Breast Cancer Cell Angiogenesis Research Articles

Lipocalin-2 promotes breast cancer brain metastasis by enhancing tumor invasion and modulating brain microenvironment.

Breast cancer is the leading cancer diagnosed in women globally, with brain metastasis emerging as a major cause of death, particularly in human epidermal growth factor receptor 2 positive and triple-negative breast cancer subtypes. Comprehensive understanding of the molecular foundations of central nervous system metastases is imperative for the evolution of efficacious treatment strategies. Lipocalin-2 (LCN2), a secreted iron transport protein with multiple functions, has been linked to the progression of breast cancer brain metastasis (BCBM). In primary tumors, LCN2 promotes the proliferation and angiogenesis of breast cancer cells, triggers the epithelial-mesenchymal transition, interacts with matrix metalloproteinase-9, thereby facilitating the reorganization of the extracellular matrix and enhancing cancer cell invasion and migration. In brain microenvironment, LCN2 undermines the blood-brain barrier and facilitates tumor seeding in the brain by modulating the behavior of key cellular components. In summary, this review meticulously examines the fuel role of LCN2 in BCBM cascade, and investigates the potential mechanisms involved. It highlights the potential of LCN2 as both a therapeutic target and biomarker, indicating that interventions targeting LCN2 may offer improved outcomes for patients afflicted with BCBM.

Nischarin inhibits the epithelial-mesenchymal transition process and angiogenesis in breast cancer cells by inactivating FAK/ERK signaling pathway via EGF like repeats and discoidin domains 3.

Our previous study has demonstrated that Nischarin (NISCH) exerts its antitumor effects in breast cancer (BC) by suppressing cell migration and invasion. This study aims to explore the underlying mechanism through which NISCH functions in BC. The relevance between EGF Like Repeats and Discoidin Domains 3 (EDIL3) mRNA expression and the overall survival of tumor patients was depicted by the Kaplan-Meier curve. The findings revealed that overexpressed NISCH attenuated cell motility and colony-forming capacities of Hs578T cells, yet silenced NISCH in MDA-MB-231 cells led to contrasting results. Western blot (WB) analysis indicated that overexpression of NISCH significantly down-regulated the Vimentin and Slug expression, and inactivated the FAK/ERK signaling pathway. RNA sequencing (RNA-seq) was performed in NISCH-overexpressed Hs578T cells and the control cells to analyze differentially expressed genes (DeGs), and the results showed a significant down-regulation of EDIL3 mRNA level upon overexpression of NISCH. Subsequent functional analyses demonstrated that overexpression of EDIL3 attenuated the inhibitory effect of NISCH on cell migration, invasion, colony formation, and tube formation. In summary, our finding preliminarily revealed that NISCH inhibits the epithelial-mesenchymal transition (EMT) process and angiogenesis in BC cells by down-regulating EDIL3 to inactivate the FAK/ERK signaling pathway, thereby suppressing the progression of BC. Our results hold promise for contributing to the deep understanding of BC pathogenesis and identifying new therapeutic strategies for clinical application.

Nanoformulated 3′-diindolylmethane modulates apoptosis, migration, and angiogenesis in breast cancer cells

