MCF-7 Human Breast Cancer Cells Research Articles

COPS6 promotes tumor progression and reduces CD8+ T cell infiltration by repressing IL-6 production to facilitate tumor immune evasion in breast cancer.

Due to poor T cell infiltration, tumors evade immune surveillance. Increased CD8+ T cell infiltration in breast cancer suggests a satisfactory response to immunotherapy. COPS6 has been identified as an oncogene, but its role in regulating antitumor immune responses has not been defined. In this study, we investigated the impact of COPS6 on tumor immune evasion in vivo. Tumor transplantation models were established in C57BL/6 J mice and BALB/c nude mice. Flow cytometry was conducted to identify the role of COPS6 on tumor-infiltrating CD8+ T cells. By analyzing the TCGA and GTEx cohort, we found that COPS6 expression was significantly up-regulated in a variety of cancers. In human osteosarcoma cell line U2OS and non-small cell lung cancer cell line H1299, we showed that p53 negatively regulated COPS6 promoter activity. In human breast cancer MCF-7 cells, COPS6 overexpression stimulated p-AKT expression as well as the proliferation and malignant transformation of tumor cells, whereas knockdown of COPS6 caused opposite effects. Knockdown of COPS6 also significantly suppressed the growth of mouse mammary cancer EMT6 xenografts in BALB/c nude mice. Bioinformatics analysis suggested that COPS6 was a mediator of IL-6 production in the tumor microenvironment and a negative regulator of CD8+ T cell tumor infiltration in breast cancer. In C57BL6 mice bearing EMT6 xenografts, COPS6 knockdown in the EMT6 cells increased the number of tumor-infiltrating CD8+ T cells, while knockdown of IL-6 in COPS6KD EMT6 cells diminished tumor infiltrating CD8+ T cells. We conclude that COPS6 promotes breast cancer progression by reducing CD8+ T cell infiltration and function via the regulation of IL-6 secretion. This study clarifies the role of p53/COPS6/IL-6/CD8+ tumor infiltrating lymphocytes signaling in breast cancer progression and immune evasion, opening a new path for development of COPS6-targeting therapies to enhance tumor immunogenicity and treat immunologically "cold" breast cancer.

Cytotoxicity and Antimicrobial efficiency of gold (Au) nanoparticles formulated by green approach using Andrographis paniculata leaf extract

Gold (Au) nanoparticles have flourished a promise for solicitations in biology due to their processability and high performance in the release of therapeutics. The use of leaf substances in the fundamental biosynthesis and manufacturing of nano-phytomedicine is the cooperative effort between plant science and nanotechnology, yielding a green nanostructured material. This study aims to synthesize AuNPs using the extract taken from Andrographis paniculata leaves and to evaluate its efficacy against MCF7 cells and chosen human pathogens. AP-AuNPs were studied with UV, FTIR, and X-ray diffraction spectroscopy, SEM and TEM (scanning and transmission electron microscopy). At 533 nm, a peak of Surface Plasmon Resonance (SPR) was identified using UV–Visible spectra of AuNPs. The XRD diffraction peaks indicated the crystalline nature of gold nanoparticles. The spherical shape was demonstrated by transmission electron microscopy, and dynamic light scattering using Particle Size Analyzer measurements revealed a mean particle size of 40 ± 18.6 nm. The research concluded that using gold nanoparticles derived from herbs is contingent on safe dosage and precise formulation, as stated in ancient literature. MCF7 human breast cancer cells have been used to evaluate the anti-proliferative activity of AuNP. The findings of an oncology investigation revealed that treating cancerous cells using AuNPs drastically diminished the viability of cells. The anti-microbial properties in the primordial herbal-based metal nanoparticles confess the conventional healthcare practice as a progressive approach for contemporary conditions. Based on the investigation, the study inferred that AP-AuNPs are therapeutic in combating human pathogens.

Size and Morphology-Mediated Antiproliferative Activity of Hydroxyapatite Nanoparticles in Human Breast Cancer Cells

Hydroxyapatite nanoparticles (nHAPs) have been recognized for potent antitumor effects in certain cancer cells, making them good candidates as drug delivery agents and tumor therapeutics with fewer than normal side effects. This study is aimed to correlate cell proliferation inhibition with the size and morphology of nHAPs in a human breast cancer cell line as well as in normal tissue cells. We present our in vitro experimental evidence that nHAPs with sizes smaller than 50 nm have high inhibitory activity against human MCF-7 breast cancer cell lines. Based on our experimental data, normal fibroblast cells (NIH 3T3) were relatively more viable upon treatment with the nanoconstructs. The present study indicates that nHAPs can be engineered as nontoxic specific inhibitors as efficient breast cancer therapeutics in humans.

