Mammary Adenocarcinoma Cell Line Research Articles

Proteomic Characterization of a 3D HER2+ Breast Cancer Model Reveals the Role of Mitochondrial Complex I in Acquired Resistance to Trastuzumab.

HER2-targeted therapies, such as Trastuzumab (Tz), have significantly improved the clinical outcomes for patients with HER2+ breast cancer (BC). However, treatment resistance remains a major obstacle. To elucidate functional and metabolic changes associated with acquired resistance, we characterized protein profiles of BC Tz-responder spheroids (RSs) and non-responder spheroids (nRSs) by a proteomic approach. Three-dimensional cultures were generated from the HER2+ human mammary adenocarcinoma cell line BT-474 and a derived resistant cell line. Before and after a 15-day Tz treatment, samples of each condition were collected and analyzed by liquid chromatography-mass spectrometry. The analysis of differentially expressed proteins exhibited the deregulation of energetic metabolism and mitochondrial pathways. A down-regulation of carbohydrate metabolism and up-regulation of mitochondria organization proteins, the tricarboxylic acid cycle, and oxidative phosphorylation, were observed in nRSs. Of note, Complex I-related proteins were increased in this condition and the inhibition by metformin highlighted that their activity is necessary for nRS survival. Furthermore, a correlation analysis showed that overexpression of Complex I proteins NDUFA10 and NDUFS2 was associated with high clinical risk and worse survival for HER2+ BC patients. In conclusion, the non-responder phenotype identified here provides a signature of proteins and related pathways that could lead to therapeutic biomarker investigation.

Phytochemical profile and determination of cytotoxicity, acute oral toxicity, genotoxicity, and mutagenicity of aqueous and ethanolic extracts of Pseudobombax marginatum (A. St.-Hil.) A. Robyns

ABSTRACT Pseudobombax marginatum, popularly known as “embiratanha,” is widely used by traditional communities as anti-inflammatory and analgesic agent. This study aimed to determine the phytochemical profile as well as cytotoxicity, acute oral toxicity, genotoxicity, and mutagenicity attributed to exposure to aqueous (AqEx) and ethanolic (EtEx) extracts of embiratanha bark. Phytochemical screening was conducted using thin-layer chromatography (TLC). Cell viability was analyzed using MTT assay with human mammary gland adenocarcinoma (MDA-MB-231) and macrophage (J774A.1) cell lines, exposed to concentrations of 12.5, 25, 50, or 100 µg/ml of either extract. For acute oral toxicity, comet assay and micronucleus (MN) tests, a single dose of 2,000 mg/kg of either extract was administered orally to Wistar rats. TLC analysis identified classes of metabolites in the extracts, including cinnamic acid derivatives, flavonoids, hydrolyzable tannins, condensed tannins, coumarins, and terpenes/steroids. In the cytotoxicity assay, the varying concentrations of extracts derived from embiratanha induced no significant alterations in the viability of MDA-MB-231 cells. The lowest concentration of EtEx significantly increased macrophage J774A.1 viability. However, the higher concentrations of AqEx markedly lowered macrophage J774A.1 viability. Animals exhibited no toxicity in the parameters analyzed in acute oral toxicity, comet assay, and MN tests. Further, EtEx promoted a significant reduction in DNA damage index and DNA damage frequency utilizing the comet assay, while the group treated with AqEx exhibited no marked differences. Thus, data demonstrated that AqEx or EtEx of embiratanha may be considered safe at a dose of 2,000 mg/kg orgally under our experimental conditions tested.

Targeting canine mammary neoplastic epithelial cells with a reengineered anthrax toxin: first study.

Mammary tumors are the most frequent type of neoplasms in intact female dogs. New therapies that target neoplastic cells without affecting normal cells are highly sought. The Bacillus anthracis toxin has been reengineered to target tumor cells that express urokinase plasminogen activators and metalloproteinases. In previous studies carried out in our laboratory, the reengineered anthrax toxin had inhibitory effects on canine oral mucosal melanoma and canine osteosarcoma cells. In this study, five canine neoplastic epithelial cell lines (four adenocarcinomas and one adenoma) and one non-neoplastic canine mammary epithelial cell line were treated with different concentrations of reengineered anthrax toxin components. Cell viability was quantified using an MTT assay and half-maximal inhibitory concentration (IC50) values. Cell lines were considered sensitive when the IC50 was lower than 5000ng/ml. One canine mammary adenocarcinoma cell line and one mammary adenoma cell line showed significantly decreased viability after treatment, whereas the non-neoplastic cell line was resistant. We conclude that the reengineered anthrax toxin may be considered a targeted therapy for canine mammary neoplasms while preserving normal canine mammary epithelial cells.

