Triple-negative Cells Research Articles

ERα status of invasive ductal breast carcinoma as a result of regulatory interactions between lysine deacetylases KAT6A and KAT6B

Breast cancer (BC) is the leading cause of death among cancer patients worldwide. In 2020, almost 12% of all cancers were diagnosed with BC. Therefore, it is important to search for new potential markers of cancer progression that could be helpful in cancer diagnostics and successful anti-cancer therapies. In this study, we investigated the potential role of the lysine acetyltransferases KAT6A and KAT6B in the outcome of patients with invasive breast carcinoma. The expression profiles of KAT6A/B in 495 cases of IDC and 38 cases of mastopathy (FBD) were examined by immunohistochemistry. KAT6A/B expression was also determined in the breast cancer cell lines MCF-7, BT-474, SK-BR-3, T47D, MDA-MB-231, and MDA-MB-231/BO2, as well as in the human epithelial mammary gland cell line hTERT-HME1 – ME16C, both at the mRNA and protein level. Statistical analysis of the results showed that the nuclear expression of KAT6A/B correlates with the estrogen receptor status: KAT6ANUC vs. ER r = 0.2373 and KAT6BNUC vs. ER r = 0.1496. Statistical analysis clearly showed that KAT6A cytoplasmic and nuclear expression levels were significantly higher in IDC samples than in FBD samples (IRS 5.297 ± 2.884 vs. 2.004 ± 1.072, p < 0.0001; IRS 5.133 ± 4.221 vs. 0.1665 ± 0.4024, p < 0.0001, respectively). Moreover, we noticed strong correlations between ER and PR status and the nuclear expression of KAT6A and KAT6B (nucKAT6A vs. ER, p = 0.0048; nucKAT6A vs. PR p = 0.0416; nucKAT6B vs. ER p = 0.0306; nucKAT6B vs. PR p = 0.0213). Significantly higher KAT6A and KAT6B expression was found in the ER-positive cell lines T-47D and BT-474, whereas significantly lower expression was observed in the triple-negative cell lines MDA-MB-231 and MDA-MB-231/BO2. The outcomes of small interfering RNA (siRNA)-mediated suppression of KAT6A/B genes revealed that within estrogen receptor (ER) positive and negative cell lines, MCF-7 and MDA-MB-231, attenuation of KAT6A led to concurrent attenuation of KAT6A, whereas suppression of KAT6B resulted in simultaneous attenuation of KAT6A. Furthermore, inhibition of KAT6A/B genes resulted in a reduction in estrogen receptor (ER) mRNA and protein expression levels in MCF-7 and MDA-MMB-231 cell lines. Based on our findings, the lysine acetyltransferases KAT6A and KAT6B may be involved in the progression of invasive ductal breast cancer. Further research on other types of cancer may show that KAT6A and KAT6B could serve as diagnostic and prognostic markers for these types of malignancies.

Synthesis of nanohybrid consisting of taurine derived carbon dots and nanoceria for anticancer applications

Here, we first synthesized carbon dots (CDs) from taurine and then a nanohybrid with ceria (CeO2) nanoparticles using thermal decomposition method for checking their antineoplastic efficacy against human triple negative mammary carcinoma cells, MDA-MB-231. The in vitro study demonstrated significant dose-dependent antineoplastic activity of the nanohybrids within a range of concentration from 10 to 50 μg/mL after 48 h of treatment. The cellular morphology analysis clearly depicted substantial amount of cell death which seems to stem from increased intracellular reactive oxygen species (ROS) production. However, the maximum anticancer activity of the nanohybrid as compared to bare CDs and CeO2 is supposed to be due to the combined anticancer effect of both CDs and CeO2 (a well-established antitumor agent). Further we have performed molecular docking study to reveal the anticancer mechanism of CDs which exhibited high binding capacity towards several proapoptotic and antiapoptotic protein molecules. The binding affinity values were found to be – 8.7 kcal/mol, – 7.9 kcal/mol, – 9.6 kcal/mol, – 9.5 kcal/mol, – 12 kcal/mol and – 11.1 kcal/mol for BAD, BCl-2, p53, Caspase-8, Caspase-9 and Caspase-3 respectively. Taken together, our synthesized CDs-CeO2 nanohybrid could be thought as a potential anticarcinogenic option in the field of breast cancer therapeutics.

Ferrocenyl β-Diketonate Compounds: Extended Ring Systems for Improved Anticancer Activity.