Antitumor and Antioxidant Activities of In Vitro Cultivated and Wild-Growing Clinopodium vulgare L. Plants.

Clinopodium vulgare L. is a valuable medicinal plant used for its anti-inflammatory, antibacterial and wound-healing properties. The present study describes an efficient protocol for the micropropagation of C. vulgare and compares, for the first time, the chemical content and composition and antitumor and antioxidant activities of extracts from in vitro cultivated and wild-growing plants. The best nutrient medium was found to be Murashige and Skoog (MS) supplemented with 1 mg/L BAP and 0.1 IBA mg/L, yielding on average 6.9 shoots per nodal segment. Flower aqueous extracts from in vitro plants had higher total polyphenol content (29,927.6 ± 592.1 mg/100 g vs. 27,292.8 ± 85.3 mg/100 g) and ORAC antioxidant activity (7281.3 ± 82.9 µmol TE/g vs. 7246.3 ± 62.4 µmol TE/g) compared to the flowers of wild plants. HPLC detected qualitative and quantitative differences in phenolic constituents between the in vitro cultivated and wild-growing plants' extracts. Rosmarinic acid was the major phenolic constituent, being accumulated mainly in leaves, while neochlorogenic acid was a major compound in the flowers of cultivated plants. Catechin was found only in cultivated plants, but not in wild plants or cultivated plants' stems. Aqueous extracts of both cultivated and wild plants showed significant in vitro antitumor activity against human HeLa (cervical adenocarcinoma), HT-29 (colorectal adenocarcinoma) and MCF-7 (breast cancer) cell lines. The best cytotoxic activity against most of the cancer cell lines, combined with the least detrimental effects on a non-tumor human keratinocyte cell line (HaCaT), was shown by the leaf (250 µg/mL) and flower (500 µg/mL) extracts of cultivated plants, making cultivated plants a valuable source of bioactive compounds and a suitable candidate for anticancer therapy.

Synthesis of Magnetic Nanocarrier Conjugated by Folate Based on Tragacanth and In Vitro Investigation of their Efficiency on Breast Cancer Cells

AbstractIn this research, a novel nanocarrier is synthesized based on tragacanth as a natural polysaccharide. For this purpose, carboxymethyl tragacanth (CMT) is first synthesized from tragacanth and then it is functionalized with isocyanate (IS) groups. In another parallel synthesis, magnetite nanoparticles (MNPs) are prepared by the co‐precipitation method and coated with SiO2 to form MNPs coated SiO2 (MNPs@SiO2). Then MNPs@SiO2 is reacted with CMT‐IS to form Fe3O4@SiO2/CMT. In the second step, the MNPs@SiO2/CMT surface is aminated by reaction with 3‐amino propyl triethoxy silane (APTES) and then it is reacted with folic acid (FA) as a targeting agent to form MNPs@SiO2/CMT/FA nanocarrier. The synthesized nanocarrier is characterized by FT‐IR, SEM, EDX, TEM, XRD, VSM, TGA, and zeta potential analysis and then it is loaded with doxorubicin (DOX) as a typical anti‐cancer drug. The in vitro drug release studies are carried out from the DOX‐loaded nanocarrier. The pH sensitivity of the DOX‐loaded nanocarrier is examined attwo different pHs. The effect of DOX‐loaded nanocarrier is investigated on MCF7 human breast cancer cells and the rate of cancer cell death is determined by MTT assay and drug IC50. The changes in the morphology of the tumor cell nuclei are observed using fluorescence microscopy.