BackgroundIt is well-established that specific herbal plants contain natural active ingredients that have demonstrated anti-cancer potential. Therefore, they are considered highly beneficial as a potential adjuvant, alternative or complementary agent in anti-cancer therapy. However, the low chemical stability and limited bioavailability of 3, 3′-Diindolylmethane (DIM), a plant-derived compound used in clinical settings, limit its therapeutic applications. To overcome this challenge, researchers have focused on developing innovative approaches to improve DIM's biological activity, such as utilizing nanoformulations. Here, we investigated the potential benefits of coating DIM nanoparticles (DIM-NPs) with PEG/chitosan in the treatment of breast cancer. Our results demonstrate the molecular mechanism underlying the activity of DIM-NPs, highlighting their potential as an effective therapeutic strategy for breast cancer treatment. MethodsDIM-PLGA-PEG/chitosan NPs were synthesised and characterised using dynamic light scattering (DLS) and evaluated the impact of these NPs on two breast cancer cell models. ResultsDIM-NPs had an average diameter of 102.3 nm and a PDI of 0.182. When treated with DIM-NPs for 48 h, both MCF7 and MDA-MB-231 cells displayed cytotoxicity at a concentration of 6.25 g/mL compared to untreated cells. Furthermore, in MDA-MB-231 cells, treatment with 2.5 μg/mL of DIM-NPs resulted in a significant decrease in cell migration, propagation, and angiogenesis which was further enhanced at 10 μg/mL. In chicken embryos, treatment with 5 μg/mL of DIM-NPs on day 2 led to a significant reduction in angiogenesis. Furthermore, this treatment induced cell death through a regulatory pathway involving the upregulation of Bax and p53, as well as the downregulation of Bcl-2. These results were supported by in-silico analysis of DIM's binding affinity to key proteins involved in this pathway, namely Bax, Bcl-2, and p53. ConclusionOur findings show that DIM-NPs induces apoptosis, inhibit migration, and reduce angiogenesis in breast cancer. However, further research using a preclinical cancer model may be necessary to determine the pharmacokinetics of DIM-NPs and ensure their safety and efficacy in vivo.

Chemotherapy-induced exosomal circBACH1 promotes breast cancer resistance and stemness via miR-217/G3BP2 signaling pathway

BackgroundChemoresistance involves metastasis and aggressiveness of breast cancer (BC). Chemotherapy-elicited exosomes have been reported to be associated with drug resistance and pro-metastatic capacity of BC cells. Non-coding RNAs (ncRNAs) are enriched in exosomes, which participated in generation, progression, and resistance of BC. However, the mechanism underlying the chemoresistance and metastasis in BC cells mediated by the BC-derived exosomal ncRNAs remained to be elucidated.MethodsThe effects of PTX-induced exosomal circBACH1 on BC cell function were assessed using RNA Binding Protein Immunoprecipitation (RIP), dual luciferase reporter gene, tube formation, CCK-8, and Western Blot assays. The circBACH1 and miR-217 expression levels were detected using quantitative real-time PCR (RT-qPCR) and Immunohistochemistry (IHC) assays in BC tissues and precancerous tissues of BC patients.ResultsCircBACH1 expression was increased in paclitaxel-treated BC-derived exosomes (PTX-EXO) and BC tissue. PTX-EXO was shown to promote PTX-resistance and angiogenesis through upregulation circBACH1. Downregulation of circBACH1 improved PTX-sensitiveness by suppressing the cell viability, stemness, migration, and angiogenesis of BC cells. Moreover, we found that miR-217 interacted with circBACH1 and targeted GTPase-activating SH3 domain-binding protein 2 (G3BP2) in BC cells. CircBACH1 combined miR-217 cotransfection suppressed the expression of G3BP2 proteins compared with circBACH1 treatment in MCF-7 cells. In addition, downregulation of G3BP2 suppressed BC cell migration.ConclusionsThese results demonstrated that PTX-induced exosomal circBACH1 promoted stemness and migration of BC cells by sponging miR-217 to upregulate the expression of G3BP2, which provided a new therapeutic target for PTX-resistance and progression of BC via circBACH1/miR-217/G3BP2 axis.

Role of the TPX2/NCOA5 axis in regulating proliferation, migration, invasion and angiogenesis of breast cancer cells.

Breast cancer is a common malignant tumor in women. Increasing evidence has demonstrated that nuclear receptor coactivator 5 (NCOA5) and targeting protein for xenopus kinesin-like protein 2 (TPX2) serve vital roles in the progression of breast cancer. However, to the best of our knowledge, the molecular mechanisms underlying the involvement of TPX2/NCOA5 in the development of breast cancer are not fully understood at present. In the present study, the expression levels of NCOA5 and TPX2 were compared between paired non-tumor and tumor tissues of patients with breast cancer using the TNMplot tool. Expression differences of NCOA5 and TPX2 in human breast epithelial cell lines (MCF10A and MCF12A) and human breast cancer cell lines (MCF7 and T47D) were assessed via reverse transcription-quantitative PCR and western blotting. Additionally, proliferation, migration and invasion of breast cancer cells were determined via Cell Counting Kit-8, would healing and transwell assays. In vitro angiogenesis was determined using a tube formation assay. Furthermore, TPX2 was identified as a high-confidence NCOA5 interactor based on BioPlex network data sets. A co-immunoprecipitation assay was adopted to confirm the interaction between TPX2 and NCOA5. The present study revealed that TPX2 and NCOA5 were highly expressed in breast cancer cells. TPX2 interacted with NCOA5 and there was a positive association between TPX2 and NCOA5 expression. NOCA5 knockdown repressed the proliferation, migration, invasion and in vitro angiogenesis of breast cancer cells. In addition, TPX2 knockdown suppressed the proliferation, migration and invasion of breast cancer cells, and inhibited in vitro angiogenesis, and all of these effects were reversed following NCOA5 overexpression. In conclusion, NCOA5 was a downstream target of TPX2 in enhancing proliferation, migration, invasion and angiogenesis of breast cancer cells.