A library of ferrocenyl β-diketonate compounds with varying degrees of aromatic functionality have been synthesized and fully characterized. This includes cyclic voltammetry and the analysis of four new structures by single crystal X-ray diffraction. The compounds cytotoxic potential has been determined by MTT screening against pancreatic carcinoma (MIA PaCa-2), ovarian adenocarcinoma (A2780), breast adenocarcinomas (MDA-MB-231 and MCF-7) and normal epithelial retinal (ARPE-19). The compounds show a general trend, where increasing the number of aromatic rings in the molecule yields an increase in cytotoxicity and follows the trend anthracenyl > naphthyl > phenyl > methyl. The compounds are particularly sensitive to the triple negative cancer cell line MDA-MB-231, and the potential modes of action have been studied by production of reactive oxygen species using fluorescence microscopy and cell morphology using Scanning Electron Microscopy. All assays highlight the ferrocenyl β-diketonate with an anthracenyl substituent to be the lead compound in this library. The decomposition was also observed within cells, to give a cytotoxic fluorescent molecule, which has been visualized by confocal microscopy.

In vitro and computational investigation of antioxidant and anticancer properties of Streptomyces coeruleofuscus SCJ extract on MDA-MB-468 triple-negative breast cancer cells

This study aimed to explore the antioxidant potential of the ethyl acetate extract of Streptomyces coeruleofuscus SCJ strain, along with its inhibitory effects on the triple-negative human breast carcinoma cell line (MDA-MB-468). The ethyl acetate extract’s total phenolic and flavonoid contents were quantified, and its antioxidant activity was investigated using DPPH (1,1-Diphenyl-2-picrylhydrazyl), ABTS (2,2’-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid), and FRAP (Ferric Reducing Antioxidant Power) assays. Furthermore, the cytotoxic effect of the organic extract from Streptomyces coeruleofuscus SCJ on MDA-MB-468 cancer cells was assessed via the crystal violet assay. In tandem, a thorough computational investigation was conducted to explore the pharmacokinetic properties of the identified components of the extract, utilizing the SwissADME and pKCSM web servers. Additionally, the molecular interactions between these components and Estrogen Receptor Beta, identified as a potential target, were probed through molecular docking studies. The results revealed that ethyl acetate extract of SCJ strain exhibited remarkable antioxidant activity, with 39.899 ± 1.56% and 35.798 ± 0.082% scavenging activities against DPPH and ABTS, respectively, at 1 mg/mL. The extract also displayed significant ferric reducing power, with a concentration of 1.087 ± 0.026 mg ascorbic acid equivalents per mg of dry extract. Furthermore, a strong positive correlation (p < 0.0001) between the antioxidant activity, the polyphenol and the flavonoid contents. Regarding anticancer activity, the SCJ strain extract demonstrated significant anticancer activity against TNBC MDA-MB-468 cancer cells, with an inhibition percentage of 62.76 ± 0.62%, 62.67 ± 0.93%, and 58.07 ± 4.82% at 25, 50, and 100 µg/mL of the extract, respectively. The HPLC-UV/vis analysis revealed nine phenolic compounds: gallic acid, sinapic acid, p-coumaric acid, cinnamic acid, trans-fereulic acid, syringic acid, chloroqenic acid, ellagic acid, epicatechin. Streptomyces coeruleofuscus SCJ showed promise for drug discovery, exhibiting antioxidant and anticancer effects.

High prevalence of chromosome 17 in breast cancer micronuclei: a means to get rid of tumor suppressors?

Micronuclei (MN), defined as small extra-nuclear chromatin bodies enclosed by a nuclear envelope, serve as noticeable markers of chromosomal instability (CIN). The MN have been used for breast cancer (BC) screening, diagnosis, and prognosis. However, more recently they have gained attention as seats for active chromosomal rearrangements. BC subtypes exhibit differential CIN levels and aggressiveness. This study aimed to investigate MN chromosomal contents across BC subtypes, exploring its potential role in aggressiveness and pathogenesis. Immunostaining of BC cells was performed with anti-centromeric antibody followed by confocal microscopy. Further, fluorescence in situ hybridization (FISH) was done to check the presence of specific chromosomes in the MN. The real time PCR was also done from the RNA isolated from MN to check the expression of TP53 gene. BC cell lines (CLs) showed the presence of both centromere-positive ( +) and -negative ( -) MN, with significant variation in frequency among hormone and human epidermal growth factor receptor positive and triple-negative (TN) BC cells. FISH targeting chromosomes 1, 3, 8, 11, and 17 detected centromeric signals for all the above chromosomes in MN with a relatively higher prevalence of chromosome 17 in all the CLs. Out of all the CLs, TNBC cells demonstrated the highest frequency of centromere + and chromosome 17 + MN. TP53 expression could also be demonstrated inside the MN by FISH and real time PCR. Patient sample imprints also confirmed the presence of chromosome 17 in MN with polysomy of the same in corresponding nuclei. The high prevalence of chromosome 17 in BC MN may connote the importance of its rearrangements in the pathogenesis of BC. Further, the higher prevalence of chromosome 17 and 1 signals in TNBC MN point towards the significance of pathogenetic events involving the genes located in these chromosomes in evolution of this more aggressive phenotype.