N-(benzazol-2-yl)-2-substituted phenylacetamide derivatives: Design, synthesis and biological evaluation against MCF7 breast cancer cell line

This work describes the straightforward and efficient one-pot synthesis of a new library of N-(benzazol-2-yl)-2-substituted phenylacetamide derivatives (19–27). Using the MTT assay, these compounds were evaluated for their in vitro anticancer activity against the MCF7 human breast cancer cell line, and the results were compared to the standard doxorubicin. The majority of compounds exhibited an inhibitory effect against the cancer cell line, with compounds 19, 22, and 26 exhibiting exceptional cytotoxicity against MCF7 cells. Using flow cytometry, the most potent compound 19 on the induction of apoptosis in the breast cancer cell line was determined. Compound 19 induced G1-phase cell cycle arrest followed by apoptotic cell death. In silico analyses of potent compounds 19, 22, and 26 were conducted to investigate their interactions with Human DNA topoisomerase II. The energy calculations were found to be in excellent agreement with the calculated IC50 values. In addition, drug similarity parameter values for the three active compounds were determined using in silico ADME prediction studies. Considering all of these results, it appears that these N-(benzazol-2-yl)-2-substituted phenylacetamide derivatives may be effective anticancer agents. This work may possibly generate new concepts for the enhancement of inhibitors of human DNA topoisomerase II for breast cancer treatment.

Abstract 3621: Identification of podocalyxin-positive tumor buds in pancreatic ductal adenocarcinoma: Implications for solid tumor collective invasion

Abstract High expression of the single-pass transmembrane sialomucin, podocalyxin, has been shown to predict poor disease outcome in a number of solid tumor types, including pancreatic ductal adenocarcinoma (PDAC) (Taniuchi et al., 2016). In addition to the classical TMN staging measures of disease progression (tumor size, lymph node involvement, and distant metastasis) local invasion at the primary tumor site is a valuable indicator of solid tumor progression. Local invasion has been observed histologically as tumor buds, defined by the presence of small cohesive clusters of tumor cells in the invasive tumor-stromal interface. To date, tumor budding has been characterized as an indicator of poor prognostic outcome in colorectal carcinoma (CRC), pancreatic ductal adenocarcinoma (PDAC), invasive ductal breast carcinoma (IDC), lung adenocarcinoma (LUA), stomach and esophageal carcinomas, oral squamous cell carcinoma, periampullary adenocarcinoma, and head and neck squamous cell carcinoma (HNSCC). Our group has previously modelled tumor budding experimentally using the MCF7 human breast cancer cell line (Graves et al., 2016). We showed that forced over-expression of podocalyxin was a driver of local collective invasion and tumor budding in vitro and in vivo. Because podocalyxin is highly expressed in PDAC we sought to determine if its expression was involved in the process of tumor budding in this tumor context as well. We assessed large format histological sections from a cohort of patient PDAC tumors and identified high podocalyxin expression in tumor buds. Further, we generated podocalyxin-null MiaPaCa2 PDAC cell lines and demonstrated that podocalyxin-expression is required for collective invasion in 3D culture conditions. We are currently evaluating in vivo xenograft tumor models with these podocalyxin-expressing and podocalyxin-null MiaPaCa2 cells to determine whether changes in 3D culture invasion are representative of tumor budding in this model. Expression of podocalyxin by tumor buds provides rationale to support investigation into the impact of therapeutically targeting these invasive cells. Citation Format: Erin M. Bell, Steve E. Kalloger, Pamela M. Dean, Janel L. Kopp, Kelly M. McNagny, Calvin D. Roskelley. Identification of podocalyxin-positive tumor buds in pancreatic ductal adenocarcinoma: Implications for solid tumor collective invasion. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3621.

Abstract 2397: An advanced approach technique for tracking point between immune check point inhibitor and cancer cell mechanism in immune response motility by infiltration a breast cancer cell

Abstract In Breast Cancer, Tumor-infiltrating lymphocyte (TIL) motility has been reported that it is reflecting tumor microenvironment response by adoptive cell therapy (ACT). ACT is well-known to be best applied for suppressing cancer cell growth with immune response and it also was well-known the immunotherapy. However, ACT response tracking by general analysis method included immunohistochemistry and flow cytometry was provided limited information for cancer therapy in recent years. Therefore, by assessing relationship immunotherapy marker, PD-L1, known as immune check point inhibitor and TIL using intravital microscopy in breast cancer model, we also imaged whether TIL response was related to immunotherapy following dependent on time point in a various breast cancer cell implanted mouse model such as 4T1 (mouse breast cancer cell), MDA-MB-231 (human triple negative breast cancer cell) and MCF7 (human breast cancer cell). These images result in PD-L1 expression was decreased in cancer cell implanted until 10days, whereas TIL expression increased in a various breast cancer cell when it is measuring displacement length, track mean velocity and confinement ratio called tumor microenvironment. These results indicated that we can imaged immune response related to TIL response in breast cancer cell line by intravital microscopy instead of general microscopy. It means that intravital microscope was an advanced approach technique to understand cancer mechanism accompanied by the immune response and established the method of high-resolution intravital cellular visualization compared to a general histopathological method for evaluating immune response in cancer tissue Citation Format: Hyunseok Kim, Soyeon Ahn, Hyungjin Kwon, Hsuan-Ju Chen, Inwon Park, Pilhan Kim, Akyildiz Kubra. An advanced approach technique for tracking point between immune check point inhibitor and cancer cell mechanism in immune response motility by infiltration a breast cancer cell [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2397.