Inhibitory Effects of Alpha-Connexin Carboxyl-Terminal Peptide on Canine Mammary Epithelial Cells: A Study on Benign and Malignant Phenotypes.

Mammary cancer is highly prevalent in non-castrated female dogs. Cell-to-cell communication is an important mechanism to maintain homeostasis, and connexins are proteins that assemble to form the communicating gap junctions. In many cancers, communication capacity is reduced; several approaches are being tested in order to increase the communication capacity in cancer cells and, therefore, alter their viability. This study analyzed the effects of the alpha-connexin carboxyl-terminal peptide (αCT1) on canine mammary non-neoplastic and neoplastic epithelial cells. Seven canine epithelial mammary cell lines were used. Among these, one was a normal canine epithelial mammary cell line (LOEC-NMG), two canine mammary adenomas (LOEC-MAd1 and LOEC-MAd2), and four canine mammary adenocarcinomas (LOEC-MCA1, LOEC-MCA2, LOEC-MCA3 and CF41). The αCT1 corresponds to a short Cx43 C-terminal sequence linked to an internalization sequence called the antennapedia. After 24 h of incubation, the medium containing different αCT1 peptide concentrations was added to the cells, and only the culture medium was used for control. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test was used to quantify cell viability before treatment and 48, 72, and 96 h after the treatment. Results showed that the normal mammary epithelial cell line (LOEC-NMG) was resistant to treatment with αCT1, which is consistent with a previous study on human mammary cell lines. One of the adenoma cell lines (LOEC-MAd2) was also resistant to treatment with αCT1, although the other (LOEC-MAd1) was susceptible to treatment, mostly at 72 h after treatment. Regarding the four canine adenocarcinoma cell lines, they differ regarding the susceptibility to the treatment with αCT1. Three cell lines, canine mixed adenocarcinoma (LOEC-MCA1), canine complex adenocarcinoma (LOEC-MCA2), and commercial canine mammary adenocarcinoma cell line CF41, were susceptible to treatment with αCT1, while one canine mammary adenocarcinoma cell line (LOEC-MCA3) was resistant to treatment. In most αCT1 treated cell lines, Cx43 was strongly detected in cell membranes by immunofluorescence. We propose that αCT1 restored the cell-to-cell communication capacity of neoplastic cells and induced inhibitory effects on cell viability.

Propranolol inhibits EMT and metastasis in breast cancer through miR-499-5p-mediated Sox6

PurposeThis study will focus on 4T1 cells, a murine mammary adenocarcinoma cell line, as the primary research subject. We aim to investigate the inhibitory effects and mechanisms of propranolol on epithelial–mesenchymal transition (EMT) in breast cancer cells, aiming to elucidate this phenomenon at the miRNA level.MethodsIn this study, the EMT inhibitory effect of propranolol was observed through in vitro and animal experiments. For the screening of potential target miRNAs and downstream target genes, second-generation sequencing (SGS) and bioinformatics analysis were conducted. Following the screening process, the identified target miRNAs and their respective target genes were confirmed using various experimental methods. To confirm the target miRNAs and target genes, Western Blot (WB), reverse transcription polymerase chain reaction (RT-PCR), and immunofluorescence experiments were performed.ResultsIn this study, we found that propranolol significantly reduced lung metastasis in 4T1 murine breast cancer cells (p < 0.05). In vitro and in vivo experiments demonstrated that propranolol inhibited the epithelial–mesenchymal transition (EMT) as evidenced by Western Blot analysis (p < 0.05). Through next-generation sequencing (SGS), subsequent bioinformatics analysis, and PCR validation, we identified a marked downregulation of miR-499-5p (p < 0.05), suggesting its potential involvement in mediating the suppressive effects of propranolol on EMT. Overexpression of miR-499-5p promoted EMT, migration, and invasion of 4T1 cells, and these effects were not reversed or attenuated by propranolol (Validated via Western Blot, wound healing assay, transwell migration, and invasion assays, p < 0.05). Sox6 was identified as a functional target of miR-499-5p, with its downregulation correlating with the observed EMT changes (p < 0.05). Silencing Sox6 or overexpressing miR-499-5p inhibited Sox6 expression, further promoting the processes of EMT, invasion, and migration in 4T1 cells. Notably, these effects were not alleviated by propranolol (validated via Western Blot, wound healing assay, transwell migration, and invasion assays, p < 0.05). The direct interaction between miR-499-5p and Sox6 mRNA was confirmed by dual-luciferase reporter gene assay.ConclusionThese results suggest that propranolol may have potential as a therapeutic agent for breast cancer treatment by targeting EMT and its regulatory mechanisms.