Anticancer Properties Against Select Cancer Cell Lines and Metabolomics Analysis of Tender Coconut Water.

Tender Coconut Water (TCW) is a nutrient-rich dietary supplement that contains in bioactive secondary metabolites and phytohormones with anti-oxidative and anti-inflammatory properties. Studies on TCW's anti-cancer properties are limited and the mechanism of its anti-cancer effects have not been defined. In the present study, we investigate TCW for its anti-cancer properties and, using untargeted metabolomics, we identify components form TCW with potential anti-cancer activity. Cell viability assay, BrdU incorporation assay, soft-agar assay, flow-cytometery, and Western blotting were used to analyze TCW's anticancer properties and to identify mechanism of action. Liquid chromatography- Tandem Mass Spectroscopy (LC-MS/MS) was used to identify TCW components. TCW decreased the viability and anchorage-independent growth of HepG2 hepatocellular carcinoma (HCC) cells and caused S-phase cell cycle arrest. TCW inhibited AKT and ERK phosphorylation leading to reduced ZEB1 protein, increased E-cadherin, and reduced N-cadherin protein expression in HepG2 cells, thus reversing the 'epithelial-to-mesenchymal' (EMT) transition. TCW also decreased the viability of Hep3B hepatoma, HCT-15 colon, MCF-7 and T47D luminal A breast cancer (BC) and MDA-MB-231 and MDA-MB-468 triplenegative BC cells. Importantly, TCW did not inhibit the viability of MCF-10A normal breast epithelial cells. Untargeted metabolomics analysis of TCW identified 271 metabolites, primarily lipids and lipid-like molecules, phenylpropanoids and polyketides, and organic oxygen compounds. We demonstrate that three components from TCW: 3-hydroxy-1-(4-hydroxyphenyl)propan-1-one, iondole-3-carbox aldehyde and caffeic acid inhibit the growth of cancer cells. TCW and its components exhibit anti-cancer effects. TCW inhibits the viability of HepG2 hepatocellular carcinoma cells by reversing the EMT process through inhibition of AKT and ERK signalling.

Molecular mechanisms of novel selective glucocorticoid receptor agonist action in breast cancer cells

Introduction. Glucocorticoids (GC) are widely used in breast cancer (BC) therapy to reduce the side effects of cytostatic drugs and may exhibit antiproliferative effects on luminal BC cells. The biological action of GC is mediated by glucocorticoid receptor (GR) by two mechanisms: transrepression, which determines the therapeutic effect of GC, and transactivation (associated with the development of side effects, resistance to cytotoxic drugs, cancer progression and metastasis). Selective GR agonists (SEGRA) which may selective activate transrepression are a promising alternative to GC to use in combination cancer therapy. One of the most studied SEGRA is Compound A (CpdA). The instability of CpdA limits its use in clinical practice. So recently we performed synthesis and evaluation of biological activities of the CpdA analogue, CpdA-03.Aim. To compare the effects of SEGRA CpdA-03 and CpdA and dexamethasone on proliferative activity of breast cancer cells, as well as receptor nuclear translocation and activation of GR-dependent genes in breast cancer cells.Materials and methods. Luminal (MCF-7) and triple negative (MDA-MB-231) BC cell lines were used. The effect of CpdA-03 on proliferation was evaluated by direct counting of viable cells with trypan blue staining. The effect of the compound on cell distribution by cell cycle phases was assessed by flow cytofluorimetry with propidium iodide staining. Changes in the expression of GR-dependent genes after incubation with CpdA-03 were evaluated by quantitative polymerase chain reaction. Additionally, the ability of the new SEGRA to induce transactivation-associated translocation of the receptor to the nucleus was evaluated by Western blotting.Results. CpdA-03 was shown to suppress proliferation of luminal and triple negative BC cells. This compound causes changes in the expression of a number of GC-inducible genes, but does not stimulate GR phosphorylation and translocation to the nucleus in BC cells.Conclusion. The observed suppression of cell proliferation, as well as the ability of CpdA-03 to reduce gene expression of proteins regulating intercellular adhesion and cell migration, intracellular signaling, stress response, and transcription in BC cells makes it relevant for further development of the drug for use in the combination therapy of cancers including breast cancer.

