Migration Of Human Breast Cancer Cells Research Articles

Caging Bioactive Triarylimidazoles: An Approach to Create Visible Light-Activatable Drugs.

Imidazoles are crucial structural components in a variety of small-molecule inhibitors designed to target different kinases in anticancer treatment. However, the effectiveness of such inhibitors is often hampered by nonspecific effects and the development of resistance. Photopharmacology provides a compelling solution by enabling external control over drug activity with spatiotemporal precision. Herein, we introduce a novel strategy for caging bioactive triarylimidazole-based drug molecules. This approach involves introducing a dialkylamino group as a photoremovable group on the carbon atom of the imidazole ring, which intrinsically modulates the core structure from planar imidazole to tetrahedral 2H-imidazole, enabling the caged compound to be selectively uncaged upon visible light exposure. We applied this innovative caging technique to SB431542, a triarylimidazole-based small-molecule inhibitor that targets the pivotal TGF-β signaling pathway, the dysregulation of which is linked to several human diseases, including cancer. Our results demonstrated the selective inhibition of human breast cancer cell migration in vitro upon light activation, highlighting the potential of our approach to transform triarylimidazole-based drug molecules into visible light-activatable drugs, thereby facilitating spatiotemporal regulation of their pharmacological activity.

MitoQ Inhibits Human Breast Cancer Cell Migration, Invasion and Clonogenicity.

Simple SummarySolid tumors comprise metabolically hostile areas adjacent to metabolically friendly ones, and their distribution fluctuates in space and time. Selection pressure is maximal in hostile areas where cancer cells develop at least four survival strategies: to adapt themselves, to modify the microenvironment, to hibernate metabolically, and to escape. Escape marks the transition from a localized to an invasive tumor that may ultimately disseminate remotely, forming metastases. Based on the hypothesis that mitochondria are metabolic sensors that control these responses, we previously established that mitochondrial superoxide is a pro-metastatic intracellular signaling agent. Here, we tested MitoQ, a mitochondria-targeted ROS inactivator that already successfully passed Phase I safety clinical trials, as a potential inhibitor of the early steps of the metastatic cascade. Using human breast cancer cells as models in anticipation of preclinical and clinical assays, we report that MitoQ inhibits cancer cell migration, invasion, clonogenicity, sphere formation and spheroid stability.To successfully generate distant metastases, metastatic progenitor cells must simultaneously possess mesenchymal characteristics, resist to anoïkis, migrate and invade directionally, resist to redox and shear stresses in the systemic circulation, and possess stem cell characteristics. These cells primarily originate from metabolically hostile areas of the primary tumor, where oxygen and nutrient deprivation, together with metabolic waste accumulation, exert a strong selection pressure promoting evasion. Here, we followed the hypothesis according to which metastasis as a whole implies the existence of metabolic sensors. Among others, mitochondria are singled out as a major source of superoxide that supports the metastatic phenotype. Molecularly, stressed cancer cells increase mitochondrial superoxide production, which activates the transforming growth factor-β pathway through src directly within mitochondria, ultimately activating focal adhesion kinase Pyk2. The existence of mitochondria-targeted antioxidants constitutes an opportunity to interfere with the metastatic process. Here, using aggressive triple-negative and HER2-positive human breast cancer cell lines as models, we report that MitoQ inhibits all the metastatic traits that we tested in vitro. Compared to other mitochondria-targeted antioxidants, MitoQ already successfully passed Phase I safety clinical trials, which provides an important incentive for future preclinical and clinical evaluations of this drug for the prevention of breast cancer metastasis.

