Soy phytoestrogens, such as daidzein and its metabolite equol, have been proposed to be
responsible for the low breast cancer rate in Asian women. Since the majority of estrogen
receptor positive breast cancer patients are treated with tamoxifen, the basic objective of this
study is to determine whether equol enhances tamoxifen’s anti-tumor effect, and to identify
the molecular mechanisms involved.
The document evaluates the potential of using statins to treat ovarian cancer. It finds that several statins inhibit the growth of ovarian cancer cell lines and spheroids, with simvastatin showing particular effectiveness. This anti-cancer activity is mediated through the mevalonate pathway and results in apoptosis. While statins reduced autophagy in some cell lines, their effects on autophagy appear complex. Combining simvastatin with chemotherapy showed antagonism, particularly with pre-treatment of simvastatin, suggesting it may be best to evaluate statins as single agents in clinical trials at high doses.
1. Several Pancratistatin (PST) analogs, including SVTH-7, SVTH-6, and SVTH-5, were found to have potent anti-cancer activity greater than PST and standard chemotherapeutics in a medium-throughput screen of various cancer cell lines.
2. The PST analogs disrupted mitochondrial function, activated the intrinsic apoptotic pathway, and reduced tumor growth in vivo. Inhibition of mitochondrial complexes II and III abrogated the pro-apoptotic effects of SVTH-7, suggesting it exploits a mitochondrial vulnerability.
3. This work identifies several PST analogs with high therapeutic potential and provides insight into distinct mitochondrial features of cancer
Calcarea carbonica induces apoptosis in cancer cells in p53-dependent manner ...home
These observations delineate the significance of immuno-modulatory circuit during calcarea carbonicamediated
tumor apoptosis. The molecular mechanism identified may serve as a platform for involving calcarea
carbonica into immunotherapeutic strategies for effective tumor regression
NMR spectroscopy techniques including 2D TOCSY, ROESY, and STD NMR were used to analyze peptide epitopes and determine their binding to monoclonal antibodies. 2D NMR was used to assign proton signals in peptide epitopes. STD NMR showed that the peptide GVTSAX3D binds to antibody SM3 through interactions of the 3-aminobenzoate substitution and methyl groups of valine, threonine, and alanine residues. Mass spectrometry and NMR spectroscopy supported the successful synthesis and characterization of peptide epitopes for studying antibody binding.
As the pioneer of spin-column based protein extraction/isolation technologies, we have successfully developed our Minute™ series extraction kits products. With years of research and development, we have discovered the fastest, easiest and most reliable ways for protein extraction and cell fractionation. For more information about Invent Biotechnologies products, please call us at 952-400-6898 or email at info@inventbiotech.com.
The National Mouse Metabolic Phenotyping Center (MMPC) at UMass provides tools for investigating transgenic mouse models of obesity, diabetes, and related complications. It consists of three phenotyping cores that assess metabolic function in mice: 1) the Metabolism Core performs non-invasive metabolic experiments; 2) the Analytical Core utilizes clinical chemistry and Luminex to measure hormones, metabolites, and cytokines; 3) the Cardiovascular Core applies imaging to assess cardiac and vascular structure and function.
This study assessed toxicities of CMF (cyclophosphamide, methotrexate, and 5-fluorouracil) chemotherapy and tamoxifen on rabbit livers and kidneys. Rabbits received either CMF alone, tamoxifen alone, or CMF plus tamoxifen. Liver enzymes and histopathology of livers and kidneys were analyzed. Rabbits that received both CMF and tamoxifen showed the highest liver enzyme levels and most histological changes in the liver including edema, infiltration, and fibrosis. They also exhibited the most kidney mesangial cell proliferation. The study concludes that CMF and tamoxifen chemotherapy can cause toxic effects in the liver and kidney.
The document describes research into creating an antibody-drug conjugate (ADC) to treat metastatic triple negative breast cancer. Key points:
- Researchers developed an antibody that recognizes HSPG2, a biomarker expressed on metastatic breast cancer cells, and aims to link this antibody to the chemotherapy drug paclitaxel.
- The ADC would be synthesized by modifying the antibody with thiol groups and paclitaxel with maleimide groups, allowing them to conjugate.
- While the antibody thiolation was successful, complications arose in activating paclitaxel for conjugation. Other drug molecules will be explored to complete the ADC synthesis.