Enhanced Synergistic Efficacy Against Breast Cancer Cells Promoted by Co-Encapsulation of Piplartine and Paclitaxel in Acetalated Dextran Nanoparticles.

Malignant breast tumors constitute the most frequent cancer diagnosis among women. Notwithstanding the progress in treatments, this condition persists as a major public health issue. Paclitaxel (PTX) is a first-line classical chemotherapeutic drug used as a single active pharmaceutical ingredient (API) or in combination therapy for breast cancer (BC) treatment. Adverse effects, poor water solubility, and inevitable susceptibility to drug resistance seriously limit its therapeutic efficacy in the clinic. Piplartine (PPT), an alkaloid extracted from Piper longum L., has been shown to inhibit cancer cell proliferation in several cell lines due to its pro-oxidant activity. However, PPT has low water solubility and bioavailability in vivo, and new strategies should be developed to optimize its use as a chemotherapeutic agent. In this context, the present study aimed to synthesize a series of acetalated dextran nanoparticles (Ac-Dex NPs) encapsulating PPT and PTX to overcome the limitations of PPT and PTX, maximizing their therapeutic efficacy and achieving prolonged and targeted codelivery of these anticancer compounds into BC cells. Biodegradable, pH-responsive, and biocompatible Ac-Dex NPs with diameters of 100-200 nm and spherical morphologies were formulated using a single emulsion method. Selected Ac-Dex NPs containing only PPT or PTX as well as those coloaded with PPT and PTX achieved excellent drug-loading capabilities (PPT, ca. 11-33%; PTX, ca. 2-14%) and high encapsulation efficiencies (PPT, ∼57-98%; PTX, ∼80-97%). Under physiological conditions (pH 7.4), these NPs exhibited excellent colloidal stability and were capable of protecting drug release, while under acidic conditions (pH 5.5) they showed structural collapse, releasing the therapeutics in an extended manner. Cytotoxicity results demonstrated that the encapsulation in Ac-Dex NPs had a positive effect on the activities of both PPT and PTX against the MCF-7 human breast cancer cell line after 48 h of treatment, as well as toward MDA-MB-231 triple-negative BC cells. PPT/PTX@Ac-Dex NPs were significantly more cytotoxic (IC50/PPT = 0.25-1.77 μM and IC50/PTX = 0.07-0.75 μM) and selective (SI = 2.9-6.7) against MCF-7 cells than all the control therapeutic agents: free PPT (IC50 = 4.57 μM; SI = 1.2), free PTX (IC50 = 0.97 μM; SI = 1.0), the single-drug-loaded Ac-Dex NPs, and the physical mixture of both free drugs. All combinations of PPT and PTX resulted in pronounced synergistic antiproliferative effects in MCF-7 cells, with an optimal molar ratio of PPT to PTX of 2.3:1. PPT/PTX-2@Ac-Dex NPs notably promoted apoptosis, cell cycle arrest at the G2/M, accumulation of intracellular reactive oxygen species (ROS), and combined effects from both PPT and PTX on the microtubule network of MCF-7 cells. Overall, the combination of PTX and PPT in pH-responsive Ac-Dex NPs may offer great potential to improve the therapeutic efficacy, overcome the limitations, and provide effective simultaneous delivery of these therapeutics for BC treatment.

Engineering Microgel Packing to Tailor the Physical and Biological Properties of Gelatin Methacryloyl Granular Hydrogel Scaffolds.

Granular hydrogel scaffolds (GHS) are fabricated via placing hydrogel microparticles (HMP) in close contact (packing), followed by physical and/or chemical interparticle bond formation. Gelatin methacryloyl (GelMA) GHS have recently emerged as a promising platform for biomedical applications; however, little is known about how the packing of building blocks, physically crosslinked soft GelMA HMP, affects the physical (pore microarchitecture and mechanical/rheological properties) and biological (in vitro and in vivo) attributes of GHS. Here, the GHS pore microarchitecture is engineered via the external (centrifugal) force-induced packing and deformation of GelMA HMP to regulate GHS mechanical and rheological properties, as well as biological responses in vitro and in vivo. Increasing the magnitude and duration of centrifugal force increases the HMP deformation/packing, decreases GHS void fraction and median pore diameter, and increases GHS compressive and storage moduli. MDA-MB-231 human triple negative breast adenocarcinoma cells spread and flatten on the GelMA HMP surface in loosely packed GHS, whereas they adopt an elongated morphology in highly packed GHS as a result of spatial confinement. Via culturing untreated or blebbistatin-treated cells in GHS, the effect of non-muscle myosin II-driven contractility on cell morphology is shown. In vivo subcutaneous implantation in mice confirms a significantly higher endothelial, fibroblast, and macrophage cell infiltration within the GHS with a lower packing density, which is in accordance with the in vitro cell migration outcome. These results indicate that the packing state of GelMA GHS may enable the engineering of cell response in vitro and tissue response in vivo. This research is a fundamental step forward in standardizing and engineering GelMA GHS microarchitecture for tissue engineering and regeneration.

