Colony Formation Of MCF-7 Cells Research Articles

Metabolic alterations and cellular responses to β-Hydroxybutyrate treatment in breast cancer cells

BackgroundThe ketogenic diet (KD), based on high fat (over 70% of daily calories), low carbohydrate, and adequate protein intake, has become popular due to its potential therapeutic benefits for several diseases including cancer. Under KD and starvation conditions, the lack of carbohydrates promotes the production of ketone bodies (KB) from fats by the liver as an alternative source of metabolic energy. KD and starvation may affect the metabolism in cancer cells, as well as tumor characteristics. The aim of this study is to evaluate the effect of KD conditions on a wide variety of aspects of breast cancer cells in vitro.MethodsUsing two cancer and one non-cancer breast cell line, we evaluate the effect of β-hydroxybutyrate (βHb) treatment on cell growth, survival, proliferation, colony formation, and migration. We also assess the effect of KB on metabolic profile of the cells. Using RNAseq analysis, we elucidate the effect of βHb on the gene expression profile.ResultsSignificant effects were observed following treatment by βHb which include effects on viability, proliferation, and colony formation of MCF7 cells, and different effects on colony formation of MDA-MB-231 cells, with no such effects on non-cancer HB2 cells. We found no changes in glucose intake or lactate output following βHb treatment as measured by LC-MS, but an increase in reactive oxygen species (ROS) level was detected. RNAseq analysis demonstrated significant changes in genes involved in lipid metabolism, cancer, and oxidative phosphorylation.ConclusionsBased on our results, we conclude that differential response of cancer cell lines to βHb treatment, as alternative energy source or signal to alter lipid metabolism and oncogenicity, supports the need for a personalized approach to breast cancer patient treatment.

Anti-tumor mechanism of total saponins of Paridis Rhizoma on inducing ferroptosis of breast cancer MCF-7 cells

This study aims to investigate the mechanism of total saponins of Paridis Rhizoma in inducing the ferroptosis of MCF-7 cells and provide a theoretical basis for the clinical treatment of breast cancer with total saponins of Paridis Rhizoma. The methyl thiazolyl tetrazolium(MTT) assay was employed to examine the effects of different concentrations of total saponins of Paridis Rhizoma on the proliferation of MCF-7 cells. A phase contrast inverted microscope was used to observe the morphological changes of MCF-7 cells. The colony formation assay was employed to test the colony formation of MCF-7 cells. The lactate dehydrogenase(LDH) release test was conducted to determine the cell membrane integrity of MCF-7 cells. The cell scratch assay was employed to examine the migration of MCF-7 cells. After that, the level of reactive oxygen species(ROS) in MCF-7 cells was observed by an inverted fluorescence microscope, and the content of Fe~(2+) in MCF-7 cells was detected by the corresponding kit. Transmission electron microscopy was employed to observe the mitochondrial ultrastructure of MCF-7 cells. Western blot was employed to determine the expression of ferroptosis-related proteins, such as p53, solute carrier family 7 member 11(SLC7A11), glutathione peroxidase 4(GPX4), acyl-CoA synthetase long-chain family member 4(ACSL4), and transferrin receptor protein 1(TFR1) in MCF-7 cells. The results showed that 1.5, 3, 4.5, 6, 7.5, and 9 μg·mL~(-1) total saponins of Paridis Rhizoma significantly inhibited the proliferation of MCF-7 cells, with the IC_(50) of 4.12 μg·mL~(-1). Total saponins of Paridis Rhizoma significantly damaged the morphology of MCF-7 cells, leading to the formation of vacuoles and the gradual shrinkage and detachment of cells. Meanwhile, total saponins of Paridis Rhizoma inhibited the colony formation of MCF-7 cells, destroyed the cell membrane(leading to the release of LDH), and shortened the migration distance of MCF-7 cells. Total saponins of Paridis Rhizoma treatment significantly increased the content of ROS, induced oxidative damage, and led to the accumulation of Fe~(2+) in MCF-7 cells. Furthermore, total saponins of Paridis Rhizoma changed the mitochondrial structure, increased the mitochondrial membrane density, led to the decrease or even disappear of ridges, promoted the expression of p53 protein, down-regulated the expression of SLC7A11 and GPX4, and up-regulated the expression of ACSL4 and TFR1. In summary, total saponins of Paridis Rhizoma can significantly inhibit the proliferation and migration of MCF-7 cells and destroy the cell structure by inducing ferroptosis.

