss significantly inhibits the expression of HK2, LDHA as well as other glycolytic enzymes, reduces glucose uptake and lactate release in breast cancer cells. In human breast cancer and hepatocellular carcinoma tissues, ETV4 expression is positively correlated with glycolytic signaling. Moreover, we confirm that breast CSCs (BCSCs) are glycolysis-dependent and show that ETV4 is required for BCSC maintenance. ETV4 is enriched in BCSCs, its knockdown and overexpression suppresses and promotes breast cancer cell stem-like traits, respectively. Mechanistically, on the one hand, we find that ETV4 may enhance glycolysis activity to facilitate breast cancer stemness; on the other, ETV4 activates Sonic Hedgehog signaling by transcriptionally promoting CXCR4 expression. A xenograft assay validates the tumor growth-impeding effect and inhibition of CXCR4/SHH/GLI1 signaling cascade after ETV4 depletion. Together, our study highlights the potential roles of ETV4 in promoting cancer cell glycolytic shift and BCSC maintenance and reveals the molecular basis.

SEC61G promotes breast cancer development and metastasis via modulating glycolysis and is transcriptionally regulated by E2F1

Breast cancer is the most common cancer in women and its incidence rates are rapidly increasing in China. Understanding the molecular mechanisms of breast cancer tumorigenesis enables the development of novel therapeutic strategies. SEC61G is a subunit of the endoplasmic reticulum translocon that plays critical roles in various tumors. We aimed to investigate the expression and function of SEC61G in breast cancer. By analyzing The Cancer Genome Atlas breast cancer cohort, we found that SEC61G was highly expressed in breast cancer and predicted poor prognosis of breast cancer patients. Overexpression of SEC61G and its prognostic role was also confirmed in the Nanjing Medical University (NMU) breast cancer cohort. Functionally, we demonstrated that knockdown of SEC61G suppressed breast cancer cell proliferation, migration, invasion, and promoted breast cancer cell apoptosis in vitro. Xenograft breast tumor model revealed that knockdown of SEC61G inhibited breast tumor development in vivo. Furthermore, we demonstrated that SEC61G positively regulated glycolysis in breast cancer cells. Mechanistically, we showed that transcription factor E2F1 directly bound to the promoter of SEC61G and regulated its expression in breast cancer cells. SEC61G overexpression antagonized the effect of E2F1 knockdown in regulating breast cancer cell proliferation, invasion, and apoptosis. Finally, we demonstrated that the E2F1/SEC61G axis regulated glycolysis and chemo-sensitivity of Herceptin in breast cancer cells. Taken together, these results of in vitro and in vivo studies demonstrate that SEC61G promotes breast cancer development and metastasis via modulating glycolysis and is transcriptionally regulated by E2F1, which might be utilized as a promising therapeutic target of breast cancer treatment.

Loss of polarity protein Par3 is mediated by transcription factor Sp1 in breast cancer

Loss of polarity protein Par3 promotes breast cancer tumorigenesis and metastasis. The underlying molecular mechanisms of Par3 down-regulation and related prognostic significance in breast cancer remain unclear. Here, we discovered that Par3 down-regulation was associated with shorter relapse-free survival in Luminal A subtype of breast cancer. Par3 knockdown promoted breast cancer cells migration and invasion. Importantly, we identified that transcription factor Sp1 bound to PARD3 promoter region and induced Par3 expression. Breast cancer patients with low Sp1 showed significantly worse RFS and low expression level of Par3. Par3 over-expression partially reversed Sp1 knockdown induced migration and invasion. Together, decreased Sp1 level mediates Par3 down-regulation, which correlated with poor prognosis of ER + breast cancer patients, via reduced binding with PARD3 promoter.

