Migration Of MCF-7 Cells Research Articles

Role of synaptotagmin 13 (SYT13) in promoting breast cancer and signaling pathways.

Breast cancer is one of the leading causes of tumor death worldwide in female, and the five-year overall survival of breast cancer patients remains poor. It is an urgent need to seek novel target for its treatment. Synaptotagmin 13 (SYT13) is a synaptic vesicle transporting protein that regulates the malignant phenotypes of various cancers. However, its role in breast cancer is still unclear. The current study aimed to investigate the effects of SYT13 on the progression of breast cancer. Twenty-five pairs of breast cancer tissues and non-tumor tissues were obtained to assess the expression of SYT13. We manually modified the expression of SYT13 in MCF-7 and MDA-MB-231 cells. CCK-8 assay, EdU staining, and cell cycle analysis were carried out to measure the proliferated ability of cells. Annexin V/PI and TUNEL assays were used to detect the apoptotic ability of cells. Wound healing and transwell assays were employed to evaluate the migrated and invasive ability of breast cancer cells. The results revealed that the mRNA and protein levels of SYT13 were higher in breast cancer tissues and cell lines. Knockdown of SYT13 inhibited the cell proliferation and induced cell cycle arrest in G1 phase of MCF-7 cells by downregulating cyclin D1 and CDK4, as well as upregulating p21. The migration and invasion of MCF-7 cells were repressed by the loss of SYT13 via the gain of E-cadherin and the loss of vimentin. Overexpression of SYT13 in MDA-MB-231 cells led to the opposite effects. Silencing of SYT13 induced the apoptosis ability of MCF-7 cells by the upregulation of bax and the downregulation of bcl-2. Moreover, we found that SYT13 depletion suppressed the FAK/AKT signaling pathway. PF573228 (a FAK inhibitor) and MK2206 (an AKT inhibitor) reversed the SYT13 overexpression-induced promotion of proliferation, migration, and invasion of MDA-MB-231 cells. The results indicated that SYT13 promoted the malignant phenotypes of breast cancer cells by the activation of FAK/AKT signaling pathway.

BML-111, the agonist of lipoxin A4, suppresses epithelial-mesenchymal transition and migration of MCF-7 cells via regulating the lipoxygenase pathway.

Introduction: Aberrant epithelial-mesenchymal transition (EMT) and migration frequently occur during tumour progression. BML-111, an analogue of lipoxin A4, has been implicated in inflammation in cancer research. Methods: 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay, western blot, Reverse Transcription Polymerase Chain Reaction (RT-PCR), transwell assay, immunofluorescence, and immunohistochemistry were conducted in this study. Results: In vitro experiments revealed that BML-111 inhibited EMT and migration in CoCl2-stimulated MCF-7 cells. These effects were achieved by inhibiting MMP-2 and MMP-9, which are downregulated by 5-lipoxygenase (5-LOX). Moreover, BML-111 inhibited EMT and migration of breast cancer cells in BALB/c nude mice inoculated with MCF-7 cells. Conclusion: Our results suggest that BML-111 may be a potential therapeutic drug for breast cancer and that blocking the 5-LOX pathway could be a possible approach for mining effective drug targets.

Effects of the Ethanol and Ethyl Acetate Extracts of Terminalia chebula Retz. on Proliferation, Migration, and HIF-1α and CXCR-4 Expression in MCF-7 Cells: an In Vitro Study.

Over recent years, much attention has been devoted to the field of screening natural products and/or their novel structures because of reversing cancer progression. The current research work was intended to explore the cytotoxic activity of ethanol and ethyl acetate extracts of dried fruit of Terminalia chebula Retz. (T. chebula) in MCF-7 cell line. High-performance thin-layer chromatographic (HPTLC) method and Folin-Ciocalteu colorimetric techniques were performed. Anti-proliferative activities of T. chebula fruit extracts on the MCF-7 cell line were evaluated using MTT assay. Effects of both extracts on the migration of MCF-7 cells and the size of MCF-7-derived spheroids were also evaluated. Moreover, antioxidant properties were measured by DPPH and FRAP methods. Western blotting was used to measure the HIF-1α and CXCR-4 protein levels. Chebulagic acid, gallic acid, chebulinic acid, and ellagic acid were found as major compounds in both extracts. The total phenolic contents based on gallic acid equivalent (GAE) in the ethanol and ethyl acetate extracts of T. chebula were found to be 453.68 ± 0.31 and 495.12 ± 0.43mg GAE/g dry weight of the extract, respectively. Both extracts exerted a significant dose- and time-dependent cytotoxicity effect on MCF-7 cells. They also had a marked negative effect on the average size of MCF-7-derived spheroids and their migration rate. None of the extracts exhibited stronger antioxidant activities than vitamin C. Furthermore, both extracts at a concentration of 125µg/ml could meaningfully decrease the expression levels of HIF-1α and CXCR-4 in MCF-7 cells. These data represent that T. chebula may be a valuable medicinal resource in the regulation of breast cancer proliferation, growth, and metastasis.