Clerodendrum chinense Stem Extract and Nanoparticles: Effects on Proliferation, Colony Formation, Apoptosis Induction, Cell Cycle Arrest, and Mitochondrial Membrane Potential in Human Breast Adenocarcinoma Breast Cancer Cells.

Breast cancer stands out as the most widespread form of cancer globally. In this study, the anticancer activities of Clerodendrum chinense (C. chinense) stem ethanolic extract were investigated. High-performance liquid chromatography (HPLC) analysis identified verbascoside and isoverbascoside as the major bioactive compounds in the C. chinense stem extract. Successfully developed nanoparticles exhibited favorable hydrodynamic diameter, polydispersity index, and surface charge, thus ensuring stability after four months of storage. The total phenolic content and total flavonoid contents in the nanoparticles were reported as 88.62% and 95.26%, respectively. The C. chinense stem extract demonstrated a dose-dependent inhibitory effect on MCF-7, HeLa, A549, and SKOV-3 cancer cell lines, with IC50 values of 109.2, 155.6, 206.9, and 423 µg/mL, respectively. C. chinense extract and NPs exhibited dose-dependent cytotoxicity and the highest selectivity index values against MCF-7 cells. A dose-dependent reduction in the colony formation of MCF-7 cells was observed following treatment with the extract and nanoparticles. The extract induced cytotoxicity in MCF-7 cells through apoptosis and necrosis. C. chinense stem extract and nanoparticles decreased mitochondrial membrane potential (MMP) and induced G0/G1 phase arrest in MCF-7 cells. In conclusion, use of C. chinense stem extract and nanoparticles may serve as a potential therapeutic approach for breast cancer, thus warranting further exploration.

Design, synthesis, and biological evaluation of harmine derivatives as topoisomerase I inhibitors for cancer treatment

A series of β-carboline derivatives were designed and synthesized by introducing the chalcone moiety into the harmine. The synthesized derivatives were evaluated their anti-proliferative activities against six human cancer cell lines (MCF-7, MDA-MB-231, HepG2, HT29, A549, and PC-3) and one normal cell line (L02). Among them, compound G11 exhibited the potent anti-proliferative activity against MCF-7 cell line, with an IC50 value of 0.34 μM. Further biological studies revealed that compound G11 inhibited colony formation of MCF-7 cells, suppressed MCF-7 cell migration by downregulating migration-associated protein MMP-2. In addition, it could induce apoptosis of MCF-7 cells by downregulating Bcl-2 and upregulating Cleaved-PARP, Bax, and phosphorylated Bim proteins. Furthermore, compound G11 can act as a Topo I inhibitor, affecting DNA synthesis and transcription, thereby inhibiting cancer cell proliferation. Moreover, compound G11 inhibited tumor growth in 4T1 syngeneic transplant mice with an inhibition rate of 43.19 % at a dose of 10 mg/kg, and 63.87 % at 20 mg/kg, without causing significant toxicity to the mice or their organs, achieving the goal of reduced toxicity and increased efficacy. All these results indicate of G11 has enormous potential as an anti-tumor agent and merits further investigation.

3‐Methoxy‐Pyrrole‐Based Small Molecule Damages Mitochondria to Instigate Apoptosis in Breast Cancer Cells

AbstractMitochondrion has been implicated in the development and progression of breast cancer, making it an unconventional target for the anti‐cancer therapy. As a result, there is a need to explore novel small molecules for next generation breast cancer therapy. Towards this endeavour, herein, we have identified a novel 3‐methoxy‐pyrrole‐based small molecule from a synthetic library. Confocal fluorescence microscopy studies revealed that, this lead small molecule induced mitochondrial damage and generated reactive oxygen species (ROS) in the MCF7 breast cancer cells. Small molecule based mitochondrial impairment subsequently triggered cell cycle arrest and nuclear DNA damage followed by apoptosis leading remarkable cell killing and inhibition in the colony formation in MCF7 cells. Moreover, this 3‐methoxy‐pyrrole killed drug resistant MDA‐MB‐231 triple negative breast cancer cells and OVCAR8 ovarian cancer cells efficiently. This novel small molecule can open a new direction towards mitochondria targeted breast cancer therapy in future.