Circ_0001667 accelerates breast cancer proliferation and angiogenesis through regulating CXCL10 expression by sponging miR-6838-5p.

An increasing number of circular RNAs (circRNAs) have been shown to play an important role in the tumorigenesis of breast cancer. The aim of this study was to investigate the expression and function of circ_0001667 and its potential molecular mechanisms in breast cancer. The expression levels of circ_0001667, miR-6838-5p and CXC chemokine ligand 10 (CXCL10) in breast cancer tissues and cells were detected by quantitative real-time PCR. Cell counting kit-8 assay, EdU assay, flow cytometry, colony formation and tube formation assays were used to detect cell proliferation and angiogenesis. The binding relationship between miR-6838-5p and circ_0001667 or CXCL10 was predicted using the starBase3.0 database and verified by dual-luciferase reporter gene assay, RIP and RNA pulldown. Animal experiments were performed to assess the function of circ_0001667 knockdown on breast cancer tumor growth. Circ_0001667 was highly expressed in breast cancer tissues and cells, and its knockdown inhibited proliferation and angiogenesis of breast cancer cells. Circ_0001667 sponged miR-6838-5p, and inhibition of miR-6838-5p reversed the inhibitory effect of silencing circ_0001667 on proliferation and angiogenesis of breast cancer cells. MiR-6838-5p targeted CXCL10, and overexpression of CXCL10 reverses the effect of miR-6838-5p overexpression on breast cancer cell proliferation and angiogenesis. In addition, circ_0001667 interference also reduced breast cancer tumor growth in vivo. Circ_0001667 is involved in breast cancer cell proliferation and angiogenesis through regulation of the miR-6838-5p/CXCL10 axis.

A20 affects macrophage polarization through the NLRP3 inflammasome signaling pathway and promotes breast cancer progression.

Breast cancer is the most common malignant tumor in females, and the majority of patients succumb to metastasis. The present study aimed to investigate the association between tumor necrosis factor alpha-induced protein 3 (A20), NOD-, LRR- and pyrin domain-containing protein 3 (NLRP3) and tumor-associated macrophage polarization, and their effects on the proliferation and metastasis of breast cancer cells. The expression of A20 in breast cancer cells was analyzed by reverse transcription-quantitative PCR (RT-qPCR) and western blotting. RT-qPCR and western blotting were also used to confirm the transfection efficiency. The viability, clone formation, migration, invasion and angiogenesis of transfected breast cancer cells were detected by Cell Counting Kit-8, colony formation, wound healing, Transwell and tube formation assays, respectively. Activated macrophages, namely M1 and M2 type macrophages, were observed by double staining immunofluorescence. The levels of M1 and M2 macrophage markers were analyzed by qPCR. The expression of angiogenesis-related proteins and NLRP3 inflammasome activation-associated proteins was detected by western blotting. The results revealed that A20 was highly expressed in breast cancer cells. Interference with A20 inhibited the proliferation, invasion, migration and angiogenesis of breast cancer cells, and inhibited the M2-like polarization of macrophages. Interference with A20 promoted the activation of the NLRP3 inflammasome. The NLRP3 inhibitor MCC950 alleviated the effect of interference with A20 to promote macrophage proliferation and recruitment, as well as M2-like polarization. In conclusion, interference with A20 inhibited macrophage proliferation and M2-like polarization through the NLRP3 inflammasome signaling pathway to inhibit breast cancer progression.