A broad-spectrum biological activities of Heracleum humile extracts: A first report of the antiviral, anti-cancer and chemical properties

The current work focused on researching the bioactive potential of ethyl acetate (EA), methanol (MeOH), and water extract of H. humile plant, specifically ethyl acetate, methanol extracts were acquired with maceration method, and water extract was obtained by the infusion method. The highest total phenolic ingredient was determined in the water extract, while the highest total flavonoid ingredient was found in the MeOH extract. The water extract displayed the most potent antioxidant activity in the assays used. The ethyl acetate extract showed the highest enzyme inhibition activity in Butyrylcholinesterase inhibition (BChE) and amylase assay. Antibacterial efficiency was analyzed using microdilution analysis. The minimum inhibitory concentration (MIC) amounts ranged from 125 to 500 μg/mL except for water extract. The antiviral potential of the extracts was assessed against Bovine herpes virus 1 (BoHV-1) (Cooper strain). Methanol extract showed more potent antiviral effect. Selectivity indices were calculated for ethyl acetate and methanol extract as 3.32 and 13.39, respectively. All extracts displayed Deoxyribonucleic Acid (DNA) protection activity at approximately 25%–35%. For cell viability evaluation, 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) test was performed against triple-negative human breast adenocarcinoma cells (MDA-MB-231) and Michigan Cancer Foundation-7 (MCF-7) cells. The most effective results were found in ethyl acetate extract at 48h. The 50% inhibitor concentrations (IC50) were determined for MDA-MB-231 and MCF-7 as 116.7 μg/mL and 247.9 μg/mL, respectively. Based on the bioactive compound content results, extracts of H. humile include valuable compounds that possess significant biological properties. In conclusion, these extracts could offer promising novel plant-derived therapeutics.

Siglec-7 and Siglec-9 expression in primary triple negative and oestrogen receptor positive breast cancer and in vitro signalling.

PD-1/PD-L1 immune checkpoint blockade can be an effective treatment for advanced breast cancer patients. However, patients with oestrogen receptor positive (ER+) tumors often display only low lymphocyte infiltration, while a large part of triple negative (TN) breast tumors does not generate an effective immunotherapy response. Therefore, new treatment strategies have to be developed. Here, we investigate Siglec-7 and Siglec-9 as novel ITIM-bearing inhibitory immune checkpoint receptors similar to PD-1, but expressed on a broader range of immune cells. We assessed Siglec-7 and Siglec-9 (ligand) expression in TN and ER+ breast cancer tumors and their breast cancer cell line-induced signalling. We report that Siglec-7 and Siglec-9 are highly expressed in TN tumors, and to a low extent in ER+ tumors. Siglec-7 was observed on myeloid cells, Tcells, and NKcells and Siglec-9 preferentially on myeloid cells. Expression of sialoglycans, including Siglec-7 and Siglec-9 ligands, was observed in both TN and ER+ breast cancer tissue sections. Expression levels of Siglec-7 and Siglec-9 ligands were higher on in vitro cultured TN cell lines than ER+ cell lines. Importantly, by applying chimeric Siglec-7 reporter cells, we showed the induction of Siglec-7 signalling by multiple TN cell lines, but only by one ER+ cell line. Moreover, Siglec-7 signalling is directly related to Siglec-7 ligand expression levels of breast cancer cell lines. These data imply that immunotherapy targeting Siglec receptors may be particularly interesting for TN breast cancer patients not responding to current treatment strategies with tumors displaying high immune cell infiltration.

A novel lipophilic amiloride derivative efficiently kills chemoresistant breast cancer cells

Derivatives of the potassium-sparing diuretic amiloride are preferentially cytotoxic toward tumor cells relative to normal cells, and have the capacity to target tumor cell populations resistant to currently employed therapeutic agents. However, a major barrier to clinical translation of the amilorides is their modest cytotoxic potency, with estimated IC50 values in the high micromolar range. Here we report the synthesis of ten novel amiloride derivatives and the characterization of their cytotoxic potency toward MCF7 (ER/PR-positive), SKBR3 (HER2-positive) and MDA-MB-231 (triple negative) cell line models of breast cancer. Comparisons of derivative structure with cytotoxic potency toward these cell lines underscore the importance of an intact guanidine group, and uncover a strong link between drug-induced cytotoxicity and drug lipophilicity. We demonstrate that our most potent derivative called LLC1 is preferentially cytotoxic toward mouse mammary tumor over normal epithelial organoids, acts in the single digit micromolar range on breast cancer cell line models representing all major subtypes, acts on cell lines that exhibit both transient and sustained resistance to chemotherapeutic agents, but exhibits limited anti-tumor effects in a mouse model of metastatic breast cancer. Nonetheless, our observations offer a roadmap for the future optimization of amiloride-based compounds with preferential cytotoxicity toward breast tumor cells.