HERPUD1, a Member of the Endoplasmic Reticulum Protein Quality Control Mechanism, may be a Good Target for Suppressing Tumorigenesis in Breast Cancer Cells.

Breast cancer is the most frequently diagnosed cancer type and the second leading cause of cancer-related death in women. Recent studies have highlighted the importance of the endoplasmic reticulum (ER) protein quality control mechanism for the survival of many cancers. It has also been recommended as a good target for the treatment of many cancer types. Homocysteine inducible ER protein with ubiquitin-like domain 1 (HERPUD1) functions as one of the main components of ER-associated degradation, which is an ER-resident protein quality mechanism. Today, the association of HERPUD1 with breast carcinogenesis is still not fully understood. Herein, we evaluated the possibility of HERPUD1 as a potential therapeutic target for breast cancer. The effects of HERPUD1 silencing on epithelial-mesenchymal transition (EMT), angiogenesis, and cell cycle proteins were analyzed by immunoblotting studies. To test the role of HERPUD1 on tumorigenic features, WST-1-based cell proliferation assay, wound-healing assay, 2D colony formation assay, and Boyden-Chamber invasion assay were performed in human breast cancer cell line MCF-7. The statistical significance of the differences between the groups was determined by Student's t-test. Our results displayed that suppressing HERPUD1 expression reduced the cell cycle-related protein levels, including cyclin A2, cyclin B1, and cyclin E1 in MCF-7 cells. Also, silencing of HERPUD1 remarkably decreased expression levels of EMT-related N-cadherin and angiogenesis marker vascular endothelial growth factor A. Moreover, we determined that cell proliferation, migration, invasion, and colony formation of MCF-7 cells were significantly limited by silencing of HERPUD1. Present data suggest that HERPUD1 may be an effective target for biotechnological and pharmacological strategies to be developed to treat breast cancer.

β-catenin promotes resistance to trastuzumab in breast cancer cells through enhancing interaction between HER2 and SRC.

More than 1 million women worldwide are diagnosed with breast cancer (BC) each year. This study aims to explore the molecular mechanisms of β-catenin affecting the trastuzumab tolerance in HER2-positive BC. β-catenin in BC and non-BC tissue samples were assessed by immunohistochemistry. β-catenin and HER2 were over-expressed and knockdown to evaluate their role in tumorigenicity and trastuzumab resistance in cell and animal models using soft-agar and xenograft assays. Confocal laser immunofluorescence assay and co-immunoprecipitation were used to assess protein-protein binding. Expression of genes was detected using Western blot analysis. β-catenin was highly expressed in primary and metastatic BC, overexpression of β-catenin increased the colony formation of MCF7 cells when it was co-expressed with HER2 and synergically increased the tumor size in immunodeficient mice. Overexpression of β-catenin also increased the phosphorylation of HER2 and HER3 and increased the size of tumor derived from HER2-elevated cells. Confocal laser immunofluorescence assay showed that β-catenin and HER2 were co-localized on the membrane of MDA-MB-231 cells, suggesting that β-catenin binds HER2 to activate the HER2 signaling pathway. Immunoprecipitation of β-catenin and HER2 also confirmed this binding. On the other hand, knockdown of β-catenin in MDA-MB-231 cell lines decreased the activity of SRC and decreased phosphorylation of HER2 at Y877 and Y1248. The interaction between HER2 and SRC was enhanced when β-catenin was overexpressed, and β-catenin increased the resistance of tumor derived from HER2 elevated BT474 cells to trastuzumab. Further analysis showed that trastuzumab inhibited the activation of HER3, but SRC was still highly expressed in cells overexpressing β-catenin. Our work demonstrates that β-catenin is highly expressed in BC and it synergically promotes formation and progress of BC with HER2. β-catenin binds with HER2 leading to enhanced interaction with SRC and resistance to trastuzumab.