Abstract PS17-55: Reactive oxygen species scavengers in triple negative breast cancer

Abstract Reactive oxygen species (ROS) are well known to play important roles in cancer. In particular, ROS of mitochondrial origin (mtROS) promote DNA mutations and cell death when produced at high levels, but rather stimulate autophagy and metastatic spread when produced at subcytotoxic levels [1]. Multiple metabolic pathways and oncogenic regulators linked to ROS have been studied in cancer cells, including MAPK/ERK, PI3K/AKT, PTEN, PKD, BREF2, HIF-1, NF-κB and p53 [2]. Abnormalities in these pathways can contribute to tumor development.By producing mtROS, mitochondria control and could even trigger metastatic spread, meaning that the use of specific mitochondria-targeted superoxide inhibitors/scavengers, such as mitoquinol mesylate (MitoQ), could reduce and/or prevent metastatic dissemination. This has been studied and reported with the use of MitoTempo, which prevents metastasis of naturally metastatic MDA-MB-231 human breast cancer cells implanted in mice [1]. Here, we show that MitoQ impairs MDA-MB-231 and SkBr3 human breast cancer cell migration and invasion in vitro, and MDA-MB-231 metastasis to the lungs in mice in vivo. In metastatic take assays, tail vein-injected MDA-MB-231 cells pretreated with MitoQ formed significantly less metastases after 4 weeks compared to vehicle. In spontaneous metastatic assays, mice bearing orthotopic MDA-MB-231 tumors were treated with ± 20 mg/kg MitoQ (per os) for 6 weeks. Primary tumors were surgically removed to allow metastatic outgrowth. In this model, MitoQ decreased primary tumor recurrence as well as the number of lung micro-metastases. Additionally, RNAseq studies in two human breast cancer cell lines identified that MitoQ significantly modulated a common set of metabolic genes in vitro and in vivo. They can be used as biomarkers of response. These results, together with a successful Phase I clinical trial and the fact that MitoQ does not interfere with the cytotoxic effects of chemotherapy, confirm that MitoQ is a potential candidate to be clinically tested in triple negative breast cancer patients.

Dieckol or phlorofucofuroeckol extracted from Ecklonia cava suppresses lipopolysaccharide-mediated human breast cancer cell migration and invasion

The phlorotannin compounds, dieckol and phlorofucofuroeckol (PFF), extracted from Ecklonia cava, have been suggested to exhibit anti-inflammatory, anti-adipogenic or anti-tumorigenic activity. However, role of dieckol or PFF in LPS/TLR-4 signaling that regulates cancer development still remains to be determined. Here we report that dieckol or PFF blocked lipopolysaccharide (LPS)-induced migration and invasion of human breast cancer MCF-7 or MDA-MB-231 cells by about 50% (p < 0.0001) or 60% (p = 0.0001), respectively. Also, PFF as well as dieckol markedly attenuated both of LPS-induced Toll-like receptor4 (TLR-4) expression and NF-κB promoter-driven transcriptional activity, which are essential for the migration and invasion. Moreover, LPS induction of NF-κB-dependent gene, matrix metalloproteinase9 (MMP-9) was markedly decreased about 2.6 or 2.5 fold by dieckol and PFF, respectively. In addition, LPS-induced MMP-2 gene expression was reduced about 1.7 or 2.1 fold by dieckol and PFF, respectively. Significantly, each inhibitor of MMP-9 or NF-κB notably reduced invasion of MDA-MB-231 cells in response to LPS by about 70% (p = 0.0115) or 40% (p < 0.0001), respectively. Collectively, these findings demonstrate that both of dieckol and PFF functions as a critical inhibitor of LPS-induced breast cancer cell invasion through downregulation of TLR-4-NF-κB-MMP-9 signaling axis, suggesting a potential role of the phlorotannins in inflammation-mediated breast cancer development.

MiR‑27a promotes human breast cancer cell migration by inducing EMT in a FBXW7‑dependent manner.