Abstract 1331: A novel approach to treating hormonal breast cancer using clinical database and 3D ex vivo model

Abstract Type 2 diabetes (T2D) female patients have a higher risk of being diagnosed and have worse survival for breast cancer compared to non-diabetic females. Although the relationship between these two female diseases has been reported, specific functional studies on which subtypes of breast cancer are affected and which genes are regulated by T2D are little known. Here, I found that Forkhead Box A1 (FOXA1) and Metformin, anti-diabetic medication for T2D, affect hormonal positive (HR+) breast cancer tumor cell growth and metastasis. 14 diabetes-related genes highly expressed in three HR+ breast cancer cell lines but not other breast cancer subtypes using a 53,805 gene database obtained from NCBI GEO. Among the diabetes-related genes, FOXA1, MTA3, PAK4, FGFR3, and KIF22 were highly expressed in HR+ breast cancer from 4,032 breast cancer patient tissue samples using the Breast Cancer Gene Expression Omnibus. Specifically, the high expression of FOXA1 correlated to a worse ER+/PR+ breast cancer patient survival rate. Consistent with this, the loss of FOXA1 inhibited the tumor proliferation and invasion of MCF-7 and T47D HR+ breast cancer cell lines using in vitro. Metformin (1,1-dimethylbiguanide hydrochloride) conspicuously inhibits the tumor cell growth in MCF-7 human hormonal breast cancer cells. Metformin or FOXA1 deletion enhanced Tamoxifen-mediated tumor growth inhibition in HR+ breast cancer cell lines through the ex vivo three-dimensional (3D) organoid model. Therefore, HR+ breast cancer is closely related to T2D, and metformin and FOXA1 inhibition might be an optimal new treatment for patients with HR+ breast cancer when combined with tamoxifen, a hormone therapy. Citation Format: Chaeun (Christine) Song, SeungBaek Lee. A novel approach to treating hormonal breast cancer using clinical database and 3D ex vivo model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1331.

Metal organic framework supported surface modification of synthesized nickel/nickel oxide nanoparticles via controlled PEGylation for cytotoxicity profile against MCF-7 breast cancer cell lines via docking analysis

Metal organic framework, (MOFs) supported nanoparticles have proven a path for the treatment of various carcinomas to be effective against MCF-7 Breast cancer cell lines. Despite the globally widespread applications of nickel/nickel oxide nanoparticles, Ni/NiO@LAA_PEG NPs with surfactant Polyethylene glycol, (PEG) and l-Acsorbic acid, (LAA) as reducing agents in industrial, commercial, and biomedical fields, their response to human cells has not been elucidated. Hence in the reported article Ni/NiO@LAA_PEG NPs with an average size range of 75–80 nm was used to study their interaction with MCF-7 Breast cancer cell lines. So the best of our understanding this is the study related to MCF-7 Human Breast cancer cell lines to show cyto-genotoxicity of Ni/NiO@LAA_PEG NPs which seems mediated through MTT analysis with rising effect surface enhancement and indexed porosiveness for the interfacial advancement than standard drug Doxorubicin for their comparison to show their best effectivity to prepared Ni/NiO@LAA_PEG NPs.The theoretical performance was simulated using Gaussian-09@Autodock 4.2 with crystal preparation and docked with MCF-7 Human Breast cancer cell lines PDBs to show their correlation with wet method of preparation. The synthesized Ni/NiO@LAA_PEG NPs was characterized against UV/Vis spectroscopy for determination of peak pattern of 395 nm, the functional group analysis for the best binding of stretching with framed metal network of nanoparticles was to shown out their scissoring, wagging, and CH stretching, etc. at 661, 834 and 3425 cm−1respectively by FTIR analysis and an energy band gap of 2.2 eV for Ni/NiO@LAA_PEG NPs by optical studies. The mesopore range and surface area measurement was calculated with BET adsorption and BJH plot to find their coercivity and retentivity via adsorption-desorption curve for their best catalytic performance. The current interpretation of Ni 2p spectra of oxides and other compounds is based on charge transfer assignment of the main peak at 856.4 eV and broad satellite cantered at about 861 eV to the cd9 L and unscreened cd8 final state configurations.