Anti-PITPNM3 small molecular compounds reverse breast cancer metastasis by targeting PITPNM3.

e15005 Background: Recent studies highlight the fundamental roles of PITPNM3 in breast cancer metastasis. PITPNM3 is identified as the functional receptor of CCL18 and promotes breast cancer cell invasion and metastasis by binding with CCL18. Since anti-CCL18 neutralized antibodies shows medium binding affinity which restricts their clinical application, small molecular inhibitors targeting PITPNM3 are needed to be further investigated. Therefore, we identified several first in class small molecular inhibitors potentially targeting PITPNM3 and can inhibit breast cancer metastasis conducted by PITPNM3 activation. Methods: We performed computer-assisted drug design by constructing PITPNM3 homology model, characterizing potential binding pockets and docking preselected high diversity structured small molecule compounds into the static PITPNM3 model. Top 100 small molecules in silico scores were selected and screened through basic experiments. After screening, the anti-metastasis effects of selected compounds were tested through transwell migration and invasion assay. Immunofluorescence and qPCR were applied to confirm the expression of vimentin and CDH1. Western blot were used to clarify the inhibition effects of selected compounds on PITPNM3 signaling pathways. Results: By using homology remodeling, we successfully constructed the PITPNM3(680-920aa) protein model. The PITPNM3(680-920aa) domain is responsible for interacting with PYK2 and phosphorylating PYK2. The phosphorylation of PYK2 conducted by PITPNM3 signaling pathway will lead to metastasis and epithelial-mesenchymal transition (EMT) of breast cancer cells. We then characterized the potential binding pockets of this static model and a druggable site was founded. More than 50K molecules with high diversity were docked into this druggable site and scored through their docking performance. Finally, top 100 scored small molecules were selected. In addition, through 1 rounds of toxicity screening, 1 round of transwell migration assay screening and 1 round of transwell invasion assay screening, 4 small molecules with higher bioactivity is identified and 1 compound with the highest bioactivity as well as docking performance among 50K small molecules is chose. This compound can inhibit CCL18 treatment as well as tumor associated macrophage co-culture mediated migration and invasion. Besides, it can also inhibit the phophorylation of PYK2 and Src without inhibition the expression of PITPNM3. Conclusions: Our findings identify the first-in-class anti-PITPNM3 small molecule inhibitors. These compounds can inhibit PITPNM3 signaling pathway and reverse breast cancer metastasis.

Coexpression of EphA10 and Gli3 promotes breast cancer cell proliferation, invasion and migration

This study investigated the influences of EphA10 and Gli3 on breast cancer (BC) cell proliferation, invasion and migration. Immunohistochemistry was used to reveal the expressions of EphA10 and Gli3 in...

MicroRNA‑454‑5p promotes breast cancer progression by inducing epithelial‑mesenchymal transition via targeting the FoxJ2/E‑cadherin axis.

MicroRNAs are important for the regulation of multiple cellular functions and are involved in the initiation and progression of various types of cancer, including breast cancer. Although microRNA (miR)‑454‑3p is reported to function as an oncogene in several types of human cancer, the role of miR‑454‑5p in breast cancer remains unknown. The present study demonstrated that miR‑454‑5p was upregulated in breast cancer and was associated with a poor prognosis in patients with breast cancer. Overexpression of miR‑454‑5p promoted breast cancer cell viability, migration and invasion invitro, whereas silencing of miR‑454‑5p inhibited breast cancer proliferation, migration and invasion invitro and suppressed tumor growth invivo. Mechanistically, forkhead box J2 (FoxJ2) was shown to be a target of miR‑454‑5p and transactivated E‑cadherin expression. Moreover, silencing of miR‑454‑5p reversed the epithelial‑mesenchymal transition phenotype through upregulation of the FoxJ2/E‑cadherin axis. Collectively, the present findings suggested that miR‑454‑5p may serve as a novel therapeutic target and prognostic predictor for patients with breast cancer.

MiR-1224-3p Promotes Breast Cancer Cell Proliferation and Migration through PGM5-Mediated Aerobic Glycolysis.

Metabolic reprogramming of aerobic glycolysis is a hallmark of cancer cells. Regulators of aerobic glycolysis have become targets for cancer diagnosis and therapy. However, the regulators of aerobic glycolysis in breast cancer development have not been well elucidated. Here, we show that the phosphoglucomutase (PGM) family member PGM5 promotes conversion of glucose-1-phosphate (G1P) into glucose-6-phosphate (G6P) and inhibits breast cancer cell proliferation and migration through regulating aerobic glycolysis. In breast cancer patients, PGM5 is significantly downregulated, and its low expression is a predictor of poor prognosis. MicroRNA-1224-3p (miR-1224-3p) inhibits the PGM5 level through directly targeting its 3'-untranslated region and suppresses PGM5-mediated breast cancer cell proliferation, migration, and glycolytic function. Moreover, the miR-1224-3p/PGM5 axis regulates the expression of cell cycle- and apoptosis-related genes and the markers of epithelial-mesenchymal transition (EMT), a process involved in migration and metastasis of cancer cells. Taken together, our results indicate that miR-1224-3p/PGM5 axis plays important roles in breast cancer cell proliferation, migration, and aerobic glycolysis and may be a potential target for breast cancer therapy.