Increasingly, evidence has revealed that aberrant microRNA (miRNA) expression is involved in breast cancer carcinogenesis and further progression, including metastasis. miRNA (miR)-27a was previously identified to be abnormally expressed and to serve pro-oncogenic functions in multiple human cancer types, including breast cancer. However, its functions and underlying mechanisms in breast cancer remain poorly understood. In the present study, it was demonstrated that miR-27a was significantly upregulated in breast cancer tissues and cell lines compared with their normal counterparts. Overexpression of miR-27a resulted in enhanced cell migration by inducing epithelial-to-mesenchymal transition, while its knockdown effectively reversed these cellular events. The present study additionally confirmed for the first time, to the best of our knowledge, that F-box and WD repeat domain containing 7 (FBXW7) is a downstream target gene of miR-27a in human breast cancer cells. FBXW7 is underexpressed in breast cancer tissues and cell lines, and is an independent positive factor for the overall survival rate of patients with breast cancer. Notably, the ectopic expression of FBXW7 may effectively suppress the epithelial-to-mesenchymal transition and migratory activity of breast cancer cells, in addition to reversing the cell migration mediated by miR-27a. Altogether, the results of the present study indicated the important function of miR-27a in regulating the metastasis of breast cancer in a FBXW7-dependent manner, and provide evidence for the potential application of miR-27a in breast cancer therapy.

Lidocaine inhibits cytoskeletal remodelling and human breast cancer cell migration

BackgroundThe metastatic potential of breast cancer cells has been strongly associated with overexpression of the chemokine CXCL12 and the activity of its receptor CXCR4. Lidocaine, a local anaesthetic that can be used during breast cancer excision, inhibits the growth, invasion, and migration of cancer cells. We therefore investigated, in a breast cancer cell line, whether lidocaine can modulate CXCL12-induced responses. MethodsIntracellular calcium, cytoskeleton remodelling, and cell migration were assessed in vitro in MDA-MB-231 cells, a human breast cancer epithelial cell line, after exposure to lidocaine (10 μM or 100 μM). ResultsLidocaine (10 or 100 μM) significantly inhibited CXCR4 signalling, resulting in reduced calcium release (Fluo 340 nm/380 nm, 0.76 mean difference, p<0.0001), impaired cytoskeleton remodelling (F-Actin fluorescence mean intensity, 21 mean difference, P=0.002), and decreased motility of cancer cells, both in the scratch wound assay (wound area at 21 h, −19%, P<0.0001), and in chemotaxis experiments (fluorescence mean intensity, 0.16, P=0.0047). The effect of lidocaine was not associated with modulation of the CD44 adhesion molecule. ConclusionsAt clinical concentrations, lidocaine significantly inhibits CXCR4 signalling. The results presented shed new insights on the molecular mechanisms governing the inhibitory effect of lidocaine on cell migration.

BCSC-1 suppresses human breast cancer metastasis by inhibiting NF-κB signaling.

Breast cancer suppressor candidate-1 (BCSC-1; also termed von Willebrand factorA domain containing5A and LOH11CR2A) is a newly identified candidate tumor suppressor gene that has been implicated in several types of cancer in previous studies. However, there have been few reports about the association between BCSC-1 and human breast cancer in recent years. In the present study, the expression of BCSC-1 in breast cancer was determined by immunohistochemistry(IHC) staining of tissue microarrays and clinical tissue specimens. Subsequently, BCSC-1 gene expression was evaluated in different breast cancer cell lines by quantitative polymerase chain reaction and the MDA-MB-231 cell line was selected for further use in subsequent experiments, due to its low BCSC-1 expression. An MDA-MB-231 cell line with stable overexpression of BCSC-1 was established through transfection with plasmid containing the BCSC-1 gene, and then screening for G418 resistance. Wound-healing, migration and invasion assays were conducted to detect the effect of BCSC-1 on MDA-MB-231 cells. Furthermore, changes in matrix metalloproteinases (MMPs), osteopontin(OPN) and the nuclear factor-κB (NF-κB) pathway were detected in the current study. Additionally, stable silencing of BCSC-1 expression in MCF-7 cells was performed using a lentivirus. The results of IHC indicated that BCSC-1 is expressed at low levels in breast cancer tissues compared with in normal breast tissue. Results of the wound healing, migration and invasion assays demonstrated that BCSC-1 overexpression reduced the metastasis ability of MDA-MB-231 cells invitro. Further research confirmed that the BCSC-1 overexpression reduced the expression levels of MMP7, MMP9 and OPN, and the phosphorylation of NF-κB p65. Furthermore, inhibition of BCSC-1 via lentivirus-mediated RNA interference revealed that the downregulation of BCSC-1 increased the invasive ability of MCF-7 cells. In summary, the results demonstrated that BCSC-1 is expressed at low levels in breast cancer tissues, and that it can suppress human breast cancer cell migration and invasion, potentially altering the expression of MMP7, MMP9, OPN, and the activity of the NF-κB pathway. Therefore, BCSC-1 may be useful as a biomarker for the treatment of breast cancer in the future.