Δ9-Tetrahydrocannabinol stimulation of estrogen receptor-positive MCF-7 breast cancer cell migration: Interfering interaction with the estrogenic milieu.

The effects of extended Δ9-tetrahydrocannabinol (Δ9-THC) exposure on estrogen receptor-positive human breast cancer MCF-7 cells have been investigated; however, the effects of Δ9-THC exposure for a shorter duration remain unclear. In this study, we sought to study whether Δ9-THC stimulates the migration of MCF-7 cells under both estrogenic and estrogen-deprived conditions over a short period (approximately 6h). MCF-7 cells were treated with Δ9-THC under estrogenic or estrogen-deprived conditions, and cell migration was subsequently analyzed. Δ9-THC-stimulated migration of MCF-7 cells 6h after exposure was only observed in the estrogen-deprived condition. However, Δ9-THC-mediated migration was counteracted under estrogenic conditions without affecting cell proliferation and estrogen receptor expression during this period. Δ9-THC can stimulate MCF-7 cell migration under estrogen-deprived conditions; however, there is an interfering interaction between Δ9-THC and the estrogenic milieu that influences the migration of MCF-7 cells.

The role of PI3K/AKT-related proteins in di(2-ethyl)hexylphthalate-induced BG-1 and MCF-7 cell proliferation, and inhibition by metformin

Objective To investigate the molecular mechanism of the proliferation and migration of BG-1 and MCF-7cells induced by DEHP, and the antagonistic effect of metformin. Methods The proliferation, cell cycle progression, migration, and invasion abilities of BG-1 and MCF-7 cancer cells were examined via Cell Counting Kit-8, flow cytometry, Transwell, and scratch assays. E2F1, SKP2, cyclin D1, vimentin, E-cadherin, and GSK-3β, all of which play key roles in cancer development via the PI3K/AKT signaling pathway, were examined by immunofluorescence and immunocytochemistry. Results Cell proliferation was significantly increased, and the wound closure and number of trans-membrane migrating cells were significantly increased, by DHEP treatment. The numbers of BG-1 and MCF-7 cells in the G0/G1 phase were significantly decreased, while those in the S phase were significantly increased. Increased E2F1, SKP2, cyclin D1, and vimentin levels and decreased E-cadherin and GSK-3β levels were detected in BG-1 and MCF-7 cells treated with DEHP compared to that in control cells. Furthermore, DEHP-induced effects were markedly inhibited by LY294002 (a PI3K/AKT inhibitor) or metformin. Conclusion We demonstrated that DEHP-induced cell proliferation and migration involves the PI3K/AKT signaling pathway and that metformin can be used to inhibit this proliferation and migration.

Thymoquinone-treated mouse mesenchymal stem cells-derived conditioned medium inhibits human breast cancer cells in vitro