PAPOLA contributes to cyclin D1 mRNA alternative polyadenylation and promotes breast cancer cell proliferation.

Poly(A) polymerases add the poly(A) tail at the 3' end of nearly all eukaryotic mRNA, and are associated with proliferation and cancer. To elucidate the role of the most-studied mammalian poly(A) polymerase, poly(A) polymerase α (PAPOLA), in cancer, we assessed its expression in 221 breast cancer samples and found it to correlate strongly with the aggressive triple-negative subtype. Silencing PAPOLA in MCF-7 and MDA-MB-231 breast cancer cells reduced proliferation and anchorage-independent growth by decreasing steady-state cyclin D1 (CCND1) mRNA and protein levels. Whereas the length of the CCND1 mRNA poly(A) tail was not affected, its 3' untranslated region (3'UTR) lengthened. Overexpressing PAPOLA caused CCND1 mRNA 3'UTR shortening with a concomitant increase in the amount of corresponding transcript and protein, resulting in growth arrest in MCF-7 cells and DNA damage in HEK-293 cells. Such overexpression of PAPOLA promoted proliferation in the p53 mutant MDA-MB-231 cells. Our data suggest that PAPOLA is a possible candidate target for the control of tumor growth that is mostly relevant to triple-negative tumors, a group characterized by PAPOLA overexpression and lack of alternative targeted therapies.

Defects in Emerin-Nucleoskeleton Binding Disrupt Nuclear Structure and Promote Breast Cancer Cell Motility and Metastasis.

Nuclear envelope proteins play an important role in regulating nuclear size and structure in cancer. Altered expression of nuclear lamins are found in many cancers and its expression is correlated with better clinical outcomes. The nucleus is the largest organelle in the cell with a diameter between 10 and 20 μm. Nuclear size significantly impacts cell migration. Nuclear structural changes are predicted to impact cancer metastasis by regulating cancer cell migration. Here we show emerin regulates nuclear structure in invasive breast cancer cells to impact cancer metastasis. Invasive breast cancer cells had 40% to 50% less emerin than control cells, which resulted in decreased nuclear size. Overexpression of GFP-emerin in invasive breast cancer cells rescued nuclear size and inhibited migration through 3.0 and 8.0 μm pores. Mutational analysis showed emerin binding to nucleoskeletal proteins was important for its regulation of nuclear structure, migration, and invasion. Importantly, emerin expression inhibited lung metastasis by 91% in orthotopic mouse models of breast cancer. Emerin nucleoskeleton-binding mutants failed to inhibit metastasis. These results support a model whereby emerin binding to the nucleoskeleton regulates nuclear structure to impact metastasis. In this model, emerin plays a central role in metastatic transformation, because decreased emerin expression during transformation causes the nuclear structural defects required for increased cell migration, intravasation, and extravasation. IMPLICATIONS: Modulating emerin expression and function represents new targets for therapeutic interventions of metastasis, because increased emerin expression rescued cancer metastasis.

Cimicifuga dahurica extract inhibits the proliferation, migration and invasion of breast cancer cells MDA-MB-231 and MCF-7 in vitro and in vivo