The GPR30 Receptor Is Involved in IL-6-Induced Metastatic Properties of MCF-7 Luminal Breast Cancer Cells.

Luminal breast cancer has a high incidence worldwide and poses a severe health threat. Estrogen receptor alpha (ER-α) is activated by 17β-estradiol (E2), and its overexpression promotes cancerous characteristics. Luminal breast cancer is an epithelial type; however, the cytokine IL-6, secreted by cells within the tumor microenvironment, stimulates the epithelial-to-mesenchymal transition (EMT) and promotes metastasis. Also, IL-6 decreases ER-α levels, favoring the tamoxifen (TMX) resistance development. However, genes under E2 regulation continue to be expressed even though this receptor is absent. GPR30 is an alternative E2 receptor present in both luminal and aggressive triple-negative breast cancer and is related to TMX resistance and cancer progression. The roles of GPR30 and IL-6 in metastasis have been individually established; however, their interplay remains unexplored. This study aims to elucidate the role of GPR30 in IL-6-induced metastatic properties of MCF-7 luminal breast cancer cells. Results showed that GPR30 contributes to the E2-induced MCF-7 proliferation because its inhibition with the antagonist G15 and the Pertussis toxin (PTX) reduced it. Besides, GPR30 upregulated vimentin and downregulated E-cadherin levels in MCF-7 and TMX-resistant (R-TMX) cells and is also involved in the IL-6-induced migration, invasion, and TMX resistance in MCF-7 cells. In addition, in MDA-MB-231 triple-negative cells, both basal and IL-6-induced metastatic properties were related to GPR30 activity. These results indicate that the GPR30 receptor regulates the EMT induced by IL-6 in breast cancer cells.

Acetylation of Steroidogenic Acute Regulatory Protein Sensitizes 17β-Estradiol Regulation in Hormone-Sensitive Breast Cancer Cells.

An imbalance in estrogen signaling is a critical event in breast tumorigenesis. The majority of breast cancers (BCs) are hormone-sensitive; they majorly express the estrogen receptor (ER+) and are activated by 17β-estradiol (E2). The steroidogenic acute regulatory protein (StAR) mediates the rate-limiting step in steroid biosynthesis. The dysregulation of the epigenetic machinery, modulating E2 levels, is a primary occurrence for promoting breast tumorigenesis. StAR expression, concomitant with E2 synthesis, was reported to be aberrantly high in human and mouse hormone-dependent BC cells compared with their non-cancerous counterparts. However, the mechanism of action of StAR remains poorly understood. We discovered StAR as an acetylated protein and have identified a number of lysine (K) residues that are putatively acetylated in malignant and non-malignant breast cells, using LC-MS/MS (liquid chromatography-tandem mass spectrometry), suggesting they differently influence E2 synthesis in mammary tissue. The treatment of hormone-sensitive MCF7 cells with a variety of histone deacetylase inhibitors (HDACIs), at therapeutically and clinically relevant doses, identified a few additional StAR acetylated lysine residues. Among a total of fourteen StAR acetylomes undergoing acetylation and deacetylation, K111 and K253 were frequently recognized either endogenously or in response to HDACIs. Site-directed mutagenesis studies of these two StAR acetylomes, pertaining to K111Q and K253Q acetylation mimetic states, resulted in increases in E2 levels in ER+ MCF7 and triple negative MB-231 BC cells, compared with their values seen with human StAR. Conversely, these cells carrying K111R and K253R deacetylation mutants diminished E2 biosynthesis. These findings provide novel and mechanistic insights into intra-tumoral E2 regulation by elucidating the functional importance of this uncovered StAR post-translational modification (PTM), involving acetylation and deacetylation events, underscoring the potential of StAR as a therapeutic target for hormone-sensitive BC.

Metabolic and Oxidative Stress Management Heterogeneity in a Panel of Breast Cancer Cell Lines.