Breast cancer is now the most prevalent cancer in females, therefore, it is essential to identify factors affecting its initiation and progression. Mesenchymal stem cells (MSCs) have received considerable attention in stem cell-based therapies and drug delivery applications. Because the therapeutic potential of MSCs is primarily achieved by their paracrine effects, thus identifying and employing bioactive molecules that promote the paracrine activity of MSCs is crucial for their efficient use in cancer treatment. Thymoquinone (TQ) has many biomedical properties, including anti-inflammatory, anti-diabetic, anti-aging, anti-cancer, etc. In addition, it has been found that TQ affects the self-renewal and immunomodulatory properties of MSCs. The present study aimed to investigate the effect of TQ-treated mouse bone marrow-derived MSCs conditioned medium (TQ-MSC-CM) on the biological characteristics of breast cancer cell line MCF7. MSCs were cultured and treated with TQ for 24 h. The TQ-MSC-CM and MSC-CM were collected, and their effects were investigated on ROS production, mitochondrial membrane potential (MMP), cell death, cell cycle, and migration of MCF7 cells by DCFDA-cellular ROS assay, Rhodamine-123 MMP assay, Annexin-PI staining and Caspase-3/7 activity assays, PI-staining and flow-cytometry, and in vitro wound healing assay, respectively. Moreover, the effects of TQ-MSC-CM and MSC-CM were studied on Cdk4, Sox2, c-Met, and Bcl2 gene expression by real-time PCR. Results demonstrated that MSC-CM and TQ-MSC-CM did not have a significant effect on the apoptosis induction in MCF7 cells; however, they significantly stimulated necrosis in the cells. Although TQ-MSC-CM promoted ROS production in MCF7 cells, it decreased the MMP of the cells. TQ-MSC-CM also induced Bcl2 anti-apoptosis gene expression and Casp-3/7 activity in cells. In addition, although MSC-CM induced MCF7 cells to enter the cell cycle, TQ-MSC-CM inhibited its progression. TQ-MSC-CM also downregulated the Cdk4 and Sox2 gene expression. Furthermore, TQ-MSC-CM induced the migration potential of MCF7 in a c-Met-independent manner. Altogether, we conclude that TQ may induce programmed necrosis and inhibits the proliferation and migration of the breast cancer cells by affecting the paracrine activity of MSCs.

Deciphering the Role and Signaling Pathways of PKCα in Luminal A Breast Cancer Cells.

Protein kinase C (PKC) comprises a family of highly related serine/threonine protein kinases involved in multiple signaling pathways, which control cell proliferation, survival, and differentiation. The role of PKCα in cancer has been studied for many years. However, it has been impossible to establish whether PKCα acts as an oncogene or a tumor suppressor. Here, we analyzed the importance of PKCα in cellular processes such as proliferation, migration, or apoptosis by inhibiting its gene expression in a luminal A breast cancer cell line (MCF-7). Differential expression analysis and phospho-kinase arrays of PKCα-KD vs. PKCα-WT MCF-7 cells identified an essential set of proteins and oncogenic kinases of the JAK/STAT and PI3K/AKT pathways that were down-regulated, whereas IGF1R, ERK1/2, and p53 were up-regulated. In addition, unexpected genes related to the interferon pathway appeared down-regulated, while PLC, ERBB4, or PDGFA displayed up-regulated. The integration of this information clearly showed us the usefulness of inhibiting a multifunctional kinase-like PKCα in the first step to control the tumor phenotype. Then allowing us to design a possible selection of specific inhibitors for the unexpected up-regulated pathways to further provide a second step of treatment to inhibit the proliferation and migration of MCF-7 cells. The results of this study suggest that PKCα plays an oncogenic role in this type of breast cancer model. In addition, it reveals the signaling mode of PKCα at both gene expression and kinase activation. In this way, a wide range of proteins can implement a new strategy to fine-tune the control of crucial functions in these cells and pave the way for designing targeted cancer therapies.