Ethnopharmacological relevanceCimicifuga dahurica (Turcz.) Maxim (C. dahurica) has a long history of treating breast cancer. From the Qing Dynasty to the Tang Dynasty and even earlier, C. dahurica has been documented in the treatment of breast carbuncle (Breast cancer is classified as breast carbuncle in Chinese medicine). In traditional prescriptions such as “Sheng Ge Decoction”, “Sheng Ma Powder” and “Breast Carbuncle Pill”, as the main medicine, C. dahurica plays an important role. At present, the systematic studies on the in vitro and in vivo effects of Cimicifuga against breast cancer are rare, especially the C. dahurica. Aim of the studyIn this article, we evaluated the in vitro activity and in vivo effects of CREE (extract of the root of C. dahurica) against breast cancer, and discussed the possible mechanism of CREE in promoting breast cancer cell apoptosis. Materials and methodsThe main component in the CREE was analyzed by HPLC. The effects of CREE on the proliferation, migration and invasion of human breast cancer cells were evaluated through SRB, colony assay, LDH release, wound healing and transwell assay. The pro-apoptotic effect of CREE was investigated in Hochest33342 and Annexin V-FITC/PI assay. To verify the results of CREE in vivo effects, we applied nude mice subcutaneous xenograft experiments. The possible mechanism of CREE treating breast cancer was investigated through mitochondrial membrane potential and western blot experiments. ResultsCREE contains cycloartane triterpene saponins. CREE can significantly inhibit the proliferation, migration and invasion of human breast cancer MCF-7 and MDA-MB-231 cells in vitro and it can effectively inhibit the growth of MDA-MB-231 cell subcutaneous tumors in vivo. Besides, we also found that CREE up-regulated the expression levels of Bax, caspase-9/3 and cytochrome C, and down-regulated the expression of Bcl-2. Therefore, regulation of the mitochondrial pathway may be one of the mechanisms by which CREE promotes breast cancer cell apoptosis. ConclusionsCREE exhibits sufficient anti-breast cancer activity in vivo and in vitro, this study provides persuasive evidence for the further research and development of C. dahurica.

Exosomal miR-222 from adriamycin-resistant MCF-7 breast cancer cells promote macrophages M2 polarization via PTEN/Akt to induce tumor progression.

Exosome-mediated intercellular communication is considered to be an effective mode for malignant cells to transform biological behaviors in stromal cells. However, the mechanisms by which exosomes modulate macrophages within tumor microenvironment remain largely unclear. In this study, we found that both adriamycin-resistant breast cancer (BCa) cells and the corresponding exosomes (A/exo) were capable of inducing macrophages M2 polarization, which promoted the mobility, proliferation, migration and invasion of BCa cells. Since exosomes deliver microRNAs to affect cellular functions in recipient cells, we confirmed that miR-222 was significantly enriched in A/exo and could be successfully transferred to macrophages. Increased miR-222 level was also detected in exosomes derived from plasma and tissues of chemoresistant patients. Moreover, exosomal miR-222 from A/exo polarized M2 macrophages by targeting PTEN and activating Akt signaling pathway, which promoted BCa cells progression in a feed back loop. Co-culture of adriamycin-resistant BCa cells with macrophages in which miR-222 was upregulated or treated with A/exo facilitated tumor growth in vivo. Collectively, our data demonstrated that chemoresistant BCa cells could remodel macrophages within tumor microenvironment by secreting exosomal miR-222, which directly targeted PTEN and caused Akt cascade activation and macrophages M2 polarization. Our findings may provide a foundation for a promising strategy of BCa treatment by targeting exosomes or exosomal miR-222.

Activation of TMEM16A Ca2+-activated Cl− channels by ROCK1/moesin promotes breast cancer metastasis

IntroductionTransmembrane protein 16A (TMEM16A) is a Ca2+-activated chloride channel that plays a role in cancer cell proliferation, migration, invasion, and metastasis. However, whether TMEM16A contributes to breast cancer metastasis remains unknown. ObjectiveIn this study, we investigated whether TMEM16A channel activation by ROCK1/moesin promotes breast cancer metastasis. MethodsWound healing assays and transwell migration and invasion assays were performed to study the migration and invasion of MCF-7 and T47D breast cancer cells. Western blotting was performed to evaluate the protein expression, and whole-cell patch clamp recordings were used to record TMEM16A Cl− currents. A mouse model of breast cancer lung metastasis was generated by injecting MCF-7 cells via the tail vein. Metastatic nodules in the lung were assessed by hematoxylin and eosin staining. Lymph node metastasis, overall survival, and metastasis-free survival of breast cancer patients were assessed using immunohistochemistry and The Cancer Genome Atlas dataset. ResultsTMEM16A activation promoted breast cancer cell migration and invasion in vitro as well as breast cancer metastasis in mice. Patients with breast cancer who had higher TMEM16A levels showed greater lymph node metastasis and shorter survival. Mechanistically, TMEM16A promoted migration and invasion by activating EGFR/STAT3/ROCK1 signaling, and the role of the TMEM16A channel activity was important in this respect. ROCK1 activation by RhoA enhanced the TMEM16A channel activity via the phosphorylation of moesin at T558. The cooperative action of TMEM16A and ROCK1 was supported through clinical findings indicating that breast cancer patients with high levels of TMEM16A/ROCK1 expression showed greater lymph node metastasis and poor survival. ConclusionOur findings revealed a novel mechanism underlying TMEM16A-mediated breast cancer metastasis, in which ROCK1 increased TMEM16A channel activity via moesin phosphorylation and the increase in TMEM16A channel activities promoted cell migration and invasion. TMEM16A inhibition may be a novel strategy for treating breast cancer metastasis.