METTL3 stabilization by PIN1 promotes breast tumorigenesis via enhanced m6A-dependent translation.

Methyltransferase-like 3 (METTL3) is the catalytic subunit of the N6-adenosine methyltransferase complex responsible for N6-methyladenosine (m6A) modification of mRNA in mammalian cells. Although METTL3 expression is increased in several cancers, the regulatory mechanisms are unclear. We explored the regulatory roles of peptidyl-prolyl cis-trans isomerase NIMA-interacting 1 (PIN1) in METTL3 stability and m6A modification of mRNA. PIN1 interacted with METTL3 and prevented its ubiquitin-dependent proteasomal and lysosomal degradation. It stabilized METTL3, which increased the m6A modification of transcriptional coactivator with PDZ-binding motif (TAZ) and epidermal growth factor receptor (EGFR) mRNA, resulting in their efficient translation. PIN1 knockout altered the distribution of TAZ and EGFR mRNA from polysomes into monosomes. Inhibition of MEK1/2 kinases and PIN1 destabilized METTL3, which impeded breast cancer cell proliferation and induced cell cycle arrest at the G0/G1 phases. METTL3 knockout reduced PIN1 overexpression-induced colony formation in MCF7 cells and enhanced tumor growth in 4T1 cells in an orthotopic mouse model. In clinical settings, METTL3 expression significantly increased with tumor progression and was positively correlated with PIN1 expression in breast cancer tissues. Thus, PIN1 plays a regulatory role in mRNA translation, and the PIN1/METTL3 axis may be an alternative therapeutic target in breast cancer.

Taxol‑resistant breast cancer cell‑derived exosome‑delivered miR‑187‑5p regulates the growth of breast cancer cells via ABCD2 and Wnt/β‑catenin signaling.

Acquired resistance to Taxol (TAX) contributes to clinical treatment failure and significantly reduces the survival rate of patients. The present study aimed to explore the effects of exosomal microRNA (miR)-187-5p on TAX resistance in breast cancer cells and its underlying mechanisms. Exosomes were isolated from MCF-7 and TAX-resistant MCF-7/TAX cells, and the miR-187-5p and miR-106a-3p levels of the cells and exosomes were determined using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Next, MCF-7 cells were treated with TAX for 48 h and either treated with exosomes or transfected with miR-187-5p mimics. Cell viability, apoptosis, migration, invasion and colony formation were determined using Cell Counting Kit-8, flow cytometry, Transwell and colony formation assays, and the expression levels of associated genes and proteins were detected by RT-qPCR and western blotting, respectively. Finally, a dual-luciferase reporter gene assay was performed to confirm the target of miR-187-5p. The results showed that miR-187-5p expression levels increased significantly in TAX-resistant MCF-7 cells and exosomes compared with normal MCF-7 cells and exosomes (P<0.05). However, miR-106a-3p was not detected in the cells or exosomes. Therefore, miR-187-5p was selected for subsequent experiments. A series of cell assays showed that TAX inhibited the viability, migration, invasion and colony formation of MCF-7 cells and promoted their apoptosis; however, these changes were reversed by resistant cell exosomes and miR-187-5p mimics. Additionally, TAX significantly upregulated ABCD2 and downregulated β-catenin, c-Myc and cyclin D1, whereas resistant exosomes and miR-187-5p mimics reversed the TAX-induced changes in expression. Finally, ABCD2 was confirmed to directly bind with miR-187-5p. It may be concluded that TAX-resistant cell-derived exosomes delivering miR-187-5p may affect the growth of TAX-induced breast cancer cells by targeting ABCD2 and c-Myc/Wnt/β-catenin signaling.

Amlodipine inhibits proliferation, invasion, and colony formation of breast cancer cells.