Effects of 6-Mercaptopurine and its Derivatives on Human Hepatocellular Carcinoma and Mammary Adenocarcinoma Cell Lines

Effects of 6-Mercaptopurine and its Derivatives on Human Hepatocellular Carcinoma and Mammary Adenocarcinoma Cell Lines

New Organotin (IV) Compounds Derived from Dehydroacetic Acid and Thiosemicarbazides: Synthesis, Rational Design, Cytotoxic Evaluation, and Molecular Docking Simulation.

Organotin complexes were prepared through a one-pot reaction with three components by reacting thiosemicarbazide or 4-methyl-3-thiosemicarbazide or 4-phenylthiosemicarbazide, dehydroacetic acid (DHA) and dibutyl, diphenyl, dicyclohexyl, and bis[(trimethylsilyl)methyl]tin(IV) oxides; all complexes were characterized by infrared (IR), ultraviolet-visible (UV-vis), mass spectrometry (MS), and nuclear magnetic resonance (NMR) spectroscopy. The 119Sn NMR revealed chemical shifts corresponding to a pentacoordinated environment in solution. The X-ray crystallography of the two complexes evidenced the formation of monomeric complexes with a pentacoordinated geometry around tin via three donor atoms from the ligand, the sulfur of the thiol, the nitrogen of the imine group, and the oxygen of the pyran ring. The geometries of the five-coordinated complexes 3a (Bu2SnL3), 3c (Ph2SnL3), and 3d (Cy2SnL3) acid were intermediate between square pyramidal and trigonal bipyramidal, and complex 1a (Bu2SnL1) adopted a bipyramidal trigonal geometry (BPT). The sulforhodamine B assay assessed the cytotoxicity of organotin(IV) complexes against the MDA-MB-231 and MCF-7 (human mammary adenocarcinoma) cell lines and one normal COS-7 (African green monkey kidney fibroblast). The IC50 values evidenced a significant antiproliferative effect on cancer cells; the complexes were more potent than the positive cisplatin control and the corresponding ligands, dehydroacetic acid thiosemicarbazone (L1), dehydroacetic acid-N(4)-methylthiosemicarbazone (L2), and dehydroacetic acid-N(4)-phenylthiosemicarbazone (L3). The IC50 values also indicated that the organotin(IV) complexes were more cytotoxic against the triple-negative breast cell line MDA-MB-231 than MCF-7, inducing significant morphological alterations. The interactions of organotin(IV) 1c (Ph2SnL1), 1d (Cy2SnL1), and 1e (((CH3)3SiCH2)2SnL1) were evaluated with ss-DNA by fluorescence; intensity changes of the fluorescence were indicative of the displacement of ethidium bromide (EB), confirming the interaction of the organotin(IV) complexes with ss-DNA; the results showed a DNA binding affinity. The thermodynamic parameters obtained through isothermal titration calorimetry showed that the interaction of 1c (Ph2SnL1), with ss-ADN, was exothermic. Molecular docking studies also demonstrated that the organotin(IV) complexes were intercalated in DNA by conventional hydrogen bonds, carbon-hydrogen bonds, and π-alkyl interactions. These complexes furthermore showed a greater affinity towards DNA than cisplatin.

Molecular Resonance Imaging of the CAIX Expression in Mouse Mammary Adenocarcinoma Cells.