The document evaluates the potential of using statins to treat ovarian cancer. It finds that several statins inhibit the growth of ovarian cancer cell lines and spheroids, with simvastatin showing particular effectiveness. This anti-cancer activity is mediated through the mevalonate pathway and results in apoptosis. While statins reduced autophagy in some cell lines, their effects on autophagy appear complex. Combining simvastatin with chemotherapy showed antagonism, particularly with pre-treatment of simvastatin, suggesting it may be best to evaluate statins as single agents in clinical trials at high doses.
1. Several Pancratistatin (PST) analogs, including SVTH-7, SVTH-6, and SVTH-5, were found to have potent anti-cancer activity greater than PST and standard chemotherapeutics in a medium-throughput screen of various cancer cell lines.
2. The PST analogs disrupted mitochondrial function, activated the intrinsic apoptotic pathway, and reduced tumor growth in vivo. Inhibition of mitochondrial complexes II and III abrogated the pro-apoptotic effects of SVTH-7, suggesting it exploits a mitochondrial vulnerability.
3. This work identifies several PST analogs with high therapeutic potential and provides insight into distinct mitochondrial features of cancer
Calcarea carbonica induces apoptosis in cancer cells in p53-dependent manner ...home
These observations delineate the significance of immuno-modulatory circuit during calcarea carbonicamediated
tumor apoptosis. The molecular mechanism identified may serve as a platform for involving calcarea
carbonica into immunotherapeutic strategies for effective tumor regression
NMR spectroscopy techniques including 2D TOCSY, ROESY, and STD NMR were used to analyze peptide epitopes and determine their binding to monoclonal antibodies. 2D NMR was used to assign proton signals in peptide epitopes. STD NMR showed that the peptide GVTSAX3D binds to antibody SM3 through interactions of the 3-aminobenzoate substitution and methyl groups of valine, threonine, and alanine residues. Mass spectrometry and NMR spectroscopy supported the successful synthesis and characterization of peptide epitopes for studying antibody binding.
As the pioneer of spin-column based protein extraction/isolation technologies, we have successfully developed our Minute™ series extraction kits products. With years of research and development, we have discovered the fastest, easiest and most reliable ways for protein extraction and cell fractionation. For more information about Invent Biotechnologies products, please call us at 952-400-6898 or email at info@inventbiotech.com.
The National Mouse Metabolic Phenotyping Center (MMPC) at UMass provides tools for investigating transgenic mouse models of obesity, diabetes, and related complications. It consists of three phenotyping cores that assess metabolic function in mice: 1) the Metabolism Core performs non-invasive metabolic experiments; 2) the Analytical Core utilizes clinical chemistry and Luminex to measure hormones, metabolites, and cytokines; 3) the Cardiovascular Core applies imaging to assess cardiac and vascular structure and function.
This study assessed toxicities of CMF (cyclophosphamide, methotrexate, and 5-fluorouracil) chemotherapy and tamoxifen on rabbit livers and kidneys. Rabbits received either CMF alone, tamoxifen alone, or CMF plus tamoxifen. Liver enzymes and histopathology of livers and kidneys were analyzed. Rabbits that received both CMF and tamoxifen showed the highest liver enzyme levels and most histological changes in the liver including edema, infiltration, and fibrosis. They also exhibited the most kidney mesangial cell proliferation. The study concludes that CMF and tamoxifen chemotherapy can cause toxic effects in the liver and kidney.
The document describes research into creating an antibody-drug conjugate (ADC) to treat metastatic triple negative breast cancer. Key points:
- Researchers developed an antibody that recognizes HSPG2, a biomarker expressed on metastatic breast cancer cells, and aims to link this antibody to the chemotherapy drug paclitaxel.
- The ADC would be synthesized by modifying the antibody with thiol groups and paclitaxel with maleimide groups, allowing them to conjugate.
- While the antibody thiolation was successful, complications arose in activating paclitaxel for conjugation. Other drug molecules will be explored to complete the ADC synthesis.
1) Plumbagin (PL), derived from plants, inhibits prostate tumor development in mice lacking the tumor suppressor PTEN (Pten-KO mice), a model for prostate cancer.
2) PL treatment decreased expression of proteins involved in prostate cancer progression (PKCε, STAT3, AKT) and epithelial-to-mesenchymal transition (EMT).
3) Dietary PL inhibited primary prostate tumor growth and castration-resistant prostate cancer growth in Pten-KO mice, potentially by inhibiting PKCε, STAT3, AKT, and EMT markers.