Calcium channel upregulation has been implicated in cancer cell proliferation and progression including in breast cancer. Fortunately, the function of calcium channels can be manipulated pharmacologically using calcium channel blockers (CCBs). Amlodipine, a dihydropyridine CCB, has been demonstrated to exert cytotoxic effects in several types of cancers. The present study evaluated the effects of amlodipine on proliferation, caspase activation, colony formation, and invasion of human breast cancer cells. Cell viability was assessed using a colorimetric MTT assay. An Apo-ONE® caspase-3/7 assay was used to measure caspase-3/7 levels. Cell invasion was evaluated using Matrigel invasion chambers. The expression of phospho-(p-)ERK1/2, Bcl-2, and integrin β1 proteins were analyzed using western blotting. A one-way ANOVA with a post-hoc Tukey's multiple comparison tests was used for statistical analysis. Amlodipine significantly inhibited the growth of both MDA-MB-231 and MCF-7 human breast cancer cells in a dose-dependent manner and inhibited colony formation of MCF-7 cells, and this was accompanied by the downregulation of p-ERK1/2 in MDA-MB-231 cells. In addition, treatment with amlodipine resulted in increased caspase-3/7 levels in MDA-MB-231 cells, which was accompanied by the downregulation of the anti-apoptotic protein, Bcl-2. Moreover, amlodipine impaired the invasive abilities of MDA-MB-231 cells, and integrin β1 expression was concurrently downregulated. The present study illustrates the anticancer effects of amlodipine on breast cancer proliferation, colony formation, and invasion in vitro and highlights the potential value of amlodipine as an anticancer agent.

Synthesis of ponatinib analogues as novel inhibitors of leukemia stem cells.

Aim: To synthesize new analogues of ponatinib and evaluate anti-leukemia cells and cytotoxicity. Methodology & results: The inhibitory activity of compounds 13a and 13c against K562 and HL60 cells was comparable to that of ponatinib (IC50=0.74, 0.88 vs 0.64nM and 0.59, 0.77 vs 0.39nM, respectively). Compounds 13a and 40b were 34-and 77-fold more potent than ponatinib against KG1a cells (IC50=0.091 and 0040 vs 3.6μM, respectively). Compounds 13a, 13c and 40b also decreased the Abl protein level in the K562 cells, inhibited colony formation in MCF-7 cells and inhibited cell migration in B16BL6 cells. Compound 13a showed low cytotoxicity in 293 cells. Conclusion: Compound 13a was the best lead compound.

Ursolic Acid Decreases the Proliferation of MCF-7 Cell-Derived Breast Cancer Stem-Like Cells by Modulating the ERK and PI3K/AKT Signaling Pathways

Cancer stem cells are strong drivers of metastasis and cancer relapse, which makes them important therapeutic targets. Ursolic acid (UA), a pentacyclic triterpenoid, has anticancer effects in various types of cancer; however, little is known about its effect on the growth of MCF-7 cell-derived breast cancer stem (BCS)-like cells in estrogen receptor positive breast cancer. In this study, the anticancer activity of UA in MCF-7 cell-derived BCS-like cells and its mechanism of action were evaluated. Furthermore, its inhibitory effects on the proliferation of MCF-7 cell-derived BCS-like cells were compared with that on MCF-7 cells. In MCF-7 cells, UA increased p53 and p21 expression but decreased cyclin D, cyclin E, CDK4, and CDK2 expression to induce cell cycle arrest in the G0/G1 phase. Moreover, UA significantly suppressed migration, invasion, and colony formation in MCF-7 cells, and suppressed mammosphere formation in a concentration- dependent manner. In MCF-7 cell-derived BCS-like cells, UA significantly decreased migration, suppressed p-PI3K, p-AKT, and p-ERK expression, and enhanced p-FoxO1/FoxO3a expression. Accordingly, in MCF-7 cell-derived BCS-like cells, UA suppressed proliferation in part by downregulating ERK and PI3K/AKT signaling pathways. These findings provide the first evidence for the selective effects of UA in BCSs.

Quercetin potentiates the chemosensitivity of MCF-7 breast cancer cells to 5-fluorouracil.