The carbonic anhydrase isoform IX (hCAIX) is one of the main players in extracellular tumor pH regulation, and it is known to be overexpressed in breast cancer and other common tumors. hCA IX supports the growth and survival of tumor cells, and its expression is correlated with metastasis and resistance to therapies, making it an interesting biomarker for diagnosis and therapy. The aim of this work deals with the development of an MRI imaging probe able to target the extracellular non-catalytic proteoglycan-like (PG) domain of CAIX. For this purpose, a specific nanoprobe, LIP_PepC, was designed by conjugating a peptidic interactor of the PG domain on the surface of a liposome loaded with Gd-bearing contrast agents. A Mouse Mammary Adenocarcinoma Cell Line (TS/A) was chosen as an in vitro breast cancer model to test the developed probe. MRI results showed a high selectivity and sensitivity of the imaging probe toward hCAI-expressing TS/A cells. This approach appears highly promising for the in vivo translation of a diagnostic procedure based on the targeting of hCA IX enzyme expression.

Schizophrenia plausible protective effect of microRNA-137 is potentially related to estrogen and prolactin in female patients.

Schizophrenia (SCZ) is a serious chronic mental disorder. Our previous case-control genetic association study has shown that microRNA-137 (miR-137) may only protect females against SCZ. Since estrogen, an important female sex hormone, exerts neuroprotective effects, the relationship between estrogen and miR-137 in the pathophysiology of SCZ was further studied in this study. Genotyping of single-nucleotide polymorphism rs1625579 of miR-137 gene in 1,004 SCZ patients and 896 healthy controls was conducted using the iMLDR assay. The effect of estradiol (E2) on the miR-137 expression was evaluated on the human mammary adenocarcinoma cell line (MCF-7) and the mouse hippocampal neuron cell line (HT22). The relationships between serum E2, prolactin (PRL), and peripheral blood miR-137 were investigated in 41 SCZ patients and 43 healthy controls. The miR-137 and other reference miRNAs were detected by real-time fluorescent quantitative reverse transcription-PCR. Based on the well-known SNP rs1625579, the distributions of protective genotypes and alleles of the miR-137 gene were not different between patients and healthy controls but were marginally significantly lower in female patients. E2 upregulated the expression of miR-137 to 2.83 and 1.81 times in MCF-7 and HT22 cells, respectively. Both serum E2 and blood miR-137 were significantly decreased or downregulated in SCZ patients, but they lacked expected positive correlations with each other in both patients and controls. When stratified by sex, blood miR-137 was negatively correlated with serum E2 in female patients. On the other hand, serum PRL was significantly increased in SCZ patients, and the female patients had the highest serum PRL level and a negative correlation between serum PRL and blood miR-137. The plausible SCZ-protective effect of miR-137 may be female specific, of which the underlying mechanism may be that E2 upregulates the expression of miR-137. This protective mechanism may also be abrogated by elevated PRL in female patients. These preliminary findings suggest a new genetic/environmental interaction mechanism for E2/miR-137 to protect normal females against SCZ and a novel E2/PRL/miR-137-related pathophysiology of female SCZ, implying some new antipsychotic ways for female patients in future.

Evaluation of coumarin-tagged deferoxamine as a Zr(IV)-based PET/fluorescence dual imaging probe

Desferoxamine (DFO) is currently the golden standard chelator for 89Zr4+, a promising nuclide for positron emission tomography imaging (PET). The natural siderophore DFO had previously been conjugated with fluorophores to obtain Fe(III) sensing molecules. In this study, a fluorescent coumarin derivative of DFO (DFOC) has been prepared and characterized (potentiometry, UV–Vis spectroscopy) for what concerns its protonation and metal coordination properties towards PET-relevant ions (Cu(II), Zr(IV)), evidencing strong similarity with pristine DFO. Retention of DFOC fluorescence emission upon metal binding has been checked (fluorescence spectrophotometry), as it would – and does – allow for optical (fluorescent) imaging, thus unlocking bimodal (PET/fluorescence) imaging for 89Zr(IV) tracers. Crystal violet and MTT assays on NIH-3 T3 fibroblasts and MDA-MB 231 mammary adenocarcinoma cell lines demonstrated, respectively, no cytotoxicity nor metabolic impairment at usual radiodiagnostic concentrations of ZrDFOC. Clonogenic colony-forming assay performed on X-irradiated MDA-MB 231 cells showed no interference of ZrDFOC with radiosensitivity. Morphological biodistribution (confocal fluorescence, transmission electron microscopy) assays on the same cells suggested internalization of the complex through endocytosis. Overall, these results support fluorophore-tagged DFO as a suitable option to achieve dual imaging (PET/fluorescence) probes based on 89Zr.