In-silico Molecular Docking and ADMET predictions of Pyrido[2,3-d]pyrimidine-2,4(1H,3H)-Dione Analogues as promising Antimicrobial, Antioxidant and Anticancer agents

Pyridopyrimidine are heterocyclic molecules enclosing fused pyridine and pyrimidine rings. Owing to its fascinating core structure and pharmacological applications a series of 7-([1,1′-biphenyl]-4-yl)-5-arylpyrido[2,3-d]pyrimidine-2,4(1H,3H)-diones were synthesized and characterized using IR, 1H, 13C NMR and Mass spectral techniques. The antibacterial, antioxidant and anticancer activities were investigated for the synthesized compound 5a-5f. Compounds with electron-donating groups showed excellent free radical scavenging activity. Halogen-substituted compounds showed more potent antimicrobial and anticancer activity than other derivatives in comparison with their respective standards. Based on the IC50 value obtained from anticancer activity, 5c was further analyzed for apoptosis by AO/EB staining method. The findings suggested early apoptosis in the MCF-7 cell line. Molecular Docking studies of the synthesized compounds were performed with Kinase 1 inhibitors (PDB id: 2YEX), 5c exposed good docking results with minimum binding energy. Further, these compounds were acknowledged as orally active drug candidates from in-silico ADMET studies. Computational analysis supports biological findings indicating compound 5c as a promising anticancer agent against the human breast cancer MCF-7 cell line.

An In Vitro Antimicrobial, Anticancer and Antioxidant Activity of N–[(2–Arylmethylthio)phenylsulfonyl]cinnamamide Derivatives

Cinnamic acid is a plant metabolite with antimicrobial, anticancer, and antioxidant properties. Its synthetic derivatives are often more effective in vitro than parent compounds due to stronger biological activities. In our study, we synthesized ten new N-(4-chloro-2-mercapto-5-methylphenylsulfonyl)cinnamamide derivatives, containing two pharmacophore groups: cinnamic acid moiety and benzenesulfonamide. The antimicrobial activity of the obtained compounds was estimated using different types of Gram-positive and Gram-negative bacteria, fungus species of Candida albicans, as well as clinical strains. The compounds were evaluated on biofilm formation and biofilm formed by Staphylococcus clinical strains (methicillin-resistance S. aureus MRSA and methicillin-resistance coagulase-negative Staphylococcus MRCNS). Furthermore, blood bacteriostatic activity test was performed using S. aureus and S. epidermidis. In cytotoxic study, we performed in vitro hemolysis assay on domestic sheep peripheral blood and MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay on human cervical HeLa, ovarian SKOV-3, and breast MCF-7 cancer cell lines. We also estimated antioxidant activity of ten compounds with 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) assays. Our results showed a significant antimicrobial activity of the compounds. All of them were active on Staphylococcus and Enterococcus species (MIC was 1-4 µg/mL). The compounds 16d and 16e were the most active on staphylococci clinical strains and efficiently inhibited the biofilm formation and biofilm already formed by the clinical staphylococci. Moreover, the hemolytic properties of the tested compounds occurred in higher quantities (>32.5 µg/mL) than the concentrations that inhibited both the growth of bacteria in the blood and the formation and growth of biofilm. The results of MTT assay showed that compounds 16c, 16d, 17a, and 17d demonstrated the best activity on the cancer cells (the IC50 values were below 10 µg/mL). Compound 16f was the least active on the cancer cells (IC50 was > 60 µg/mL). Antiradical tests revealed that compounds 16f and 17d had the strongest antioxidant properties within the tested group (IC50 was 310.50 ± 0.73 and 574.41 ± 1.34 µg/mL in DPPH, respectively, and 597.53 ± 1.3 and 419.18 ± 2.72 µg/mL in ABTS assay, respectively). Our study showed that the obtained cinnamamide derivatives can be used as potential antimicrobial therapeutic agents.