Bioinformatics analysis reveals the potential target of rosiglitazone as an antiangiogenic agent for breast cancer therapy

BackgroundSeveral studies have demonstrated the antitumor activity of rosiglitazone (RGZ) in cancer cells, including breast cancer cells. However, the molecular targets of RGZ in the inhibition of angiogenesis in breast cancer cells remain unclear. This study aimed to explore the potential targets of RGZ in inhibiting breast cancer angiogenesis using bioinformatics-based analysis.ResultsVenn diagram analysis revealed 29 TR proteins. KEGG pathway enrichment analysis demonstrated that TR regulated the adipocytokine, AMPK, and PPAR signaling pathways. Oncoprint analysis showed genetic alterations in FABP4 (14%), ADIPOQ (2.9%), PPARG (2.8%), PPARGC1A (1.5%), CD36 (1.7%), and CREBBP (11%) in patients with breast cancer in a TCGA study. The mRNA levels of FABP4, ADIPOQ, PPARG, CD36, and PPARGC1A were significantly lower in patients with breast cancer than in those without breast cancer. Analysis of gene expression using bc-GenExMiner showed that the mRNA levels of FABP, ADIPOQ, PPARG, CD36, PPARGC1A, and CREBBP were significantly lower in basal-like and triple-negative breast cancer (TNBC) cells than in non-basal-like and non-TNBC cells. In general, the protein levels of these genes were low, except for that of CREBBP. Patients with breast cancer who had low mRNA levels of FABP4, ADIPOQ, PPARG, and PPARGC1A had lower overall survival rates than those with high mRNA levels, which was supported by the overall survival related to DNA methylation. Correlation analysis of immune cell infiltration with TR showed a correlation between TR and immune cell infiltration, highlighting the potential of RGZ for immunotherapy.ConclusionThis study explored the potential targets of RGZ as antiangiogenic agents in breast cancer therapy and highlighted FABP4, ADIPOQ, PPARG, PPARGC1A, CD36, and CREBBP as potential targets of RGZ. These findings require further validation to explore the potential of RGZ as an antiangiogenic agent.

A miR-34a-guided, tRNAiMet-derived, piR_019752-like fragment (tRiMetF31) suppresses migration and angiogenesis of breast cancer cells via targeting PFKFB3

Although we recently demonstrated that miR-34a directly targets tRNAiMet precursors via Argonaute 2 (AGO2)-mediated cleavage, consequently attenuating the proliferation of breast cancer cells, whether tRNAiMet fragments derived from this cleavage influence breast tumor angiogenesis remains unknown. Here, using small-RNA-Seq, we identified a tRNAiMet-derived, piR_019752-like 31-nt fragment tRiMetF31 in breast cancer cells expressing miR-34a. Bioinformatic analysis predicted 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3) as a potential target of tRiMrtF31, which was validated by luciferase assay. tRiMetF31 was downregulated, whereas PFKFB3 was overexpressed in cancer cell lines. Overexpression of tRiMetF31 profoundly inhibited the migration and angiogenesis of two breast cancer cell lines while slightly inducing apoptosis. Conversely, knockdown of tRiMetF31 restored PFKFB3-driven angiogenesis. miR-34a was downregulated, whereas tRNAiMet and PFKFB3 were upregulated in breast cancer, and elevated PFKFB3 significantly correlated with metastasis. Our findings demonstrate that tRiMetF31 profoundly suppresses angiogenesis by silencing PFKFB3, presenting a novel target for therapeutic intervention in breast cancer.

LncRNA LINC01857 reduces metastasis and angiogenesis in breast cancer cells via regulating miR-2052/CENPQ axis.