Inhibitory effect of interleukin -11 receptor α-targeted 131I-CGRRAGGSC on human breast cancer cells

Objective To establish interleukin -11 receptor α (IL-11Rα)-based 131I-CGRRAGGSC by chloramines-T method, and explore the inhibitory effects of 131I-CGRRAGGSC on human breast cancer cells overexpressing IL-11Rα. Methods The tyrosine modified CGRRAGGSC protein was labeled with 131I by chloramines-T method. Methyl thiazol tetrazolium (MTT), Transwell and wound-healing assay were used to detect the effects of 131I-CGRRAGGSC on the migration and proliferation of human breast cancer cells. Western blotting was used to detect the protein expression of IL-11Rα, phosphorylated signal transducer and activators of transcription 3 (p-STAT3) and signal transducer and activators of transcription 3 (STAT3) in breast cancer cells after treatment with 0, 2.775 and 5.550 MBq/ml 131I-CGRRAGGSC. Results 131I-CGRRAGGSC was radiolabeled successfully and radiochemical purity after purification was over 95%. The inhibition ratio of breast cancer cells (MDA-MB-231 and MCF-7) was (21.46±1.08)%, (37.95±2.59)%, (49.18±0.77)%, (60.34±0.19)% (F=389.4, P=0.000) and (27.59±2.15)%, (41.67±0.92)%, (54.53±1.07)%, (64.84±1.69)% (F=328.6, P=0.000) after 131I-CGRRAGGSC treatment (4.625, 9.250, 18.500 and 37.000 MBq/ml, respectively). Transwell demonstrated that the migration ratios of 2.775, 5.55 MBq/ml 131I-CGRRAGGSC were (32.05±4.44)%, (45.92±2.55)% (t=4.688, P=0.009) for MDA-MB-231 cells and (21.99±1.18)%, (39.45±1.36)% (t=16.760, P=0.000) for MCF-7 cells, respectively. Wound-healing assay also suggested that 131I-CGRRAGGSC inhibited MDA-MB-231 cells, but not MCF-7 cells. The cellular IL-11Rα and p-STAT3 levels were decreased in a dose-dependent manner whereas the STAT3 level had no significant change in 131I-CGRRAGGSC-treated breast cancer cells after 24 h in compared to those in the control cells. Conclusion 131I-CGRRAGGSC could inhibit the migration and proliferation of human breast cancer cells by down-regulating the IL-11Rα to inhibit STAT3 activation. Key words: Breast cancer; Radiolabeling; Migration; 131I

The LPI/GPR55 axis enhances human breast cancer cell migration via HBXIP and p-MLC signaling.

The G protein-coupled receptor 55 (GPR55) is expressed in multiple tissues, and has been implicated in cancer pathogenesis, but little is known about its role in the migratory behavior of cancer cells, particularly breast cancer cells. In this study we first showed that GPR55 expression levels in 38 metastatic lymph nodes of breast cancer patients were profoundly elevated, and were positively associated in human breast cancer cells with their migratory ability. Moreover, the plasma levels of GPR55 endogenous agonist L-a-lysophosphatidylinositol (LPI) were significantly increased in breast cancer patients compared with healthy individuals. In human breast cancer LM-MCF-7 and MDA-MB-231 cells, treatment with LPI (2.5 μmol/L) significantly increased filopodia formation and resulted in cell migration, which could be blocked either by the GPR55 antagonist CID16020046 or by siRNA-mediated GPR55 knockdown. Furthermore, dual-luciferase report gene assays showed that GPR55 upregulated HBXIP at the promoter; GPR55 expression levels were positively correlated with HBXIP expression levels in breast cancer tissues and 8 breast cancer cell lines. We also showed that the LPI/GPR55 axis promoted the migration of breast cancer cells via two mutually exclusive pathways - the HBXIP/p-ERK1/2/Capn4 and MLCK/MLC signaling pathways. In xenograft nude mouse model, loss of GPR55 mainly affected breast cancer cell metastasis and the formation of metastatic foci. Thus, GPR55 is involved in the migratory behavior of human breast cancer cells and could serve as a pharmacological target for preventing metastasis.