The Effect of Cholesterol in MCF7 Human Breast Cancer Cells

In the last decade, cholesterol level has been implicated in several types of cancer, including breast cancer. In the current study, we aimed to investigate the condition of lipid depletion, hypocholesterolemia or hypercholesterolemia reproduced in vitro to analyze the response of different human breast cancer cells. Thus, MCF7 as the luminal A model, MB453 as the HER2 model and MB231 as the triple-negative model were used. No effect on cell growth and viability was detected in MB453 and MB231 cells. In MCF7 cells, hypocholesterolemia (1) reduced cell growth, and Ki67 expression; (2) increased ER/PgR expression; (3) stimulated the 3-Hydroxy-3-Methylglutaryl-CoA reductase and neutral sphingomyelinase and; (4) stimulated the expression of CDKN1A gene coding cyclin-dependent kinase inhibitor 1A protein, GADD45A coding growth arrest and DNA-damage-inducible alpha protein and, PTEN gene coding phosphatase and tensin homolog. All these effects were exacerbated by the lipid-depleted condition and reversed by the hypercholesterolemic condition. The relationship between cholesterol level and sphingomyelin metabolism was demonstrated. In summary, our data suggest that cholesterol levels should be controlled in luminal A breast cancer.

Establishment and verification of a digital PCR assay for the detection of CK19 expression in quantitative analysis of circulating tumor cell.

The objective of this study was to establish and verify a digital PCR assay for the detection of CK19 gene expression, and to use it to detect circulating tumor cells (CTC) by taking advantages of its ultra-high sensitivity and absolute quantitation. Firstly, the primers and probes were designed according to the mRNA sequence of CK19 gene, and housekeeping gene ABL1 was used as the internal control. The best candidate was screened by human breast cancer MCF7 cells and healthy human leukocytes from 13 sets of primer and probes and verified by direct sequencing. Secondly, after the reaction conditions of the selected primers and probes were optimized, limit of blank (LOB) analysis were performed with different concentrations of cDNAs as templates from healthy human leukocytes. The results revealed the LOB of CK19 with ABL1 copy numbers of 20,000, 15,000, 10,000, 5000 and 2500 were 9.24, 8.93, 3.12, 3.17 and 2.53 copies, respectively. Thirdly, the different concentrations of cDNAs from MCF7 cells and healthy human leukocytes were premixed and used in the limit of detection (LOD) analysis, which showed that the CK19 gene could be effectively detected at the concentration ratio of 50%, 10%, 5%, 1%, 0.5% and 0.1%, and the linear R2 value was 0.9998. Finally, the preliminary results of digital PCR in clinical samples indicated that CK19 copy numbers were higher in advanced breast cancer patients than healthy controls. The above results demonstrated the advantages of our CK19 digital PCR assay in sensitivity, specificity, and accurate quantification. If verified further, the assay is expected to play significant roles in the quantitative analysis of CTC in breast cancer with a good application prospect.

Cytotoxic potential activity of quercetin derivatives on MCF-7 breast cancer cell line

Many previous investigations have found quercetin to be a powerful antioxidant and antitumor flavonoid, but its poor bioavailability has limited its use. This current study investigated the effects of two newly synthesized Quercetin Schiff bases containing 2-amino thiadiazole-5-thiol (Q1), and its benzyl derivatives (Q2) on MCF-7 human breast cancer cells. Cell viability and apoptosis were assessed to determine the toxic effects of Q1 and Q2. Cytotoxicity valuation showed that both compounds inhibited MCF-7 cell growth, and lactate dehydrogenase (LDH) activity increased in a dose-dependent aspect compared to the control group. Comet assay results observed that Q1 and Q2 induce more serious DNA damage than the control (untreated cell); however, in all curried experiments, Q2 showed higher effects than Q1. Hence two synthesized quercetin Schiff bases can take action as a promising anticancer agent. Keywords: quercetin derivatives, Schiff base, breast cancer, MCF-7 Cytotoxic.