Breast cancer is one of the leading causes of cancer mortality worldwide. 5-fluorouracil (5-FU) is one of the chemotherapy drugs to treat breast cancer, but it is associated with several side effects. Combination therapy is a way to increase the effectiveness of chemo drugs and decrease their usage dose. Quercetin (Quer) is one of the natural polyphenols with anti-cancer properties. This study investigated the apoptotic effect of 5-FU in combination with Quer compared with 5-FU alone on MCF-7 breast cancer cells. Different single and combined concentrations of 5-FU and Quer were applied to MCF 7 cells for 48h. Cell viability, apoptosis, gene expression of Bax, Bcl2, and p53, caspase activity, and colony number were assessed using MTT assay, flow cytometry, quantitative real-time PCR, enzyme-linked immunosorbent (ELISA), and Colony formation assay, respectively. The combination of 5-FU and Quer compared to 5-FU alone improved apoptosis by increasing the gene expression of Bax and p53 and caspase-9 activity and decreasing the Bcl2 gene expression. Colony formation in MCF-7 cells significantly decreased in the combined state compared to 5-FU alone. Quer potentiates the sensitivity of breast cancer to 5-FU so that this combination may be proposed as a treatment for breast cancer. Therefore, this combination can be suggested for future in vivo studies.

Synthesis of novel sulfonamide derivatives containing pyridin-3-ylmethyl 4-(benzoyl)piperazine-1-carbodithioate moiety as potent PKM2 activators

Pyruvate kinase M2 isoform (PKM2) plays a key role in cancer progression through both metabolic and non-metabolic functions, thus it is recognized as a potential target for cancer diagnosis and treatment. In this study, we discovered a sulfonamide-dithiocarbamate compound 8a as a novel PKM2 activator from a random screening of an in-house compound library. Then, a series of lead compound 8a analogs were designed and synthesized for screening as potent PKM2 activators. Among them, compound 8b (AC50 = 0.136 µM) and 8k (AC50 = 0.056 µM) showed higher PKM2 activation activities than positive control NZT (AC50 = 0.228 µM), and they (IC50 < 1 µM) exhibited more significant anti-proliferative activities against human tumor cell lines than NZT (IC50 > 10 µM). Especially, compound 8k inhibited the proliferation of multiple cancer cells, but showed little toxicity on normal cells. In addition, we found that compound 8k inhibit the colony formation of MCF7 cells. Western blot analysis demonstrated that 8k could reduce PKM2 nuclear localization and block the downstream signaling pathway of PKM2, resulting in suppression of tumor cell proliferation. Overall, compound 8k may be a promising candidate for further mechanistic investigation of PKM2 and cancer therapy.

Comparative Study of Lipophilic Statin Activity in 2D and 3Din vitro Models of Human Breast Cancer Cell Lines MDA-MB-231 and MCF-7.

IntroductionStatins are a type of drugs that are used to lower cholesterol level in blood. Since the early 1990s, it has been known that statins could be beneficial in cancer therapy. However, data remain controversial, especially regarding estrogen receptors status. Despite many studies in breast cancer models in vitro, the correlations of effects of separate statins in various model systems remain unclear.AimOur aim was to evaluate the anticancer activity of lovastatin, mevastatin, pitavastatin and simvastatin on different subtypes of human breast cancer (MDA-MB-231 and MCF-7 cell lines) in spatially different 2D and 3D cultures in vitro.Materials and MethodsCell viability was tested using MTT assay. Effect of statins on cell colony formation was evaluated by calculating breast cancer cell colony area and number. The effect on cell migration was estimated by “wound healing” assay. The activity of compounds in 3D cultures was evaluated by measuring the spheroid size changes during incubation.ResultsAmong the tested statins, pitavastatin had the greatest effect on the viability of breast cancer MDA-MB-231 and MCF-7 cell lines. The mevastatin and pitavastatin mostly decreased the MDA-MB-231 and MCF-7 cell colony formation. All statins at 90% of their estimated effective concentration (EC50) and simvastatin at 10% of its EC50 concentration suppressed the MCF-7 cells migration in “wound healing” assay. Only higher concentrations of mevastatin and pitavastatin slowed down the MDA-MB-231 cell migration. Statins showed different activity on 3D cell cultures growth. Lovastatin and simvastatin delayed the growth of MDA-MB-231 cell spheroids, while mevastatin and pitavastatin reduced the growth of MCF-7 spheroids.ConclusionStatins possess different anticancer activity in human breast cancer MDA-MB-231 and MCF-7 cell cultures. Pitavastatin and simvastatin showed the highest activity in most tested assays, especially against MCF-7 cell line.