Molecular characterization of feline immune checkpoint molecules and establishment of PD-L1 immunohistochemistry for feline tumors.

Spontaneous tumors are a major cause of death in cats. Treatment of human tumors has progressed dramatically in the past decade, partly due to the success of immunotherapies using immune checkpoint inhibitors, such as anti-programmed death 1 (PD-1) and anti-PD-ligand 1 (PD-L1) antibodies. However, little is known about the PD-1 pathway and its association with tumor disease in cats. This study investigated the applicability of anti-PD-1/PD-L1 therapy in feline tumors. We first determined the complete coding sequence of feline PD-L1 and PD-L2, and found that the deduced amino acid sequences of feline PD-L1/PD-L2 share high sequence identities (66-83%) with orthologs in other mammalian species. We prepared recombinant feline PD-1, PD-L1, and PD-L2 proteins and confirmed receptor-ligand binding between PD-1 and PD-L1/PD-L2 using flow cytometry. Next, we established an anti-feline PD-L1 monoclonal antibody (clone CL1Mab-7) to analyze the expression of PD-L1. Flow cytometry using CL1Mab-7 revealed the cell surface expression of PD-L1 in a feline macrophage (Fcwf-4) and five mammary adenocarcinoma cell lines (FKNp, FMCm, FYMp, FONp, and FONm), and showed that PD-L1 expression was upregulated by interferon-γ stimulation. Finally, immunohistochemistry using CL1Mab-7 also showed PD-L1 expression in feline squamous cell carcinoma (5/5, 100%), mammary adenocarcinoma (4/5, 80%), fibrosarcoma (5/5, 100%), and renal cell carcinoma (2/2, 100%) tissues. Our results strongly encourage further investigations of the PD-1/PD-L1 pathway as a potential therapeutic target for feline tumors.

Cytotoxic effects of Curcuma longa leaves on MCF-7 and HepG2 cells

Curcuma longa (tumeric) is listed in several therapeutic regimens used in the management of cancer. The present study investigated the potential cytotoxic effects of ethanol extract of Curcuma longa (turmeric) leaves on HepG2 (a human hepatocellular carcinoma cell line), and MCF-7 (a human mammary gland adenocarcinoma cell line) cells. The HepG2 and MCF-7 cells were exposed to different concentrations; 2, 0.2, 0.02, 0.002, 0.0002, 0.00002, 0.000002 μg/ml, of the ethanol extract of C. longa leaves. The Water-Soluble tetrazolium salt (WST-1) assay was used to estimate the cell viability of the cells after which the percentage proliferation was computed. The results of this study indicated that C. longa had cytotoxic effect on the different cell lines, caused a decrease in proliferation and it is more selective to breast cancer cell lines based on the results of WST-1 analysis on the HepG2 and MCF-7 cells. These results suggest that C. longa leaf possesses cytotoxic properties, which can be exploited to develop novel cytotoxic drugs for cancer management. Keywords: Curcuma longa, turmeric, HepG2, MCF-7, WST-1, proliferation, cytotoxic, cancer

Effect Of Shikimic Acid And Acetylsalicylic Acid On Biofilm Formation In Staphylococcus Epidermidis