Macrophage-derived interleukin-1beta promotes human breast cancer cell migration and lymphatic adhesion in vitro

Lymphovascular invasion (LVI), encompassing blood and lymphatic vessel invasion, is an important event in tumourigenesis. Macrophages within the tumour microenvironment are linked to the presence of LVI and angiogenesis. This study investigates the role of macrophage-derived, caspase-1-dependent interleukin-1beta (IL-1β) in an in vitro model of LVI. IL-1β significantly augmented the adhesion and transmigration of breast cancer cell lines MCF7 and MDA-MB-231 across endothelial cell barriers. MDA-MB-231 and MCF7 showed a higher percentage of adhesion to lymphatic endothelial cells than blood endothelial cells following endothelial cell IL-1β stimulation (P < 0.001 and P < 0.0001, respectively). Supernatants from activated macrophages increased the adhesion of tumour cells to lymphatic and blood endothelium. Secretion of IL-1β was caspase-1 dependent, and treatment with caspase-1 inhibitor reduced IL-1β production by 73% and concomitantly reduced tumour cell adhesion to levels obtained with resting macrophages. Transmigration of MDA-MB-231 cells across blood and lymphatic endothelial monolayers was significantly increased following IL-1β stimulation. Furthermore, supernatants from activated macrophages increased transmigration of MDA-MB-231 cells across endothelial monolayers, which was abolished by caspase-1 inhibition. IL-1β stimulation of tumour cells significantly increased their migratory ability and a significant increase in migration was observed when MDA-MB-231 cells were stimulated with macrophage conditioned media (two of three donors). Results demonstrate that macrophage production of IL-1β plays an important role in the migration of breast cancer cells and their adhesion to, and transmigration across, blood and lymphatic endothelial cells. Results suggest that IL-1β may play a role in the adhesion to lymphatic endothelial cells in particular.

Inhibitory Effects on Human Breast Cancer Cells Migration of Small Black Bean according to the Cooking Methods

Inhibitory Effects on Human Breast Cancer Cells Migration of Small Black Bean according to the Cooking Methods

Hexachlorobenzene modulates the crosstalk between the aryl hydrocarbon receptor and transforming growth factor-β1 signaling, enhancing human breast cancer cell migration and invasion.

Given the number of women affected by breast cancer, considerable interest has been raised in understanding the relationships between environmental chemicals and disease onset. Hexachlorobenzene (HCB) is a dioxin-like compound that is widely distributed in the environment and is a weak ligand of the aryl hydrocarbon receptor (AhR). We previously demonstrated that HCB acts as an endocrine disruptor capable of stimulating cell proliferation, migration, invasion, and metastasis in different breast cancer models. In addition, increasing evidence indicates that transforming growth factor-β1 (TGF-β1) can contribute to tumor maintenance and progression. In this context, this work investigated the effect of HCB (0.005, 0.05, 0.5, and 5μM) on TGF-β1 signaling and AhR/TGF-β1 crosstalk in the human breast cancer cell line MDA-MB-231 and analyzed whether TGF-β1 pathways are involved in HCB-induced cell migration and invasion. RT-qPCR results indicated that HCB reduces AhR mRNA expression through TGF-β1 signaling but enhances TGF-β1 mRNA levels involving AhR signaling. Western blot analysis demonstrated that HCB could increase TGF-β1 protein levels and activation, as well as Smad3, JNK, and p38 phosphorylation. In addition, low and high doses of HCB were determined to exert differential effects on AhR protein levels, localization, and activation, with a high dose (5μM) inducing AhR nuclear translocation and AhR-dependent CYP1A1 expression. These findings also revealed that c-Src and AhR are involved in HCB-mediated activation of Smad3. HCB enhances cell migration (scratch motility assay) and invasion (Transwell assay) through the Smad, JNK, and p38 pathways, while ERK1/2 is only involved in HCB-induced cell migration. These results demonstrate that HCB modulates the crosstalk between AhR and TGF-β1 and consequently exacerbates a pro-migratory phenotype in MDA-MB-231 cells, which contributes to a high degree of malignancy. Taken together, our findings help to characterize the molecular mechanism underlying the effects of HCB on breast cancer progression.