Metabolic alterations are recognized as one of the hallmarks of cancer. Among these, alterations in mitochondrial function have been associated with an enhanced production of Reactive Oxygen Species (ROS), which activate ROS-regulated cancer cell signaling pathways. Breast cancer is the main cancer-related cause of death for women globally. It is a heterogeneous disease with subtypes characterized by specific molecular features and patient outcomes. With the purpose of identifying differences in energy metabolism and the oxidative stress management system in non-tumorigenic, estrogen receptor positive (ER+) and triple negative (TN) breast cancer cells, we evaluated ROS production, protein enzyme levels and activities and profiled energy metabolism. We found differences in energetic metabolism and ROS management systems between non-tumorigenic and cancer cells and between ER+ and TN breast cancer cells. Our results indicate a dependence on glycolysis despite different glycolytic ATP levels in all cancer cell lines tested. In addition, our data show that high levels of ROS in TN cells are a result of limited antioxidant capacity in the NADPH producing and GSH systems, mitochondrial dysfunction and non-mitochondrial ROS production, making them more sensitive to GSH synthesis inhibitors. Our data suggest that metabolic and antioxidant profiling of breast cancer will provide important targets for metabolic inhibitors or antioxidant treatments for breast cancer therapy.

Synergistic Treatment of Breast Cancer by Combining the Antimicrobial Peptide Piscidin with a Modified Glycolipid.

Piscidin 3 (P3), a peptide produced by fish, and a hexyl ester-modified sophorolipid (SL-HE), have individually shown promise as antimicrobial and anticancer drugs. A recent report by our team revealed that combining P3 with SL-HE in a 1:8 molar ratio resulted in an 8-fold enhancement in peptide activity, while SL-HE improved by 25-fold its antimicrobial activity against the Gram-positive microorganism Bacillus cereus. Extending these findings, the same P3/SL-HE combination was assessed on two breast cancer cell lines: BT-474, a hormonally positive cell line, and MDA-MB-231, an aggressive triple-negative cell line. The results demonstrated that the 1:8 molar ratio of P3/SL-HE synergistically enhances the anticancer effects against both tumorigenic breast cell lines. Mechanistic studies indicate the activation of an intrinsic apoptotic cell death mechanism through an increase in reactive oxygen species and mitochondrial dysfunction and a secondary programmed necrotic pathway that involves pore formation in the plasma membrane. When a fibroblast cell line, CCD1065SK HDF, was utilized to determine selectivity, the synergistic SL-HE/P3 combination exhibited a protective property compared to the use of SL-HE alone and therefore afforded vastly improved selectivity indices. Given the promising results reported herein, the synergistic combination of P3/SL-HE constitutes a novel strategy that merits further study for the treatment of breast cancer.

An Electrochemical Immuno-cytosensor Modified with Nanofibers for the Determination of a Carcinoembryonic Antigen

In this study, La0.25Fe0.75FeO3 (PNp)perovskite nanoparticle was synthesized using the sol–gel method. PNp-coated polyacrylonitrile (PAN) nanofibers were prepared by electrospinning on the pencil graphite electrode (PGE) surface. In another step, carcinoembryonic antigen (CEA) was loaded with CEA antibodies (Anti-CEA) as a biomarker receptor. Finally, PGE/PAN@PNp/Anti-CEA was used for CEA detection. Optimization steps and cell culture steps were performed using differential pulse voltammetry (DPV). The use of this composite system is a novel immunosensor development approach for label-free detection of CEA. Under optimum conditions, detection limit (LOD) of PGE/PAN@PNp/Anti-CEA immunosensor LOD 1.48 ng/mL, limit of quantification (LOQ) = 4.94 ng/mL, reproducibility 1.46% (n = 5) and R2 = 0.9984 for antigen concentration within a linear working range of 0.1–10 ng/mL. Also, immunosensor recovery in real serum samples containing dopamine and ascorbic acid was found as 98.94 ± 7.43. It has great potential in clinical screening of different cancer biomarkers. The number of cells attached to the PGE/PAN@PNp/Anti-CEA/BSA(bovine serum)/CEA surface decreased in RT-4(bladder cancer), MDA-MB-231 (triple-negative breast adenocarcinoma cell line), and T98G cells (glioblastoma multiforme cell line), which are known as CEA-negative cell lines, whereas the number of MCF-7 cells (estrogen-sensitive human breast cancer cell line, known to be CEA positive) attached to the PGE/PAN@PNp/Anti-CEA/BSA/CEA surface increased, indicating higher affinity to the immunosensor surface. As a result, while MCF-7, which is CEA positive, can be determined best when using an immune-cytosensor, the cell that can be best determined with cytosensors was found to be RT-4.