Senescent Secretome of Blind Mole Rat Spalax Inhibits Malignant Behavior of Human Breast Cancer Cells Triggering Bystander Senescence and Targeting Inflammatory Response

Subterranean blind mole rat, Spalax, has developed strategies to withstand cancer by maintaining genome stability and suppressing the inflammatory response. Spalax cells undergo senescence without the acquisition of senescence-associated secretory phenotype (SASP) in its canonical form, namely, it lacks the main inflammatory mediators. Since senescence can propagate through paracrine factors, we hypothesize that conditioned medium (CM) from senescent Spalax fibroblasts can transmit the senescent phenotype to cancer cells without inducing an inflammatory response, thereby suppressing malignant behavior. To address this issue, we investigated the effect of CMs of Spalax senescent fibroblasts on the proliferation, migration, and secretory profile in MDA-MB-231 and MCF-7 human breast cancer cells. The results suggest that Spalax CM induced senescence in cancer cells, as evidenced by increased senescence-associated beta-galactosidase (SA-β-Gal) activity, growth suppression and overexpression of senescence-related p53/p21 genes. Contemporaneously, Spalax CM suppressed the secretion of the main inflammatory factors in cancer cells and decreased their migration. In contrast, human CM, while causing a slight increase in SA-β-Gal activity in MDA-MB-231 cells, did not decrease proliferation, inflammatory response, and cancer cell migration. Dysregulation of IL-1α under the influence of Spalax CM, especially the decrease in the level of membrane-bound IL1-α, plays an important role in suppressing inflammatory secretion in cancer cells, which in turn leads to inhibition of cancer cell migration. Overcoming of SASP in tumor cells in response to paracrine factors of senescent microenvironment or anti-cancer drugs represents a promising senotherapeutic strategy in cancer treatment.

Effects of siRNA-mediated Silencing of ERBB2, IGF-1R, and ITGB1 in HER2-positive Breast Cancer Cells.

One of the hallmarks of cancer is deregulation of multiple signaling pathways, which can lead to uncontrolled proliferation and migration of cells. Over-expression and mutations in human epidermal growth factor receptor 2 (HER2) can lead to overactivation of these pathways, potentially developing cancer in different tissues, including breast tissue. IGF-1R and ITGB-1 are two receptors that have been linked to cancer development. Therefore, the aim of this study was to investigate the effects of silencing of the corresponding genes using specific siRNAs. Transient silencing of HER2, ITGB-1, and IGF-1R was conducted using siRNAs and expression was quantified by reverse transcription-quantitative polymerase chain reaction. Viability in human breast cancer cells SKBR3, MCF-7, and HCC1954 and cytotoxicity in HeLa cells were tested using WST-1 assay. The use of anti-HER2 siRNAs in a breast cancer cell line over-expressing HER2 (SKBR3) led to a decrease in cell viability. However, silencing of ITGB-1 and IGF-1R in the same cell line had no significant effects. Silencing of any of the genes encoding any of the three receptors in MCF-7, HCC1954, and HeLa had no significant effects. Our results provide evidence towards using siRNAs against HER2-positive breast cancer. Silencing of ITGB-1 and IGF-R1 did not significantly inhibit the growth of SKBR3 cells. Therefore, there is need for testing the effect of silencing ITGB-1 and IGF-R1 in other cancer cell lines over-expressing these biomarkers and explore their potential use in cancer therapy.

Abstract P6-14-17: Endomucins and their expression in breast cancer and the cellular and therapeutic impact