The aim of the current study was to evaluate the activity of Shikimic acid (SA) and Acetylsalicylic acid (ASA) against S. epidermidis biofilms formation in different conditions. A total of (231) samples were collected from patients who attended different hospitals in Ramadi and Faluja cities during the period from December(2020) to the end of July (2021). Samples were cultured on blood, and manitol salt agar and identification was performed by the VITEK 2 system. Antibiofilm activity of SA was evaluated by two methods, modified crystal violet, and MTT assay. The SA was tested in five MIC (0.25 MIC, 0.5 MIC, MIC2, MIC, and 4 MIC) and was tested during different incubation periods (2 hr, 4 hr, 6 hr, 12 hr, and 24 hr), Shikimic acid gave highest percentage inhibition of biofilm formation was 93% at 12hr and ,94% at 24hr , while Acetyl salicylic acid was gave 85% at 12hr and 91% at 24hr incubation period . Then evaluated the synergistic activity of SA and ASA acid with ceftriaxone and azithromycin against biofilm formation also then determination of the antibiofilm activity of SA and ASA acid against bacteria adherence to Foley balloon latex catheter (FBLC) pieces by REMA assay.In addition, determine the efficacy of SA and ASA acid against cell adherence using murine mammary adenocarcinoma cell line (amn3)cells. The results showed high percentage inhibition of biofilm formation as the concentration of SA and ASA increased . In conclusion, SA and ASA have the potential to be further developed as a new antitumor agent due to their impressive antibiofilm action against bacteria. shikimic acid , acetyle salicylic acid were gave up regulation of atlE gene , 2 ( −ΔΔCT) was (2)for shikimic acid while (3)for acetyl salicylic acid , aap,icaA genes, all antibiofilm agents were gave down regulation result 2 ( −ΔΔCT) was (0.2)for shikimic acid while (0.1)for acetyl salicylic acid.

Enhancing Cancer Cell Membrane Permeability by Application of Tumor Treating Fields (TTFields)

<h3>Purpose/Objective(s)</h3> Tumor Treating Fields (TTFields) are alternating electric fields that display anti-mitotic effects on cancer cells. TTFields therapy is approved for treatment of glioblastoma (GBM) and unresectable malignant pleural mesothelioma. It has recently been shown that treatment of GBM cells with TTFields increases cell membrane permeability. The current study aimed to further explore this effect, testing the potential of TTFields to facilitate cellular accumulation of the anticancer agent doxorubicin (DOX) <i>in vitro</i> and <i>in vivo</i>. <h3>Materials/Methods</h3> Breast mammary carcinoma (4T1), breast adenocarcinoma (MCF-7), uterine sarcoma (MES-SA), and lung fibroblast (MRC-5) cell lines were treated with TTFields for 72 h at an intensity of 1.7 V/cm RMS across a range of frequencies (100-400 kHz). Membrane permeability was determined by 7-aminoactinomycin D (7-AAD) intracellular accumulation, and cytotoxicity was examined by cell counts, both measured by flow cytometry. TTFields at the frequency inducing highest permeability were applied together with DOX to DOX-sensitive and matched DOX-resistant 4T1 cells, followed by flow cytometry measurements of intracellular DOX accumulation and cytotoxicity. Mice orthotopically inoculated with 4T1 cells were treated with TTFields for 72 h, with DOX injected 24 h before end of treatment. DOX florescence was measured using flow cytometry for detection at the single-cell level and <i>in vivo</i> imaging system (IVIS) for whole tumor assessment. <h3>Results</h3> While TTFields increased 7-AAD intracellular accumulation in 4T1, MCF-7, and MES-SA cancer cells, there was no such effect when TTFields were applied to the non-cancer MRC-5 cells. Maximal permeability was induced in 4T1 cells with 300 kHz TTFields, whereas highest TTFields-induced cytotoxicity was observed at 150 kHz. TTFields application allowed for DOX accumulation in DOX-resistant cells to the same extent as in DOX-sensitive cells. TTFields delivery also sensitized both cell types to DOX, with cytotoxicity observed at low drug concentrations. <i>In vivo</i>, DOX accumulation was elevated by 2- to 3-fold in tumors isolated from mice treated with TTFields relative to control mice. <h3>Conclusion</h3> TTFields elevated the permeability of 4T1 breast cancer cells, allowing for enhanced intracellular accumulation of DOX and improved drug efficacy, even in cells resistant to DOX. TTFields-induced accumulation of DOX was also demonstrated <i>in vivo</i>.

Lanostane triterpenoids from the fruiting bodies of Ganoderma hainanense and their cytotoxic activity

Three undescribed lanostane triterpenoids, 24E-en-11-oxo-ganoderiol D (1), 11β-hydroxy-ganoderiol D (2), and 11β-hydroxy-lucidone H (3) were isolated from the 80% EtOH extract of the fruiting bodies of Ganoderma hainanense. Structural elucidation of all the compounds were performed by spectral methods such as 1 D and 2 D (1H–1H COSY, HMQC, and HMBC) NMR spectroscopy. All the triterpenoids were in vitro evaluated for their cytotoxic activities against six mammary adenocarcinoma cell lines (MCF7, MDA-MB-231, SK-BR-3, BT-20, HCC38, and AU565). As a result, compound 3 exhibited significant cytotoxic activities against all tested cell lines with IC50 values less than 20 μM.