Sulforaphene Interferes with Human Breast Cancer Cell Migration and Invasion through Inhibition of Hedgehog Signaling.

Although inhibition of mammary tumorigenesis by isothiocyanates has been widely studied, little is known about the effects of sulforaphene on invasiveness of breast cancer. Here, sulforaphene significantly inhibited the migration and invasion of triple-negative SUM159 human breast cancer cells and suppressed the expression and activity of matrix metalloproteinases 2 and 9 (MMP-2 and MMP-9). The Hedgehog (Hh) pathway, as an upstream signaling modulator, was significantly suppressed by sulforaphene. In particular, ciliary localization of Gli1 and its nuclear translocation were blocked by sulforaphene in a time-dependent manner. Consistently, downregulation of Hh signaling by vismodegib and Gli1 knockdown reduced the cellular migration and invasion as well as the expression of MMP-2 and MMP-9. These results indicate that the suppression of Hh/Gli1 signaling by sulforaphene may reduce the MMP-2 and MMP-9 activities and cellular invasiveness of human breast cancer cells, suggesting the potential efficacy of sulforaphene against breast cancer invasion and metastasis.

Inhibition of p38 activity reverses claudin-6 induced cell apoptosis, invasion, and migration.

Claudin-6 is a protein component of tight junctions and its expression could downregulate the malignant phenotype of breast carcinoma. Here we investigated the mechanisms of claudin-6 induced human MCF-7 breast cancer cells apoptosis, invasion, and migration. Terminal deoxyribonucleotide transferase-mediated nick-end labeling assay and Annexin-V/PI double stain assay were carried out to evaluate apoptosis. Inhibitors of each pathway were used to inactivate the signaling pathways. The expression of claudin-6 and phosphate p38, Erk 1/2 and Akt protein levels was confirmed by Western blotting analysis. Invasive and migratory traits of claudin-6 expressing cells were determined by Boyden chamber invasion assay and monolayer wound-healing assay. Cells with high-level expression of claudin-6 had a higher rate of apoptosis than control cells. Western blotting assay showed that by contrast to control groups, p38 pathways were more activated in claudin-6 expressing cells. However, after inhibitor SB203580 treatment, the activation status could be significantly counteracted. Furthermore, by applying inhibitors to the apoptotic rate, invasive and migratory traits were also recovered in cells with claudin-6 expression. Claudin-6 may function through p38 mitogen-activated protein kinase pathway, of which inhibition may reverse claudin-6-induced cell apoptosis, invasion, and migration.