Tryptanthrin exerts anti-breast cancer effects both in vitro and in vivo through modulating the inflammatory tumor microenvironment.

Tryptanthrin is an indole quinazoline alkaloid from the indigo-bearing plants, such as Isatis indigotica Fort. Typically, this natural compound shows a variety of pharmacological activities such as antitumor, antibacterial, anti-inflammatory and antioxidant effects. This study was conducted to assess the antitumor activity of tryptanthrin in breast cancer models both in vitro and in vivo, and to explore the important role of the inflammatory tumor microenvironment (TME) in the antitumor effects of tryptanthrin. Human breast adenocarcinoma MCF-7 cells were used to assess the antitumor effect of tryptanthrin in vitro. MTT assay and colony formation assay were carried out to monitor the antiproliferative effect of tryptanthrin (1.56~50.0 μmol L-1) on inhibiting the proliferation and colony formation of MCF-7 cells, respectively. The migration and invasion of MCF-7 cells were evaluated by wound healing assay and Transwell chamber assay, respectively. Moreover, the 4T1 murine breast cancer model was established to examine the pharmacological activity of tryptanthrin, and three groups with different doses of tryptanthrin (25, 50 and 100 mg kg-1) were set in study. Additionally, tumor volumes and organ coefficients were measured and calculated. After two weeks of tryptanthrin treatment, samples from serum, tumor tissue and different organs from tumor-bearing mice were collected, and the enzyme-linked immunosorbent assay (ELISA) was performed to assess the regulation of inflammatory molecules in mouse serum. Additionally, pathological examinations of tumor tissues and organs from mice were evaluated through hematoxylin and eosin (H&E) staining. The expression of inflammatory proteins in tumor tissues was measured by immunohistochemistry (IHC) and Western blotting. Tryptanthrin inhibited the proliferation, migration and invasion of MCF-7 cells, up-regulated the protein level of E-cadherin, and down-regulated those of MMP-2 and Snail, as suggested by the MCF-7 cell experiment. According to the results from in vivo experiment, tryptanthrin was effective in inhibiting tumor growth, and it showed favorable safety without inducing the fluctuations of body mass and organ coefficient (p > 0.05). In addition, tryptanthrin also suppressed the expression levels of NOS1, COX-2 and NF-κB in mouse tumor tissues, and regulated those of IL-2, IL-10 and TNF-α in the serum of tumor cells-transplanted mice. Tryptanthrin exerted its anti-breast cancer activities through modulating the inflammatory TME both in vitro and in vivo.

In Vivo and In Vitro Evaluation of the Protective Effects of Hesperidin in Lipopolysaccharide-Induced Inflammation and Cytotoxicity of Cell

(1) Background: Plant flavonoids are efficient in preventing and treating various diseases. This study aimed to evaluate the ability of hesperidin, a flavonoid found in citrus fruits, in inhibiting lipopolysaccharide (LPS) induced inflammation, which induced lethal toxicity in vivo, and to evaluate its importance as an antitumor agent in breast cancer. The in vivo experiments revealed the protective effects of hesperidin against the negative LPS effects on the liver and spleen of male mice. (2) Methods: In the liver, the antioxidant activity was measured by estimating the concentration of glutathione (GSH) and catalase (CAT), whereas in spleen, the concentration of cytokines including IL-33 and TNF-α was measured. The in vitro experiments including MTT assay, clonogenity test, and sulforhodamine 101 stain with DAPI (4′, 6-diamidino-2-phenylindole) were used to assess the morphological apoptosis in breast cancer cells. (3) Results: The results of this study revealed a significant increase in the IL-33 and TNF-α cytokine levels in LPS challenged mice along with a considerable elevation in glutathione (GSH); moreover, the catalase (CAT) level was higher compared to that of the control group. Cytotoxicity of the MCF-7 cell line revealed significant differences among the groups treated with different concentrations when compared to the control groups, in a concentration-dependent manner. Hesperidin significantly inhibited the colony formation of MCF7 cells when compared to that of control. Clear changes were observed in the cell shape, including cell shrinkage and chromatin condensation, which were associated with a later apoptotic stage. (4) Conclusion: The results indicate that hesperidin might be a potential candidate in preventing diseases.