ODP562 ZnT5 Involvement in EMT in Breast Cancer

Abstract The zinc levels in breast cancer tissue are reportedly higher than in normal tissue. In addition, the expression levels of zinc transporters, including ZnT5 and ZnT6, in breast cancer tissue are higher than in normal breast tissue. Moreover, ZnT5 and ZnT6 contribute to the formation of heterodimers and are involved in different biological functions. In vitro studies using chicken DT40 cells showed that ZnT5 and ZnT6 heterodimers are required to activate several proteins that play important roles in tumorigenesis, infiltration, and metastasis. However, the functions of ZnT5 and ZnT6 heterodimers in breast cancer remain unknown. Therefore, in this study, we first immunolocalized ZnT5 and ZnT6 in 113 pathological specimens of breast cancer. We then analyzed the interaction between ZnT5 and ZnT6 in breast cancer tissue using proximity ligation assay (PLA). Further, we evaluated cell migration and the expression of epithelial mesenchymal transition (EMT) markers as well as vimentin and E-cadherin in ZnT5 knockdown MCF-7 cells using siRNA. Next, we utilized human phosphorylation multipathway profiling array to explore the underlying mechanism of ZnT5 knockdown-induced cell migration. Immunohistochemical analysis revealed that the number of ZnT5-positive breast cancer cells was significantly higher in patients with a low pathologic N factor, and the number of ZnT6-positive breast cancer cells was significantly higher in patients with a low histological grade. The number of ZnT5 and ZnT6 double-positive breast cancer cells was significantly higher in patients with low Ki-67 expression than those with ZnT5-positive and ZnT6-negative breast cancer cells. In addition, ZnT5 and ZnT6 heterodimers were detected using PLA in breast cancer tissues with high expression of both ZnT5 and ZnT6. Treatment with 100 µM ZnCl 2 inhibited migration of MCF-7 cells, but ZnT5 knockdown promoted cell migration. ZnT5 knockdown induced higher levels of vimentin and lower levels of E-cadherin and activated SMAD1 in the presence of 100 µM ZnCl 2 . These results indicate that the involvement of ZnT5 in the inhibition of EMT through SMAD1 inactivation. These results also indicate that the role of ZnT5 and ZnT6 heterodimers differs from that of the ZnT5 homodimer in cell proliferation. However, further studies are required to clarify the role of zinc transporters and zinc signaling in breast cancer. Presentation: No date and time listed

L-arginase from Streptomyces diastaticus MAM5 as a potential therapeutic agent in breast cancer: Purification, characterization, G1 phase arrest and autophagy induction

Targeting cancer metabolic processes has increased interest over the last century. Cancer cells have an enhanced proliferation rate that requires high quantities of amino acids, including arginine. Therefore, arginine deprivation by L-arginase impairs tumor growth resulting in cell death. In the present study, L-arginine amidinohydrolase (L-arginase) from Streptomyces diastaticus was purified successfully by heat treatment, ethanol precipitation, and Sephadex G75–120 column. The molecular mass of the purified enzyme was 39 kDa. It showed maximum activity at pH 9.0 and a temperature of 50 °C. Moreover, the enzyme stability was observed at temperatures up to 50 °C and a pH range of 7.5 to 9.0. Then, the potential cytotoxicity of L-arginase was examined. L-arginase has an IC50 value of 595 μg/ml for MCF-7 (breast adenocarcinoma cells), 915 μg/ml for HepG2 (hepatocellular carcinoma cells), and 1200 μg/ml for SW620 cells (colorectal carcinoma cells) at 72 h post-treatment. Noteworthy, MCF-7 showed the lowest IC50 value of arginase treatment, therefore was further investigated for the underlying cytotoxic mechanisms using flow cytometric analysis of cell-cycle distribution, apoptosis, and autophagy. Moreover, SI values indicating a high selective cytotoxicity of arginase toward MCF-7 cells. L-arginase induced significant cell cycle arrest at the G1 phase, and no apparent apoptosis was detected. Interestingly, arginine deprivation by arginase leads to a prominent activation of autophagy in the apoptosis defected MCF-7 cells. Moreover, treatment with arginase significantly attenuated MCF-7 cell migration compared with control medium-treated cells. Collectively, L-arginase might potentially be involved in treating breast cancer.

MiR-219a-5p Inhibits Michigan Cancer Foundation-7 (MCF-7) Cell Migration by Regulating Zinc Finger E-Box Binding Homeobox 1 (ZEB1) Expression

The potential role of miR-219a-5p in the migration of breast cancer has not been fully elucidated. In this study, bioinformatic analysis revealed that high miR-219a-5p expression in breast cancer tissue was associated with good survival of breast cancer patients. RT-qPCR analysis indicated that miR-219a-5p expression is significantly lower in MDA-MB-231 triple-negative breast cancer (TNBC) cells. In addition, pre-miR-219a overexpression inhibited MCF-7 cell migration and inhibited ZEB1, Twist1 and vimentin expression but promoted the expression of E-cadherin. Moreover, miR-219a-5p mimics inhibited MCF-7 cell migration, whereas MCF-7 cell migration was promoted by the miR-219a-5p inhibitor. Furthermore, miR-219a-5p was found to inhibit the translation of ZEB1 expression by targeting the 5′-ACAAUCA-3′ motif of the ZEB1 3′UTR, and the binding motif is conserved in multiple species. ZEB1 overexpression rescued the inhibition of cell migration induced by miR-219a-5p. Finally, an inverse correlation of miR-219a-5p and ZEB1 expression was observed in four breast cancer cell lines. Thus, miR-219a-5p inhibits MCF-7 cell migration by regulating ZEB1 expression, and ZEB1 is the target gene of miR-219a-5p.