Klotho Regulation of TRPC Channels Involving Human Breast Cancer Cell Migration

Klotho is a mammalian aging‐suppression gene and function as a suppressor of cancer. Here, we investigated the molecular mechanism of Klotho regulation of TRPC channels involving human breast cancer cell migration. Serum‐induced activation of TRPC and cell migration was specifically blocked by TRPC channel inhibitor, SKF96365, but not by voltage‐gated Ca2+ channel inhibitor, nimodipine. Treatment of purified Klotho protein inhibited TRPC6 channel activity. Interestingly, serum treatment increased the surface expression of TRPC6 channel. Klotho suppressed cell surface abundance of TRPC6 channels, whereas did not affect gating and activation kinetics of the channel. In addition, treatment of PI3K inhibitor (Wortmannin) or Erk1/2 inhibitor (U0126) blocked the serum‐induced augmentation of TRPC6. Suppression of vesicular trafficking by treatment of brefeldin A or tetanus toxin significantly decreased TRPC6 activity and Klotho regulation.Taken together, our data suggest that Klotho decreases surface expression of TRPC6 probably by reducing serum growth factor‐induced secretory pathway of the channel. Klotho may function as suppressor for human breast cancer cell migration caused by upregulation of TRPC6.[This research was supported by Basic Science Research Program through the National Research Foundation of Korea funded by the Ministry of Education, Science and Technology (2010–0024789)]

Cytotoxic 3,4,5-trimethoxychalcones as mitotic arresters and cell migration inhibitors

Based on classical colchicine site ligands and a computational model of the colchicine binding site on beta tubulin, two classes of chalcone derivatives were designed, synthesized and evaluated for inhibition of tubulin assembly and toxicity in human cancer cell lines. Docking studies suggested that the chalcone scaffold could fit the colchicine site on tubulin in an orientation similar to that of the natural product. In particular, a 3,4,5-trimethoxyphenyl ring adjacent to the carbonyl group appeared to benefit the ligand–tubulin interaction, occupying the same subcavity as the corresponding moiety in colchicine. Consistent with modeling predictions, several 3,4,5-trimethoxychalcones showed improved cytotoxicity to murine acute lymphoblastic leukemia cells compared with a previously described parent compound, and inhibited tubulin assembly in vitro as potently as colchicine. The most potent chalcones inhibited the growth of human leukemia cell lines at nanomolar concentrations, caused microtubule destabilization and mitotic arrest in human cervical cancer cells, and inhibited human breast cancer cell migration in scratch wound and Boyden chamber assays.

The effects of annexin a2 expression on the proliferation, migration and invasion of human breast cancer cells

The effects of annexin a2 expression on the proliferation, migration and invasion of human breast cancer cells

On-chip cell migration assay for quantifying the effect of ethanol on MCF-7 human breast cancer cells

Ethanol consumption is associated with the risk of breast cancer progression; however, the mechanism of relationship has not yet been fully explained. Research on breast cancer cell migration after ethanol stimulation may give hope for a better understanding of the disease and oncotherapy. Conventional cell migration assays such as Boyden chamber and wound-healing assays are easy to conduct for this purpose; however, these assays have inherent limitations. In this study, we quantified the effect of ethanol on MCF-7 hunam breast cancer cells using a microfluidics-based wound-healing assay. Wounds were prepared by partially digesting a confluent cell sheet using parallel laminar flows in the presence of protease trypsin. The cells at the leading edge of the wound remained intact. Cell image analysis indicates that all the cells cultured in the microdevice took on a good morphology and monolayer growth status. Cell viability assay demonstrates that cell viability decreased with the increase in ethanol concentration and treatment time. For 0, 22, 43, and 65 mmol/l of ethanol, cell viability after being cultured for 24 h was 100%, 99.6%, 99.4%, and 98.4%, respectively. Studying MCF-7 human breast cancer cell migration when treated with different ethanol concentrations revealed that the cell migration distance is directly proportional with ethanol concentration. After being cultured for 24 h at 37°C and 5% CO2, the maximal cell migration distance was 231, 283, and 332 μm for 22, 43, and 65 mmol/l ethanol, respectively; all results were higher than the blank test (i.e., ethanol-free test, 218 μm). These findings will be beneficial in developing microfluidic device applications for future research on breast tumor therapy in a biomimetic microenvironment and for developing new methods for breast cancer therapy.