Hydrogen sulfide decreases photodynamic therapy outcome through the modulation of the cellular redox state

Photodynamic therapy (PDT) is a non-surgical treatment that has been approved for its human medical use in many cancers. PDT involves the interaction of a photosensitizer (PS) with light. The amino acid 5- aminolevulinic acid (ALA) can be used as a pro-PS, leading to the synthesis of Protoporphyrin IX.Hydrogen sulfide (H2S) is an endogenously produced gas that belongs to the gasotransmitter family, which can diffuse through biological membranes and have relevant physiological effects such as cardiovascular functions, vasodilatation, inflammation, cell cycle and neuro-modulation. It was also proposed to have cytoprotective effects. We aimed to study the modulatory effects of H2S on ALAPDT in the mammary adenocarcinoma cell line LM2. Exposure of the cells to NaHS (donor of H2S) in concentrations up to 10 mM impaired the response to ALA-PDT in a dose-dependent manner. The addition of 3 doses of NaHS showed the highest effect. This decreased response to the photodynamic treatment was correlated to an increase in the GSH levels, catalase activity, a dose dependent reduction of PpIX and increased intracellular ALA, decreased levels of oxidized proteins and a decrease of PDT-induced ROS. NaHS also reduced the levels of singlet oxygen in an in vitro assay. H2S also protected other cells of different origins against PDT mediated by ALA and other PSs.These results suggest that H2S has a role in the modulation of the redox state of the cells, and thus impairs the response to ALA-PDT through multifactor pathways. These findings could contribute to developing new strategies to improve the effectiveness of PDT particularly mediated by ALA or other ROS-related treatments.

Investigation of the Lethal Effect of Purified Arginine Deiminase Purified from Lactobacillusplantarum p5 on Murine Mammary Adenocarcinoma and Vero cell Lines.

Cancer is one of the most serious diseases facing humanity; accordingly, it is urgent to find a cure that is rarely harmful to the patient as much as possible. It has been approved that arginine deiminase (ADI) can hydrolyze the plasma arginine to citrulline. This hydrolysis activity and reduction in the amount of intercellular arginine suppress lipopolysaccharide-induced nitric oxide synthesis. On the other hand, arginine depletion arrests the cell cycle at the G1 phase; therefore, ADI has been considered a powerful anticancer agent. The current study aimed to investigate the lethal effects of ADI purified from the Lactobacillus plantarum p5 strain on murine mammary adenocarcinoma and Vero cell lines. Anti-proliferative activity of ADI against murine mammary adenocarcinoma) AMN3) cell line was evaluated after different incubation times (3, 6, 24, 48, and 72 h) of exposure to 1 µg/mL of ADI, compared to Vero (non-cancer cell line) transformed cell line with same conditions. The autophagy process in cancer cells was recognized after three hours of incubation with ADI which was clearly observed in the AMN3 cell line under an inverted microscope. The first stages of the programmed cell death (apoptosis) pathway were only observed in AMN3 cells after 24 h of incubation with ADI, and this process continued with the time until they reached the last stages of apoptosis after 72 h of incubation. The results of the current study showed that the AMN3 cell line was auxotrophic for arginine because it could not produce it in the presence of enzyme which had a robust activity to kill these cancer cells; however, Vero non-cancer cell line survived in the presence of ADI because it had the ability to produce arginine.

In Vitro Cytotoxic Effects of Secondary Metabolites Present in Sarcopoterium Spinosum L.