BRCA1 overexpression attenuates breast cancer cell growth and migration by regulating the pyruvate kinase M2-mediated Warburg effect via the PI3K/AKT signaling pathway.

This work explored the mechanism of the effect of breast-cancer susceptibility gene 1 (BRCA1) on the metabolic characteristics of breast cancer cells, including the Warburg effect and its specific signaling. We transfected MCF-7 cells with a BRCA1-encoding LXSN plasmid or PKM2 siRNA and examined cancer cell metabolism using annexin V staining, inhibitory concentration determination, Western blotting, glucose uptake and lactic acid content measurements, and Transwell assays to assess glycolytic activity, cell apoptosis, and migration, and sensitivity to anti-cancer treatment. The BRCA1-expressing MCF-7 cells demonstrated low PKM2 expression and decreased glycolytic activity (downregulated hexokinase 2 (HK2) expression, upregulated isocitrate dehydrogenase 1 (IDH1) expression, and reduced O2 and glucose consumption and lactate production) via regulation of PI3K/AKT pathway compared with the empty LXSN group. BRCA1 transfection slightly increased apoptotic activity, decreased cell migration, and increased the IC50 index for doxorubicin, paclitaxel, and cisplatin. Inhibiting PKM2 using siRNA attenuated the IC50 index for doxorubicin, paclitaxel, and cisplatin compared with the control. Inhibiting PKM2 activated PI3K/AKT signaling, increased apoptosis, and decreased MCF-7 cell migration. Our data suggest that BRCA1 overexpression reverses the Warburg effect, inhibits cancer cell growth and migration, and enhances the sensitivity to anti-cancer treatment by decreasing PKM2 expression regulated by PI3K/AKT signaling. These novel metabolic findings represent a potential mechanism by which BRCA1 exerts its inhibitory effect on breast cancer.

A significantly non-toxic novel Cobalt(III) Schiff base complex induces apoptosis via G2-M cell cycle arrest in human breast cancer cell line MCF-7

AimsMetal complexes have ignited considerable interest in the field of chemotherapy after the serendipitous discovery of cisplatin but the severe toxicity of these platinum-based drugs compelled researchers to search for newer, more effective lesser toxic anticancer drugs. Materials and methodsStructural analysis is done by different physicochemical techniques including X-ray single crystallography. Toxicity study has been done in normal Swiss albino mice. MTT assay assessed cell viability. Apoptosis, cell cycle arrest, and cell proliferation were assessed by FACS using Annexin V-PI, PI, and CFSE staining respectively. Western blot quantifies protein expression. While cell migration was studied by wound healing assay. Key findingsOne-pot synthesis of a novel mononuclear cobalt(III)-Schiff base complex (1) (>99 % purity) and its complete characterization have been done. Cell viability assay showed that 1 (IC50 = 16.81 ± 1.33 μM) exhibits cytotoxicity at much lower concentration in comparison to oxaliplatin (IC50 = 31.4 ± 0.69 μM) against MCF-7 cells for 24 h of therapy without being overly toxic to human PBMCs (IC50 ≥ 60 μM). Additional in vitro studies demonstrated that 1 induces apoptosis via G2-M cell cycle arrest and reduces cell proliferation as well as cell migration in MCF-7 cells. In vivo subacute toxicity (28 days) and systemic chronic toxicity (40 days) studies were carried out in normal Swiss albino mice showed 1 is significantly nontoxic to the host. SignificanceThe readily synthesizable, significantly nontoxic cobalt complex with appreciable anticancer activity implies that it might be an effective chemotherapeutic agent for new-age anti-tumor medication.

Fucoxanthin prevents breast cancer metastasis by interrupting circulating tumor cells adhesion and transendothelial migration.