Tumour suppressor function of MDA-7/IL-24 in human breast cancer

IntroductionMelanoma differentiation associated gene-7 (MDA-7), also known as interleukin (IL)-24, is a tumour suppressor gene associated with differentiation, growth and apoptosis. However, the mechanisms underlying its anti-neoplastic activity, tumour-specificity and efficacy across a spectrum of human cancers have yet to be fully elucidated. In this study, the biological impact of MDA-7 on the behavior of breast cancer (BC) cells is evaluated. Furthermore, mRNA expression of MDA-7 is assessed in a cohort of women with BC and correlated with established pathological parameters and clinical outcome.MethodsThe human BC cell line MDA MB-231 was used to evaluate the in-vitro impact of recombinant human (rh)-MDA-7 on cell growth and motility, using a growth assay, wounding assay and electric cell impedance sensing (ECIS). Localisation of MDA-7 in mammary tissues was assessed with standard immuno-histochemical methodology. BC tissues (n = 127) and normal tissues (n = 33) underwent RNA extraction and reverse transcription, MDA-7 transcript levels were determined using real-time quantitative PCR. Transcript levels were analyzed against tumour size, grade, oestrogen receptor (ER) status, nodal involvement, TNM stage, Nottingham Prognostic Index (NPI) and clinical outcome over a 10 year follow-up period.ResultsExposure to rh-MDA-7 significantly reduced wound closure rates for human BC cells in-vitro. The ECIS model demonstrated a significantly reduced motility and migration following rh-MDA-7 treatment (p = 0.024). Exposure to rh-MDA-7 was only found to exert a marginal effect on growth. Immuno-histochemical staining of human breast tissues revealed substantially greater MDA-7 positivity in normal compared to cancer cells. Significantly lower MDA-7 transcript levels were identified in those predicted to have a poorer prognosis by the NPI (p = 0.049) and those with node positive tumours. Significantly lower expression was also noted in tumours from patients who died of BC compared to those who remained disease free (p = 0.035). Low levels of MDA-7 were significantly correlated with a shorter disease free survival (mean = 121.7 vs. 140.4 months, p = 0.0287) on Kaplan-Meier survival analysis.ConclusionMDA-7 significantly inhibits the motility and migration of human BC cells in-vitro. MDA-7 expression is substantially reduced in malignant breast tissue and low transcript levels are significantly associated with unfavourable pathological parameters, including nodal positivity; and adverse clinical outcomes including poor prognosis and shorter disease free survival. MDA-7 offers utility as a prognostic marker and potential for future therapeutic strategies.

MiR-17-5p promotes human breast cancer cell migration and invasion through suppression of HBP1

MicroRNAs have been implicated in regulating diverse cellular pathways. Emerging evidence indicate that the miR-17-92 cluster may have a causal role in breast cancer tumorigenesis as a novel class of oncogenes, but the role of these miRNAs in breast cancer invasion and migration remains unexplored. The aims of this study were to verify the effect of miR-17-5p (an important member of the miR-17-92 cluster) on the invasive and migratory ability of breast cancer cells. The matching of miR-17-5p and HMG box-containing protein 1 (HBP1) was predicted by TargetScan and confirmed by DNA constructs and luciferase target assay. The expression levels of miR-17-5p and its candidate target-HBP1 in MCF7 and MDA-MB-231 breast cancer cells were measured by real-time PCR and western blotting. Effects of miR-17-5p in cell cycle progression, proliferation, invasion and migration were evaluated by flow cytometry assay, 3-(4,-dimethy -lthiazol-2-yl)-2,-diphenyl -tetrazoliumbromide assay, soft-agar colony formation assay, and transwell invasive and migratory assay, respectively. The results showed that miR-17-5p was highly expressed in high-invasive MDA-MB-231 breast cancer cells but not in low-invasive MCF-7 breast cancer cells. Over-expression of miR-17-5p in MCF-7 cells rendered them the invasive and migratory abilities by targeting HBP1/β-catenin pathway. On the other hand, down-regulation of endogenous miR-17-5p suppressed the migration and invasion of MDA-MB-231 cells in vitro. These findings suggest that miR-17-5p plays an important role in breast cancer cell invasion and migration by suppressing HBP1 and subsequent activation of Wnt/β-catenin.