This document evaluates the effects of tamoxifen and paclitaxel (taxol) in treating breast cancer. It begins with background on breast anatomy and cancer types. Tamoxifen is a selective estrogen receptor modulator that blocks estrogen's effects in breast tissue. Paclitaxel disrupts microtubule function, inhibiting cell division. The document reviews literature on paclitaxel's mechanism of action and structure. It will present results from a study on tamoxifen and paclitaxel's effects, and discuss methodology, interpretation and conclusions.
Current Perspective of Natural Alkaloid Carbazole and its Derivatives as Anti...Hanif Shaikh
This document summarizes research on natural and synthetic carbazole alkaloids and their potential as antitumor agents. Several studies are described that tested various carbazole derivatives for cytotoxicity against different cancer cell lines. Many of the carbazole compounds showed cytotoxicity in the low micromolar or nanomolar range against cell lines like MCF-7, A549, HCT-116, and others. The carbazoles are proposed to exert their antitumor effects through mechanisms like DNA intercalation or inhibition of DNA-dependent enzymes. The review concludes that natural carbazole alkaloids and their derivatives show promise as antitumor agents and warrant further investigation.
Propolis with its active component CAPE (Caffeic Acid Phenethyl Ester) stops breast cancer cell growth. These results of CAPE are present in the naturopathic formulation
of propolis, a widely available natural substance with an extended safety record, making it a naturally-occurring and readily available epigenetic agent with great potential in breast cancer and oncology in general. The ability to link the biological effects of a naturopathic remedy to the pharmacologic effects seen with an exciting class of drugs in the treatment of cancer opens the door to a host of new therapeutic opportunities for patients.
This document analyzes the kinetic and mechanical properties of interactions between MUC16 and PODXL (glycoproteins expressed on pancreatic cancer cells) and E- and L-selectins. Single molecule force spectroscopy was used to characterize the binding interactions and determine that MUC16- and PODXL-E-selectin interactions are mechanically stronger than the respective L-selectin interactions. Microfluidic assays were also used to study the interactions under flow, finding that the single molecule properties and contact time regulate rolling behavior on selectin surfaces under shear stress. Understanding the biophysics of these selectin-ligand interactions could aid in developing diagnostic tools and preventing cancer metastasis.
A New Platform for Combining the ‘Bottom-Up’ PBPK Paradigm and POPPK Data Ana...mjamei
The document discusses combining bottom-up physiologically-based pharmacokinetic (PBPK) modeling with top-down population pharmacokinetic (POPPK) data analysis. It describes current and previous Simcyp team members and grants received. It then contrasts top-down and bottom-up approaches, highlighting how the bottom-up approach can incorporate more physiological covariates compared to the empirical top-down approach. The document outlines Simcyp's parameter estimation module, which allows fitting simulations to clinical data to estimate population and drug parameters.
Activation of AMPK inhibits cervical cancer cell growth through AKT/FOXO3a/FO...Enrique Moreno Gonzalez
Although advanced-stage cervical cancer can benefit from current treatments, approximately 30% patients may fail after definitive treatment eventually. Therefore, exploring alternative molecular therapeutic approaches is imperatively needed for this disease. We have recently shown that activation of AMP-activated protein kinase (AMPK), a metabolic sensor, hampers cervical cancer cell growth through blocking the Wnt/β-catenin signaling activity. Here, we report that activated AMPK (p-AMPK) also inhibits cervical cancer cell growth by counteracting FOXM1 function.
Background and objectives: Iron is one of the major component
of hemoglobin, is required for transport of oxygen, ferritin is a
protein for storage iron, and transferring is a protein for iron
transport, all these it may be change in breast cancer.
The aim of present study was to measure the serum ferritin, iron,
total iron binding capacity, transferring, and C-reactive protein
concentration in breast tumors.
Material and method: A prospective study was carried out from
April 2013 to August 2014 by clinical biochemistry department in
College of Pharmacy-University of Sulaimani on (45) healthy
female individuals, (group 1) and (50) females with breast tumor
(group 2).
Results: The mean value of serum iron, transferring, total iron
binding capacity were significantly lower in females with breast
tumors (group 2), than that of healthy female individuals, (group
1), while serum ferritin was significantly higher in females with
breast tumors (group2), than that of healthy individuals (group
1).
Conclusion: Based on findings of the present study it can be
concluded that breast tumors can cause deficient of all iron
profile except ferritin will be increase in female breast cancers.
Key words- Serum iron, S.Ferritin, S. Total iron binding, S.
Transferrin, S.C-Reactive protein, breast tumors.