Sarcopoterium spinosum (L.) is a medicinal plant traditionally used for the treatment of various diseases including cancer in the Near- and Middle East. The fractions and constituents of the ethanol extract of S. spinosum were screened for in vitro cytotoxic activities on Jurkat (acute T-lymphoblastic leukemia), HeLa (cervical adenocarcinoma), MCF-7 (mammary gland adenocarcinoma), Caco-2 (human colorectal adenocarcinoma), and MDA-MB-231 (mammary gland adenocarcinoma) cell lines using the MTT (3-(dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. The ethanol extract was subsequently re-extracted with ethyl acetate and in its sub-fraction obtained by column chromatography three compounds (stachydrine, benzalkonium chloride and rutine) were the first time identified by nuclear magnetic resonance (NMR) analyses. The most active subfraction showed cytotoxic activity against HeLa, MCF-7, and Caco-2 cell lines. The three compounds mentioned, as standards of high-performance liquid chromatography (HPLC) quality, were studied individually and in combination. Cytotoxic activity observed might be due to the presence of benzalkonium chloride and rutin. Benzalkonium chloride showed the strongest growth suppression effect against HeLa cells (IC50 8.10−7 M) and MCF-7 cells (IC50 5.10−6 M). The mixture of stachydrine and benzalkonium chloride allowed a synergistic cytotoxic effect against all tested cancer and normal cells to be obtained. Anti-cancer activity of the plant extract of S. spinosum remains under-investigated, so this research describes how the three major compounds identified in the ethyl acetate extract can exert a significant dose dependent in vitro cytotoxicity.

In vitro estrogenic, cytotoxic, and genotoxic profiles of the xenoestrogens 8-prenylnaringenine, genistein and tartrazine

Phytoestrogens have been widely praised for their health-promoting effects, whereas synthetic environmental estrogens are considered a toxicological risk to human health. The aim of this study was therefore to compare in vitro the estrogenic, cytotoxic, and genotoxic profiles of three common estrogen-like endocrine-disrupting chemicals: the phytoestrogens 8-prenylnaringenine (8-PN) and genistein and the synthetic xenoestrogen tartrazine. As assessed by a yeast bioreporter assay and estrogen-dependent proliferative response in human mammary gland adenocarcinoma cell line (MCF-7), 8-PN showed the highest estrogen-like activity of the three compounds, followed by tartrazine and genistein. After 24-h incubation on MCF-7 cells, all three compounds exhibited low cytotoxicity in the lactate dehydrogenase assay and no genotoxicity in the micronucleus assay. These results demonstrate that 8-PN, genistein and tartrazine possess variable estrogenic activity but display little cellular toxicity in short-term tests in vitro. No difference between phytoestrogens and a synthetic xenoestrogen could be established.

Type 1 conventional dendritic cells and interferons are required for spontaneous CD4+ and CD8+ T-cell protective responses to breast cancer.

ObjectivesTo better understand how immune responses may be harnessed against breast cancer, we investigated which immune cell types and signalling pathways are required for spontaneous control of a mouse model of mammary adenocarcinoma.MethodsThe NOP23 mammary adenocarcinoma cell line expressing epitopes derived from the ovalbumin model antigen is spontaneously controlled when orthotopically engrafted in syngeneic C57BL/6 mice. We combined this breast cancer model with antibody‐mediated depletion of lymphocytes and with mutant mice affected in interferon (IFN) or type 1 conventional dendritic cell (cDC1) responses. We monitored tumor growth and immune infiltration including the activation of cognate ovalbumin‐specific T cells.ResultsBreast cancer immunosurveillance required cDC1, NK/NK T cells, conventional CD4+ T cells and CD8+ cytotoxic T lymphocytes (CTLs). cDC1 were required constitutively, but especially during T‐cell priming. In tumors, cDC1 were interacting simultaneously with CD4+ T cells and tumor‐specific CTLs. cDC1 expression of the XCR1 chemokine receptor and of the T‐cell‐attracting or T‐cell‐activating cytokines CXCL9, IL‐12 and IL‐15 was dispensable for tumor rejection, whereas IFN responses were necessary, including cDC1‐intrinsic signalling by STAT1 and IFN‐γ but not type I IFN (IFN‐I). cDC1 and IFNs promoted CD4+ and CD8+ T‐cell infiltration, terminal differentiation and effector functions. In breast cancer patients, high intratumor expression of genes specific to cDC1, CTLs, CD4+ T cells or IFN responses is associated with a better prognosis.ConclusionInterferons and cDC1 are critical for breast cancer immunosurveillance. IFN‐γ plays a prominent role over IFN‐I in licensing cDC1 for efficient T‐cell activation.