Long non-coding RNAs have been confirmed closely related to the metastasis and angiogenesis of breast cancer (BC). LINC01857 can promote the growth and metastasis of BC cells. The present work focused on exploring the role of LINC01857 in BC metastasis and angiogenesis and investigating the possible mechanisms. The results showed that LINC01857 and CENPQ were highly expressed in BC tissues and cells, while miR-2052 was contrarily expressed. In vitro study showed that low expression of linc01857 could inhibit the migration ability and vascularization of BC cells, and mir-2052 inhibitor partially restored the effect of si-LINC01857 on the migration ability and vascularization of BC cells. Likewise, inhibition of CENPQ can partially rescue the effects of miR-2052 inhibitor on the migration ability and vascularization of BC cells. In vivo studies showed that down-regulation of LINC01857 notably suppressed tumor growth and angiogenesis in nude mice. The miR-2052 inhibitor partially restored the effects of si-LINC01857. CENPQ suppression partially rescued the effects of the miR-2052 inhibitor. To conclude, LINC01857/miR-2052/CENPQ is the potential novel target for BC treatment.

A fungicide, fenhexamid, is involved in the migration and angiogenesis in breast cancer cells expressing estrogen receptors

Fenhexamid (Fen) is used to eradicate gray mold of fruits and vegetables leading to greater detection of its residual concentration in wine than other fungicides. Here, we further investigated the malign influence of Fen on the migration and angiogenesis via regulation of the estrogen receptor (ER) and phosphoinositide 3-kinase (PI3K) pathways in breast cancer models. ER-positive MCF-7 and ER-negative MDA-MB-231 breast cancer cells were exposed to 17β-estradiol (E2, 10−9 M), Fen (10−5 M and 10−7 M), ICI 182,780 (ICI; an ER antagonist, 10−8 M) or/and Pictilisib (Pic; a PI3K inhibitor, 10−7 M), and subsequently subjected to migration assay, live cell motility monitoring, trans-chamber assay, immunofluorescence, angiogenesis assay, tumor spheroid formation, and Western blot analysis. In MCF-7 cells, E2 and Fen induced cell migration by regulating the cell migration-related proteins. Although expressions of N-cadherin and Vimentin remained unchanged E2 and Fen induced the decrease of E-cadherin and Occludin in the immunofluorescence assay and Western blot analysis. In addition, Fen increased vessel formation in HUVEC cells. Furthermore, Fen treatment induced the formation of larger and denser tumor spheroids in MCF-7 cells. Western blot further confirmed the increased expressions of vascular endothelial growth factor (VEGF) and sex-determining region Y-box 2 (SOX2) after exposure to Fen. We conclude that Fen plays an important role as an endocrine-disrupting chemical in breast cancer migration and metastasis through the regulation of ER and PI3K signaling pathways.

Stabilization of Notch1 and β-catenin in response to ER- breast cancer-specific up-regulation of PSAT1 mediates distant metastasis

Breast cancer has the highest incidence in women worldwide, with a mortality rate second only to lung cancer. Distant metastasis is the major cause of breast cancer-induced death. While upregulation of phosphoserine aminotransferase 1 (PSAT1) has been reported in several cancer types, its specific roles in breast cancer and potential involvement in distant metastasis remain unclear. In our study, PSAT1 was upregulated in metastatic breast cancer and promoted distant metastasis both in vitro and in vivo. Data obtained from transwell and wound healing, colony, sphere assays and detection of various malignant phenotypic markers showed that PSAT1 mediates distant metastasis by promoting invasion, migration, proliferation, anti-apoptosis, stemness and angiogenesis in breast cancer cells. Mechanistically, PSAT1 activated Notch and β-catenin signaling pathways, leading to enhanced distant metastasis. The clinical relevance of PSAT1 in breast cancer was additionally investigated, which revealed associations of poorer patient prognosis with high PSAT1 mRNA and protein expression. In summary, PSAT1 is a critical molecular regulator of distant metastasis that may effectively serve as a marker of poor prognosis in breast cancer.

Endoglin Modulates TGFβR2 Induced VEGF and Proinflammatory Cytokine Axis Mediated Angiogenesis in Prolonged DEHP-Exposed Breast Cancer Cells.