LINC00052 promotes breast cancer cell progression and metastasis by sponging miR‑145‑5p to modulate TGFBR2 expression

Long non-coding RNAs (lncRNAs) may participate in biological regulatory mechanisms of tumors. The aim of the present study was to uncover the molecular mechanism of the lncRNA LINC00052 in the tumorigenesis of breast cancer (BC). LINC00052 expression in BC tissues and cell lines was detected by reverse transcription-quantitative PCR analysis. The Cell Counting Kit-8, proliferation, Transwell and wound healing assays were employed to confirm the effect of LINC00052 on cell proliferation, migration and invasion. The cell localization of LINC00052 was estimated by cytoplasmic nuclear separation assay. Finally, the potential regulatory mechanism of LINC00052 in BC was detected by western blot analysis. The expression levels of LINC00052 were found to be significantly higher in BC tissues compared with those in the adjacent normal tissues. Downregulation of LINC00052 expression in vitro significantly suppressed the proliferation, migration and invasion of BC cells. LINC00052 was mainly expressed in the cytoplasm and was considered to bind with microRNA (miR)-145-5p based on various databases. Notably, the high expression levels of LINC00052 led to the low expression levels of miR-145-5p and high expression levels of TGF-β receptor II (TGFBR2). In conclusion, the findings of the present study demonstrated that LINC00052 may sponge miR-145-5p to upregulate TGFBR2 expression in order to promote the proliferation and metastasis of BC cells. Therefore, LINC00052 may be an effective potential target for the diagnosis and treatment of BC.

Long Noncoding RNA TCONS_00068220 Promotes Breast Cancer Progression by Regulating Epithelial-Mesenchymal Transition Marker E-Cadherin.

BackgroundLong noncoding RNAs (lncRNAs) play essential roles in the regulation of breast cancer development. We herein investigated the potential role of lncRNA TCONS_00068220 in breast cancer pathogenesis.Material/MethodsThe expression levels of TCONS_00068220 in breast cancer tissues were measured by qRT-PCR. Afterwards, TCONS_00068220 was (1) overexpressed in MCF-7 breast cancer cells, and (2) silenced in MDA-MB-231 cells. Then, CCK-8 and transwell assays were conducted to detect the impact of TCONS_00068220 on cell proliferation, migration, and invasion. The expression of the epithelial-mesenchymal transition (EMT) marker E-cadherin was detected by western blot assay after upregulation or downregulation of TCONS_00068220.ResultsTCONS_00068220 was remarkably upregulated in breast cancer tissues compared with non-cancerous tissues. In addition, TCONS_00068220 level was significantly correlated with lymphatic metastasis, Ki67 index, clinical stage, and differentiation grade. All breast cancer cell lines displayed a higher expression level of TCONS_00068220 compared with the normal breast epithelial cell line MCF-10A. Furthermore, enhanced expression of TCONS_00068220 in MCF-7 cells promoted cell proliferation, migration, invasion, and EMT, whereas TCONS_00068220 knockdown in MDA-MB-231 cells led to the opposite results. E-cadherin was negatively regulated by TCONS_00068220 in both breast cancer tissues and cell lines. Finally, TCONS_00068220 regulated MCF-7 and MDA-MB-231 cell behaviors by downregulating E-cadherin.ConclusionsTCONS_00068220 promotes breast cancer cell proliferation, migration, and invasion, while facilitating the process of EMT by interacting with E-cadherin and suppressing its expression. Therefore, it may potentially serve as an oncogene in breast cancer progression.