Abstract Introduction. Endomucin-1 and endomucin-2 are two proteins structurally related, yet with low homology. The membrane bound sialoglycoproteins appear to play a key role in interfering with the formation of focal adhesion complexes (FAC) and matrix adhesiveness of cells, by mechanisms independent of the MUC1 repeat, which the endomucins do not possess. Endomucins are thought to be expressed at high levels in endothelium and haematopietic cell lineages, although the levels in mammary tissues also seem high. Despite the seeming importance of endomucins in the adhesion and migration of cells, including cancer cells, the clinical value of endomucin in clinical cancer, including breast cancer, is largely unknown. The present study examined the expression profile of endomucins, together with the focal adhesion kinase FAK, in breast cancer and aimed to explore the cellular impact of endomucin on cancer cells. Methods. Human breast cancer cells MCF-7 and MDA MB-231 and a range of other cell types were used. An endomucin overexpression cell model was created and subsequently used to evaluate the function of the cells. The expression profile of the endomucin-1 and endomucin-2 transcripts and FAK, in an existing fresh frozen breast cancer tissue cohort, were quantified. Results. High levels of endomucins, particularly endomucin-1, are good indicators for the overall survival of the patient, p=0.021 for endomucin-1 and p=0.15 for endomucin-2. When expression levels of FAK were integrated into the survival analysis model, patients with high levels of both endomucins and low levels of FAK had the most favourable outcome, compared with those with most unfavourable outcome who had low level of endomucin and high FAK (survival during the follow up period respectively at 100% and 54%, p=0.013, Harzoud Ratio 0.298). Together with the Nottingham Prognostic Index, which independently predicts a poor outcome (p=0.009, HR=7.6), the integrated expression profile of endomucin/FAK represents an independent prognostic indicator for favourable overall survival (p=0.003, HR=0.13), and indeed for a favourable disease free survival (p=0.008, HR=0.17). Mammary tissues, and indeed breast cancer cell lines, expressed high levels of endomucin-2 transcripts and low levels of endomucin-1 transcripts. High levels of endomucin-2 were also seen in fibroblasts and vascular endothelial cells. We created a breast cancer cell submodel with MCF-7, by overexpressing endomucin. It was shown that although endomucin over-expression had some marginal impact on the adhesiveness of breast cancer cells, the over-expression however, had significant impact on cells’ sensitivity to FAK inhibitor, with a markedly reduced adhesiveness to matrix (p< 0.001 control versus endomucin overexpression cells). Discussion. Endomucins have a reduced expression in breast cancers and the reduction, together with low levels of focal adhesion kinase, facilitate a favourable outcome for the patients. Together with the findings of in vitro cell models, it would suggest that the expression profile of endomucins and FAK may be a good indicator, not only for evaluating clinical outcomes, but also for choice of target therapies. Citation Format: Amber Xinyu Li, Tracey A. Martin, Lin Ye, Andrew J. Sanders, Fiona Ruge, Jane Lane, Eleri Davies, Wen G. Jiang. Endomucins and their expression in breast cancer and the cellular and therapeutic impact [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P6-14-17.

Retracted: Antiproliferative and apoptotic activity of glycyrrhizinic acid in MCF-7 human breast cancer cells and evaluation of its effect on cell cycle, cell migration and m-TOR/PI3K/Akt signalling pathway.

[This retracts the article DOI: 10.5114/aoms.2018.79429.].

Abstract P6-12-02: Connecting Fluctuations in Estrogen Production, Release, and Action with Hormone-Dependent Gene Expression in Breast Cancers

Abstract Estrogens are steroid hormones that play a key role in a wide range of physiological and pathological processes. Estrogen receptor-alpha, ERɑ, functions primarily as a nuclear transcription factor (TF) through ligand-dependent activation by 17β-estradiol (E2), the predominant naturally-occurring estrogen. This results in dimerization, DNA binding to cis-regulatory elements called enhancers, and gene regulatory function of ERɑ. In vivo, E2 production and action fluctuates during reproductive cycles and possibly even in a circadian manner. In tissues and cells, E2-regulated gene expression is dynamic, with rapid induction of enhancer formation and target gene expression (or repression), followed by enhancer “decommissioning” and cessation of the gene regulatory effects. The effects of sustained E2 production and signaling, fluctuations in E2 levels, and removal of E2 on estrogen-regulated gene expression are unknown. Classical experiments exploring the gene regulatory effects of E2 involve stimulation of cultured cells throughout the duration of the experiment with a continuous and typically high dose of hormone, with the cells harvested at the end of the stimulation period. While this has allowed us to understand the molecular underpinnings of signal-dependent transcription, it has not allowed analysis of physiological fluctuations in estrogen signaling involving periods of stimulation (i.e., the presence of hormone) and non-stimulation (i.e., the absence of hormone). In our studies, we are applying a combination of next generation sequencing techniques and fluorescence microscopy to investigate signal-dependent responses to fluctuating and short duration E2 exposures using ERɑ-positive MCF-7 human breast cancer cells after E2 treatment and removal. Our initial results suggest that different E2 target genes respond differently to low dose E2 treatment and E2 withdrawal. Moreover, for some E2 target genes, sustained E2 treatment may not be required for a complete E2 transcriptional response. With these studies, we hope to connect physiological aspects of E2 production, release, and action with the molecular mechanisms of E2-dependent gene expression - an aspect of our understanding of estrogen signaling that has been lacking. Citation Format: Hyung Bum Kim, William L. Kraus. Connecting Fluctuations in Estrogen Production, Release, and Action with Hormone-Dependent Gene Expression in Breast Cancers [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P6-12-02.