Angiogenesis is the process of vascular network development and plays a crucial role in cancer growth, progression, and metastasis. Phthalates are a class of environmental pollutants that have detrimental effects on human health and are reported to increase cancer risk. However, the interplay between phthalate exposure and angiogenesis has not been investigated thoroughly. In this study, we investigated the effect of prolonged di (2-ethylhexyl) phthalate (DEHP) treatment on the angiogenic potential of triple-negative breast cancer. MDA-MB-231 cells were exposed to physiological concentrations of DEHP for more than three months. Prolonged DEHP exposure induced angiogenesis in breast cancer cells. Endoglin (ENG)/CD105 is a membrane glycoprotein and an auxiliary receptor of the TGFβ receptor complex. In endothelial cells, ENG is highly expressed and it is a prerequisite for developmental angiogenesis. A literature review highlights endoglin as a well-known mesenchymal stem cell marker responsible for vascular development and angiogenesis. NGS analysis showed that endoglin overexpression in DEHP-exposed MDA-MB-231 cells correlated with tumor development and growth. An in vivo zebrafish xenograft assay showed that VEGFA induced sprouting of the subintestinal vein (SIV) in embryos injected with DEHP-exposed cells. Endoglin knockdown reduced SIV sprouting and VEGFA expression in zebrafish embryos. An in vitro HUVEC tube formation assay showed that endoglin depletion reversed DEHP-induced VEGF-mediated HUVEC tube formation in coculture. DEHP-induced endoglin activated TGFβ/SMAD3/VEGF and MAPK/p38 signaling in MDA-MB-231 cells. A cytokine angiogenesis antibody array showed induced expression of the inflammatory cytokines IL1α, IL1β, IL6, and IL8, along with GMCSF and VEGF. Endoglin knockdown reversed DEHP-induced activation of the TGFβ/SMAD3/VEGF signaling axis, MAPK/p38 signaling, and cytokine regulation, limiting angiogenesis potential both in vivo and in vitro. Targeting endoglin might serve as a potential alternative treatment to control angiogenesis, leading to metastasis and limiting cancer progression.

Sophoridine Inhibits Breast Cancer Cell Migration and Angiogenesis by Regulating Vascular Endothelial Growth Factor Receptor 2/Extracellular Signal-Regulated Kinase 2 Pathways

Breast cancer is the most common form of cancer in women in need for better treatment. Sophoridine, a plant-derived quinoxidine alkaloid, has been shown to exhibit pharmacological effects on several diseases including lung and gastric cancers. To further extend these beneficial effects we have explored the role of sophoridine in breast cancer progression. Our results show that sophoridine suppressed the proliferation of breast cancer cells and inhibited the migration as well as invasion. In addition, sophoridine inhibited the angiogenesis of breast cancer cells. Mechanically, our data confirmed that sophoridine suppressed the vascular endothelial growth factor receptor 2/extracellular signal-regulated kinase 2 pathway, and therefore suppressed breast cancer progression. We therefore suggest that sophoridine could serve as a promising drug for the treatment of breast cancer.

A Fungicide, Fenhexamid, is Involved in the Migration and Angiogenesis in Breast Cancer Cells Expressing Estrogen Receptors

A Fungicide, Fenhexamid, is Involved in the Migration and Angiogenesis in Breast Cancer Cells Expressing Estrogen Receptors

Nanoencapsulation of Hirudo medicinalis proteins in liposomes as a nanocarrier for inhibiting angiogenesis through targeting VEGFA in the Breast cancer cell line (MCF-7).

Introduction: Breast cancer is the most serious cause of women’s death throughout the world. Using nanocarrier vehicles to the exact site of cancer upgrades the therapeutic efficiency of the drugs. Capsulation of active proteins in the vesicular liposomes’ hydrophilic core is essential to develop a therapeutic protein carrier system. We aimed to encapsulate the medicinal leech saliva extract (LSE) and assess the inhibition of angiogenesis of breast cancer cells by targeting vascular endothelial growth factor A (VEGFA). Methods: In this research, enhanced formulation of liposomal protein was determined by zeta potential analysis, droplet size, drug release assay, and transmission electron microscopy (TEM). Furthermore, a cytotoxicity assay of liposomal LSE was performed to determine the cytotoxic activity of components. For assessing the expression of VEGFA, P53, and hypoxia-inducible factor subunit alpha (HIF1a) genes, Real-Time PCR was applied. Results: Nano liposome was chosen as an enhanced formulation due to its much smaller size (46.23 nm). Liposomal LSE had more practical actions on the MCF-7 cells. As noticed by DAPI staining, apoptosis was extensively greater in treated MCF-7 cells. Wound healing assay demonstrated that MCF-7 cells could not sustain growth at the presence of liposomal LSE and expression of the VEGFA gene was declined in treated cells. Downregulation of VEGFA was evaluated with western blotting technique. Conclusion: It can be concluded that our investigation of the tests confirmed the fact that nano liposomal LSE is a novel promising formulation for anticancer drugs and can significantly improve the penetration of protein drugs to cancer cells.