Metastasis is the leading cause of cancer-related death and a critical challenge in improving cancer treatment today. Circulating tumor cells (CTCs) adhesion to and across the vascular endothelium are critical steps in the establishment of micrometastatic foci away from the primary tumor. Therefore, we believe that interrupting CTCs adhesion to endothelium and transendothelial migration may efficiently prevent cancer metastasis. Fucoxanthin (Fx) is an algal carotenoid widely distributed in brown algae, macroalgae, and diatoms. Previous studies have found that Fx has various pharmacological activities, including antidiabetic, antioxidant, anti-inflammatory, anti-obesity, antimalarial, anticancer, and so on. However, it remains unclear whether Fx has a preventive effect on cancer metastasis. Here, we found that Fx interrupts breast cancer cells MCF-7 adhesion to endothelium and transendothelial migration, thus inhibiting CTCs-based pulmonary metastasis in vivo. The hetero-adhesion assay showed that Fx significantly inhibited the expression of inflammatory factor-induced cell adhesion molecules (CAMs) and the resulting adhesion between MCF-7 cells and endothelial cells. The wound-healing and transwell assays showed that Fx significantly inhibited the motility, invasion, and transendothelial migration abilities of MCF-7 cells. However, the same concentration of Fx did not significantly alter the cell viability, cell cycle, apoptosis, and ROS of breast cancer cells, thus excluding the possibility that Fx inhibits MCF-7 cell adhesion and transendothelial migration through cytotoxicity. Mechanistically, Fx inhibits the expression of CAMs on endothelial cells by inhibiting the NF-кB signaling pathway by down-regulating the phosphorylation level of IKK-α/β, IкB-α, and NF-кB p65. Fx inhibits transendothelial migration of MCF-7 cells by inhibiting Epithelial-to-mesenchymal transition (EMT), PI3K/AKT, and FAK/Paxillin signaling pathways. Moreover, we demonstrated that Fx significantly inhibits the formation of lung micrometastatic foci in immunocompetent syngeneic mouse breast cancer metastasis models. We also showed that Fx enhances antitumor immune responses by substantially increasing the subsets of cytotoxic T lymphocytes in the peripheral immune system. This new finding provides a basis for the application of Fx in cancer metastatic chemoprevention and suggests that interruption of the CTCs adhesion to endothelium and transendothelial migration may serve as a new avenue for cancer metastatic chemoprevention.

Green synthesized nickel doped cobalt ferrite nanoparticles exhibit antibacterial activity and induce reactive oxygen species mediated apoptosis in MCF-7 breast cancer cells through inhibition of PI3K/Akt/mTOR pathway.

This study aimed to synthesize the nickel (Ni) doped cobalt ferrite (CFO) nanoparticles and to determine its anticancer effect on breast cancer MCF-7 cells. The various concentrations of Ni (0.2%, 0.4%, 0.6%, 0.8%, and 1%) doped CFO nanoparticles were synthesized using Coriandrum sativum extracts by precipitation technique. The synthesized Ni-CFO was characterized by X-ray diffraction, Fourier transform infrared spectroscopic, transmission electron microscopy, and vibrating sample magnetometer analyses. The results show that 0.8% Ni-CFO nanoparticles showed good magnetic properties and antioxidant activity than other concentrations of Ni-CFO. The results showed that the administration of 0.8% Ni-CFO nanoparticles promoted apoptosis, and reduced cell adhesion and migration of MCF-7 cells, as demonstrated by increased lipid peroxidation, decreased antioxidant levels such as superoxide dismutase, catalase, and glutathione peroxidase, and increased formation of reactive oxygen species. Moreover, administration of 0.8% Ni-CFO nanoparticles decreased the Bcl-2 expression while activating the expression of p53, Bax, and cleaved caspase 3, 8 and 9 protein expression. Notably, 0.8% Ni-CFO treatment reduced phospho-PI3K, phospho-Akt, and phospho-mTOR expression levels. As a result, via altering apoptotic related proteins, 0.8% Ni-CFO induced cell death. Therefore, the 0.8% Ni-CFO could be utilized to treat breast cancer.