Structure, antiproliferative and cancer preventive properties of sulfated α-d-fucan from the marine bacterium Vadicella arenosi

Sulfated fucose-containing glycopolymers are currently of great interest because of their wide spectrum of bioactivity, including anti-tumor properties. In this study, the structure of O-polysaccharide (OPS) of the marine bacterium Vadicella arenosi KMM 9024T, its effect on the proliferation of human breast cancer MCF-7 cells and cancer preventive properties were investigated. Two OPS fractions with different molecular weights were isolated and purified from the lipopolysaccharide by mild acid hydrolysis followed by anion-exchange chromatography. The OPS was found to consist of α-(1→3)-linked 2-O-sulfate-d-fucopyranosyl residues, whose structure was deduced by sugar analysis along with 2D NMR spectroscopy. The biological assay indicated that polysaccharide significantly reduced the proliferation and inhibited colony formation of MCF-7 cells in a dose-dependent manner. Besides, the experiment indicated the inhibitory role of polysaccharide on EGF-induced neoplastic cell transformation in mouse epidermal cells. The investigated polysaccharide is the first sulfated fucan isolated from the bacteria.

2-Pyridineformamide N(4)-ring incorporated thiosemicarbazones inhibit MCF-7 cells by inhibiting JNK pathway.

In an effort to develop a more potent anticancer therapeutic agent, a series of 2-pyridineformamide thiosemicarbazones (R = H, 4-CH3, 5-F, 6-CH3 and ) have been synthesized and evaluated for their anti-cancer activities against the cancer cells MCF-7 (breast cancer cell line), A-431 and A375 (epidermoid carcinoma cell line), and HeLa (cervical cancer cell line) using MTT assay. All these 2-pyridineformamide thiosemicarbazones exhibited anti-proliferative activities towards these cell lines. 5FAmPyrr possess most profound effects against MCF-7 cells with IC50 of 0.9 µM. In flow cytometry using Propidium Iodide, 5FAmPyrr was found to induce cell death significantly in a dose dependent manner (100 nM-3 µM) and inhibited colony formation of MCF-7 cells. This compound induced pro-apoptotic protein Bax and inhibited anti apoptotic protein Bcl-2 as well as both c-Jun and Jun N-terminal kinase (abbreviated as JNK) in concentration dependent manner. Further pro-caspase 3 and PARP were inhibited by 5FAmPyrr at concentration of 3 µM. The results suggest that 5FAmPyrr exhibit anticancer potency and induced cell death by inhibiting MAPK signaling and inducing intrinsic apoptotic pathway. All these indicate that 2-pyridineformamide thiosemicarbazones could be developed as future therapeutics agents to treat cancer.

Knockdown of Rab7a suppresses the proliferation, migration, and xenograft tumor growth of breast cancer cells.

Breast cancer is a common invasive cancer in women. Ras-related protein Rab-7a (Rab7a) is involved in late endocytic trafficking, while its role in breast cancer is largely unclear. In the present study, we investigated the role of Rab7a in breast cancer. Comparing with adjacent breast tissues, Rab7a expression was increased in breast cancer tissues. Using lentivirus-mediated knockdown strategy, we found that Rab7a silencing inhibited the proliferation and colony formation of MDA-MB-231 cells. Apoptosis and G2 cell cycle arrest were induced in Rab7a knockdown. By contrast, Rab7a suppressed the apoptosis and promoted proliferation and colony formation of MCF-7 cells. The migration of MDA-MB-231 cells was suppressed by Rab7a knockdown. In vivo, depletion of Rab7a inhibited the xenograft tumor development of MDA-MB-231 cells. Altogether, our results highlight the novel function of Rab7a in the proliferation, invasion, and xenograft tumor development of breast cancer cells.