Inhibitory effect of flavonoid xanthomicrol on triple-negative breast tumor via regulation of cancer-associated microRNAs.

Breast cancer is the leading cause of cancer death in women worldwide. Due to the side effects of current chemo-reagents on healthy tissues, it is essential to search for alternative compounds with less toxicity and better efficacy. In the present study, we have investigated the anticancer effects of flavonoid xanthomicrol on the mice breast cancer model using MTT assay, cell cycle and Annexin/PI analysis, colony formation assay, H&E staining, immunohistochemistry, and miRNA analysis. Our results demonstrated that xanthomicrol decreased the cell viability and clonogenic capability, induced G1-arrest and apoptosis in the breast cancer cells in vitro, and caused a significant reduction in the volume and weight of mice tumors in vivo. In addition, xanthomicrol reduced the expression of TNFα, VEGF, MMP9, and Ki67, while upregulating the expression of apoptotic markers such as Bax, caspase3, and caspase9. Finally, the expression of miR21, miR27, and miR125, known as oncomirs, decreased significantly after xanthomicrol administration, while the expression of miR29 and miR34, functioning as tumor suppressors, increased significantly (p < .001). Our data demonstrated that xanthomicrol can induce apoptosis and suppress angiogenesis in breast cancer cells due to its inhibitory effect on oncomirs and its stimulatory effect on tumor suppressor miRNAs.

MiR-183-5p Promotes Proliferation, Metastasis and Angiogenesis in Breast Cancer Cells through Negatively Regulating Four and a Half LIM Protein 1.

PurposeFour and a half LIM protein 1 (FHL1) is involved in breast cancer (BC) development, but the regulatory mechanism involved remain unclear. In the present study, we examined the role of FHL1 in BC development.MethodsThe expression of FHL1, miR-183-5p, and miR-96-5p in BC tissues was analyzed using StarBase analysis. FHL1 expression in BC tissues, a normal human breast epithelial cell line, and BC cell lines was detected using quantitative reverse transcription polymerase chain reaction (qRT-PCR). The relationship between FHL1 and miR-183-5p/miR-96-5p was analyzed via Pearson's rank correlation, TargetScan, and a dual-luciferase reporter assay. BT549 and MDA-MB-231 cells were transfected with either FHL1 and miR-183-5p mimics, or siFHL1 and a miR-183-5p inhibitor, respectively. The viability, colony number, migration, invasion, and tube length of BT549 and MDA-MB-231 cells were examined using cell counting kit-8, colony formation, wound-healing, Transwell, and tube formation assays, respectively. The levels of FHL1, vascular endothelial growth factor (VEGF), p53, E-cadherin, N-cadherin, and vimentin were quantified using western blotting and qRT-PCR.ResultsFHL1 expression was downregulated in BC tissues and cells, whereas miR-183-5p and miR-96-5p were upregulated in BC tissues (negative correlation with FHL1 expression). FHL1 overexpression inhibited the viability, colony number, migration, and invasion of BC cells and the expression of VEGF, N-cadherin, and vimentin, and increased the expression of FHL1, p53, and E-cadherin in BT549 cells. Furthermore, a miR-183-5p mimic reversed these effects of FHL1 overexpression, whereas FHL1 silencing caused opposite results to those observed in MDA-MB-231 cells; however, this was reversed by a miR-183-5p inhibitor.ConclusionOur study suggests that miR-183-5p promotes cell proliferation, metastasis, and angiogenesis by negatively regulating FHL1 in BC.