Discovery of novel tetrahydropyrido[4,3-d]pyrimidine analogs as potent autotaxin regulators with impressive tumor suppression effects

Aberrant ATX-LPA signaling turns out to be a powerful driver for the development of various cancers. Herein, a series of unique tetrahydropyrido[4,3-d]pyrimidine compounds were designed and synthesized via a hybrid strategy to serve as novel ATX inhibitors. Benzamide fragment was introduced perpendicular to the 4-positon of tetrahydropyrido[4,3-d]pyrimidine skeleton to obtain compounds 7∼12c. To retrieve satisfying druggable profile, 4-aminopiperidine analogs (14a∼15b) were investigated in parallel. ATX enzymatic assay indicated the series of compounds endowed with moderate to satisfying ATX inhibitory activity. Delightfully, the benzamide 11c bearing a N6-benzyl group was identified as the optimal compound with an excellent IC50 value of 3.72 nM against ATX enzyme, accompanied by impressive cytotoxicity, especially on MCF-7 (0.67 µM) and RAW264.7 (0.83 µM) cell lines. Furthermore, wound healing assays and colony formation assays demonstrated 11c could significantly suppress the MCF-7 cell migration rate and colony formation in a dose dependent manner. Meanwhile, 11c was capable of inducing apoptosis on MCF-7 cells in the immunofluorescence assays. Taken together, the novel compound 11c with favorable druggability may serve as a promising candidate in cancer treatment.

Steviol glycosides affect functional properties and macromolecular expression of breast cancer cells.

Steviol glycosides, the active sweet components of stevia plant, have been recently found to possess a number of therapeutic properties, including some recorded anticancer ones against various cancer cell types (breast, ovarian, cervical, pancreatic, and colon cancer). Our aim was to investigate this anticancer potential on the two most commonly used breast cancer cell lines which differ in the phenotype and estrogen receptor (ER) status: the low metastatic, ERα+ MCF-7 and the highly metastatic, ERα-/ERβ+ MDA-MB-231. Specifically, glycosides' effect was studied on cancer cells': (a) viability, (b) functionality (proliferation, migration, and adhesion), and (c) gene expression (mRNA level) of crucial molecules implicated in cancer's pathophysiology. Results showed that steviol glycosides induced cell death in both cell lines, in the first 24 hr, which was in line with the antiapoptotic BCL2 decrease. However, cells that managed to survive showcased diametrically opposite behavior. The low metastatic ERα+ MCF-7 cells acquired an aggressive phenotype, depicted by the upregulation of all receptors and co-receptors (ESR, PGR, AR, GPER1, EGFR, IGF1R, CD44, SDC2, and SDC4), as well as VIM and MMP14. On the contrary, the highly metastatic ERα-/ERβ+ MDA-MB-231 cells became less aggressive as pointed out by the respective downregulation of EGFR, IGF1R, CD44, and SDC2. Changes observed in gene expression were compatible with altered cell functions. Glycosides increased MCF-7 cells migration and adhesion, but reduced MDA-MB-231 cells migratory and metastatic potential. In conclusion, the above data clearly demonstrate that steviol glycosides have different effects on breast cancer cells according to their ER status, suggesting that steviol glycosides might be examined for their potential anticancer activity against breast cancer, especially triple negative breast cancer (TNBC).

46P Inhibition of cell invasion and migration by targeting matrix metalloproteinase-9 expression via sirtuin 6 silencing in human breast cancer cells

Sirtuin 6 (SIRT6) regulation is involved in carcinogenesis. However, its role in breast cancer (BC) metastasis remains unclear. We investigated the effects of SIRT6 on protein kinase C activator- and cytokine-mediated cancer cell invasion and migration in MCF-7 and MDA-MB-231 cells and the association between SIRT6 and matrix metalloproteinase-9 (MMP-9) expression.

Antipyrine derived-Schiff base copper complex: Synthesis, characterization, and in vitro evaluation

The copper(II) complex (NMPAAP-Cu) of 1,5-dimethyl-4-(((1-methyl-1H-pyrrol-2-yl)methylene)amino)-2-phenyl-1,2-dihydro-3H-pyrazole-3-one (NMPAAP) was synthesized and analyzed by elemental analysis, magnetic susceptibility molar conductivity, thermal analysis, and defined spectral features using MS, IR, UV–vis, 1H, 13C NMR. In vitro biological activity of the complex was also investigated. Antimicrobial potency was evaluated by Disc Diffusion Method. Antioxidant activity was investigated by using DPPH radical scavenging and metal chelating activity tests. The complex exhibited a dose-dependent increase in its antioxidant capacity. The in vitro cytotoxic and anticancer potentials were evaluated in the human breast cancer cell line MCF-7. Data showed that NMPAAP-Cu significantly decreased the cell viability of MCF-7 cells in a concentration-dependent manner. AO/EB staining showed that the complex significantly increased apoptosis in MCF-7 cells. 50 µM NMPAAP-Cu also inhibited the migration of MCF-7 cells revealed in the wound healing assay. The study results indicated that the NMPAAP-Cu showed very high anticancer activity against MCF-7 carcinoma cells.