The Effect of Photobiomodulation on the Conditioned Media of 3T3-L1 Cells in the Treatment of Breast Cancer.

Introduction: Breast cancer ranks among the most prevalent malignancies, and its prompt diagnosis significantly amplifies the prospects of successful treatment. Approximately one in seven women will experience a breast cancer diagnosis in their lifetime. Stromal cells and their secreted factors exert various effects on tumor growth, impacting proliferation, invasion, and metastasis. Research has emphasized the significant impact of proteins secreted by adipose tissue on breast cancer proliferation, surpassing the influence of factors released by other cell types. Yet, the specific transcription factors and cofactors involved in adipokine expression in the tumor microenvironment remain enigmatic. Methods: In this study, adipocyte cells were cultured and exposed to 980 nm and 650 nm Photobiomodulation. The MDA-MD-231 cells (triple negative cancer cell line) were cultured with a conditioned medium from laser-treated cells. The real-time assay was employed to analyze the gene expression level changes involved in apoptosis. Results: Results showed that the irradiated conditioned medium at 980 nm and 650 nm caused a reduction in cell viability of cancer cells. Conversely, the conditioned medium from the irradiated cells triggered an increase in the expression of Caspase 3, Caspase 9, and BAX2 genes, alongside a decrease in BCL2 gene expression. Conclusion: The findings highlighted the potential of the laser-treated conditioned medium to induce apoptosis pathways in cancer cells, demonstrating a promising avenue for further research in utilizing low-level laser therapy in breast cancer treatment.

The Imidazacridine Derivative LPSF/AC-05 Induces Apoptosis, Cell Cycle Arrest, and Topoisomerase II Inhibition in Breast Cancer, Leukemia, and Lymphoma.

Cancer is the major cause of morbidity and mortality worldwide. Current treatments for both solid and hematological tumors are associated with severe adverse effects and drug resistance, necessitating the development of novel selective antineoplastic drugs. The present study describes the antitumor activity of the imidazacridine derivative 5-acridin-9-ylmethylidene-2-thioxoimidazolidin-4-one (LPSF/AC05) in breast cancer, leuke-mia, and lymphoma cells. Cytotoxicity assays were performed in PBMC and in breast cancer, leukemia, and lymphoma cell lines using the MTT method. Changes in cell cycle progression and apoptosis were assessed using flow cytometry. Moreover, topoisomerase II inhibition as-says were performed. LPSF/AC05 exhibited cytotoxicity in six of the nine cell lines tested. The best results for leukemia and lymphoma were observed in the Toledo, Jurkat, and Raji cell lines (IC50 = 27.18, 31.04, and 33.36 M, respectively). For breast cancer, the best re-sults were observed in the triple-negative cell line MDA-MB-231 (IC50 = 27.54 μM). The compound showed excellent selectivity, with no toxicity to normal human cells (IC50 > 100M; selectivity index > 3). Cell death was primarily induced by apoptosis in all cell lines. Furthermore, LPSF/AC05 treatmentinduced cell cycle arrest at the G0/G1 phase in leuke-mia/lymphoma and at the G2/M phase in breast cancer. Finally, topoisomerase II was inhibited. These results indicate the potential ap-plication of LPSF/AC05 in cancer therapy.

Structure-Activity Relationship Study Identifies a Novel Lipophilic Amiloride Derivative that Efficiently Kills Chemoresistant Breast Cancer Cells.

Derivatives of the potassium-sparing diuretic amiloride are preferentially cytotoxic toward tumor cells relative to normal cells, and have the capacity to target tumor cell populations resistant to currently employed therapeutic agents. However, a major barrier to clinical translation of the amilorides is their modest cytotoxic potency, with estimated IC 50 values in the high micromolar range. Here we report the synthesis of ten novel amiloride derivatives and the characterization of their cytotoxic potency toward MCF7 (ER/PR-positive), SKBR3 (HER2-positive) and MDA-MB-231 (triple negative) cell line models of breast cancer. Comparisons of derivative structure with cytotoxic potency toward these cell lines underscore the importance of an intact guanidine group, and uncover a strong link between drug-induced cytotoxicity and drug lipophilicity. We demonstrate that our most potent derivative called LLC1 is preferentially cytotoxic toward mouse mammary tumor over normal epithelial organoids, acts in the single digit micromolar range on breast cancer cell line models representing all major subtypes, acts on cell lines that exhibit both transient and sustained resistance to chemotherapeutic agents, but exhibits limited anti-tumor effects in a mouse model of metastatic breast cancer. Nonetheless, our observations offer a roadmap for the future optimization of amiloride-based compounds with preferential cytotoxicity toward breast tumor cells.