IN-VITRO AND IN-VIVO STUDIES OF THE POSSIBLE CHEMOPREVENTIVE FUNCTIONS OF SOYBEAN ISOFLAVONE AND FLAVONOIDS ON BREAST CANCER

Objectives: In this study, we assess the possible influence of soybean isoflavone (genistein) and other flavonoids (quercetin and catechin) on breast cancer chemoprevention. We design in-vitro and in-vivo experiments to analyze the effect of genistein, quercetin and catechin on cell proliferation, cell migration, and angiogenesis of breast cancer cells. Methods: In cell proliferation experiment, MCF-7 cells, SKBR-3 cells, and HUVEC cells were treated with genistein and other flavonoids (catechin and/or quercetin) for 48[Formula: see text]h to assess the influence on cell growth of normal and breast cancer cells. In cell motility test, we analyze the effect of isoflavone and flavonoids on migration ability of MCF-7 cells by 16[Formula: see text]h and SKBR-3 cells by 24[Formula: see text]h in two different concentrations (1.25[Formula: see text][Formula: see text]g/ml and 2.5[Formula: see text][Formula: see text]g/ml). In the in-vivo experiment, SKBR-3 cells mixed with PBS and catechin, respectively, were injected subcutaneously into nude mice, then we investigated the effect of catechin on cell growth by observing subcutaneous tumor size changes after 15 days. Results: The results suggest that genistein and quercetin can significantly inhibit proliferation of breast cancer cells, and their inhibitory effects are independent of estrogen receptor. In cell motility tests, all of the three phytochemicals were effective in the inhibition of cell migration on two breast cancer cell lines, except for quercetin on cell migration of SKBR-3 cell line. In the in-vitro experiment, catechin showed stimulatory effect on cell proliferation of HUVEC cell line, which may consider positive effect on angiogenesis, rather than inhibitory effect. However, in the in-vivo experiment, it showed no significant change in tumor size between the groups of with and without catechin treatment. Conclusions: According to our study, the results suggest that isoflavone and flavonoids tend to inhibit cell growth and metastasis of breast cancer cells. Our in-vivo experiment does not reach a significant result, and it may be due to lower catechin concentration. Under in-vivo environment, we should also consider the possible metabolic forms of catechin that cause different result from the in-vitro study.

Rhizoma Pleionis inhibits of breast cancer epithelial-mesenchymal transition by VEGF-A related angiogenesis

Objective To investigate the Rhizoma Pleionis mediating vascular endothelial growth factor A (VEGF-A)-related angiogenesis in breast cancer cells to inhibit epithelial-mesenchymal transition (EMT) in breast cancer. Methods Firstly, the proliferation activity of MDA-MB-231 and 4T1 of breast cancer cells was detected by methyl thiazolyl tetrazolium (MTT) method at different concentrations (0.2 g/ml, 0.4 g/ml, 0.6 g/ml, 0.8 g/ml, 1.0 g/ml); secondly, the specific concentration of Rhizoma Pleionis was selected and tested by Transwell test for its effect on breast cancer cell metastasis and invasion. The formation of human umbilical vein endothelial cells (HUVEC) capillary wall was measured to detect the effect of Rhizoma Pleionis on cell angiogenesis. The EMT-related protein and angiogenesis-related molecules were detected by Western blot. Results MTT results showed that Rhizoma Pleionis of 0.8 g/ml and 1.0 g/ml can inhibit the MDA-MB-231 and 4T1 cells proliferation in a dose-dependent manner, with statistically significant difference (P<0.05). Rhizoma Pleionis can inhibit the invasion of breast cancer cells and inhibit the formation of blood vessel walls in HUVECs. Western blot confirmed that the expression level of EMT-related proteins: E-cadherin was significantly increased and the expression level of N-cadherin, Vimentin and Snail was significantly decreased after treatment with Rhizoma Pleionis (P<0.05); and the level of the related molecule hypoxia inducible factor-1α (HIF-1α), VEGF-A and vascular endothelial growth factor receptor-2 (VEGFR-2) was significantly reduced (P<0.05). Conclusions Rhizoma Pleionis inhibits the epithelial-mesenchymal transition of breast cancer by influencing the angiogenesis of VEGF-A, and inhibits the invasion and metastasis of breast cancer. Key words: CREMASTRA VARIABILIS; Vascular endothelial growth factor A; Breast neoplasms; Tumor cells, cultured; Epithelial-mesenchymal transition