Long Non-Coding RNA LINC00052 Targets miR-548p/Notch2/Pyk2 to Modulate Tumor Budding and Metastasis of Human Breast Cancer.

Abnormal expression of long non-coding RNAs (lncRNAs) is involved in many pathological processes of cancers. However, the role of lncRNA LINC00052 in breast cancer progression is still unclear. Here, LINC00052 expression was detected by in situ hybridization and quantitative real-time PCR assays. Cell Counting Kit-8, wound healing, and transwell assays were used to investigate changes in the proliferation, migration, and invasion of breast cancer cells. MiR-548p was found associated with LINC00052 or Notch2 by RNA pull-down, dual-luciferase reporter, and qRT-PCR assays. The effect of LINC00052 on lung metastasis was explored through in vivo experiments. High LINC00052 expression was observed in breast cancer tissues and cells. LINC00052 silencing inhibited the proliferation, migration, and invasion of MCF7 cells, and LINC00052 overexpression produced the opposite results. MiR-548p, a target gene of LINC00052, partially rescued the effects of LINC00052 on proliferation, migration, and invasion of MCF7. Notch2 was the target of miR-548p and LINC00052 could promote Notch2 expression. Moreover, the phosphorylation of proline-rich tyrosine kinase 2 (Pyk2), a downstream factor of Notch2, was increased by LINC00052, and a Pyk2 mutant could inhibit the cell migration and invasion induced by LINC00052 overexpression in MDA-MB-468 cells, which was similar to the function of the miR-548p mimic. We further demonstrated that LINC00052 exacerbated the metastases of breast cancer cells in vivo. Our research demonstrated that LINC00052 is highly expressed in breast cancer and promotes breast cancer proliferation, migration, and invasion via the miR-548p/Notch2/Pyk2 axis. LINC00052 could serve as a potential therapeutic target for breast cancer.

Inhibition of cell invasion and migration by targeting matrix metalloproteinase-9 expression via sirtuin 6 silencing in human breast cancer cells

Sirtuin 6 (SIRT6) regulation is involved in carcinogenesis. However, its role in breast cancer (BC) metastasis remains unclear. We investigated the effects of SIRT6 on protein kinase C activator- and cytokine-mediated cancer cell invasion and migration in MCF-7 and MDA-MB-231 cells and the association between SIRT6 and matrix metalloproteinase-9 (MMP-9) expression. To assess MMP-9 and SIRT6 expression in patients, protein levels in BC tissues were analyzed. MCF-7 and MDA-MB-231 cell viability was analyzed using MTT assays. SIRT6 was silenced in both cell lines and protein secretion, expression, and mRNA levels were analyzed. Transcription factor DNA activity was investigated using luciferase assays. Matrigel invasion assays were used to assess the effects of SIRT6 in both cell lines. SIRT6 and MMP-9 expression in cancer tissues was significantly higher than in paired normal breast tissues. 12-O-tetradecanoylphorbol-13-acetate (TPA) or tumor necrosis factor-α (TNF-α) increased MMP-9 expression and cell invasion and migration, but SIRT6 knockdown abolished these effects. SIRT6 overexpression additively increased TPA- and TNF-α-induced MMP-9 expression. SIRT6 knockdown suppressed the mitogen-activated protein kinase (MAPK) signaling pathway and thus TPA- and TNF-α-induced MMP-9 expression. SIRT6 silencing suppressed TPA- and TNF-α-induced nuclear factor-κB (NF-κB) and activator protein-1 (AP-1) expressions in both cell lines, and treatment with MAPK, NF-κB, and AP-1 inhibitors reduced MMP-9 expression. The anti-invasive effects of SIRT6 in BC cells might be mediated by suppression of MAPK phosphorylation and reduction in NF-κB and AP-1 DNA activities, leading to MMP-9 downregulation, suggesting that SIRT6 modulation has the potential to target BC metastasis.