IPI-549 is a potent and selective inhibitor of PI3K-γ that has been shown to inhibit tumor growth in mouse models. IPI-549 blocks macrophage M2 polarization and migration in vitro. In mouse tumor models, IPI-549 treatment reduced tumor growth, decreased tumor-associated M2 macrophages and myeloid cells, and increased tumor-associated CD8+ T cells in a T cell-dependent manner. Combination of IPI-549 with checkpoint inhibitors showed enhanced anti-tumor effects over monotherapy in mouse models. These results suggest that targeting PI3K-γ with IPI-549 can alter the immune suppressive tumor microenvironment to support immune-mediated tumor growth inhibition.
This study evaluated the effects of Amgen 386, a novel angiopoietin inhibitor, alone and in combination with a c-MET inhibitor (Compound A) on metastasis and survival in a patient-derived xenograft model of clear cell renal cell carcinoma. Treatment with Amgen 386 alone and in combination with Compound A significantly reduced the number and size of lung metastases and prolonged survival compared to the control. The combination treatment was more effective at reducing metastases than either drug alone, suggesting potential clinical benefit from combination therapy for treating metastatic kidney cancer.
This document summarizes a study examining EphB4 as a potential therapeutic target in mesothelioma. The study found that EphB4 is overexpressed in 72% of human mesothelioma tissue samples, particularly in epithelioid subtypes. Treatment with an EphB4 inhibitor called sEphB4-HSA significantly inhibited tumor growth in mesothelioma xenograft models by inducing apoptosis and inhibiting PI3K and Src signaling. Combination treatment with sEphB4-HSA and an anti-VEGF antibody was more effective than either treatment alone and led to complete tumor regression in some cases. These results suggest that targeting EphB4, particularly with s
Nanoparticle (NP) Delivery of Chemotherapy
Drugs to Prostate Cancer Patients by Toluleke Oloruntobi Famuyiwa* and James Kwasi Kumi-Diaka in Integrative Journal of Conference Proceedings
The vitamin E phosphate nucleoside prodrug (VEPNP) platform is designed to bypass two major mechanisms of tumor resistance to nucleosides: nucleoside transport and kinase downregulation. Testing showed that VEPNPs NUC050 and NUC024 were relatively unaffected by the presence of dipyridamole, an inhibitor of nucleoside transport, while the activity of gemcitabine decreased significantly. Further experiments found that NUC050 was able to deliver gemcitabine-monophosphate intracellularly in deoxycytidine kinase-deficient cells, bypassing this mechanism of resistance. A pilot study in mice found that NUC050 had a half-life 13.9 times longer
The document summarizes the synthesis of new triazenoazaindole compounds with potential antitumor activity. It introduces that new chemotherapeutic agents are still needed to treat cancer. It describes that a new class of compounds bearing a triazenoazaindole scaffold were synthesized with the aim of identifying new antiproliferative agents. The antiproliferative activity of the compounds was evaluated in five human tumor cell lines and results were expressed as GI50 values, with dacarbazine as a reference, showing the compounds have cytotoxic potential and warrant further investigation.
1) The study investigated the signaling pathways associated with the reduction in mammary cancer burden from dietary common bean consumption.
2) The study found that common bean consumption reduced levels of factors that promote cancer cell growth and survival like cyclin D1 and increased apoptotic factors like Bax/Bcl-2 ratio.
3) Bean consumption also modified key metabolic signaling networks linked to cell growth and survival, especially the mTOR pathway, which was reduced in a dose-dependent manner corresponding to bean consumption levels.
Molecular Modeling of Metalloreductase STEAP2 Protein and Docking Interaction...BRNSS Publication Hub
This gene is an individual from the STEAP family and encodes a multipass film protein that confines to the Golgi complex, the plasma layer, and the vesicular cylindrical structures in the cytosol. A very comparative protein in mouse has both ferrireductase and cupric reductase action and invigorates the cell take-up of both iron and copper in vitro. Expanded transcriptional articulation of the human quality is related with prostate malignant growth movement. Substitute transcriptional graft variations, encoding distinctive isoforms, have been described. Therefore, in the present study, we generated a precise three-dimensional (3D) model of metalloreductase STEAP2 protein using MODELLER 9.21 and validated its structure using PROCHECK software. Modeled protein contains more than 94.5% of amino acids in core region. We interpreted the action of natural compounds docking against the modeled metalloreductase STEAP2 protein. Three compounds (ginkgetin, medicagenin, and erybraedin A) showed lower binding affinity values toward metalloreductase STEAP2 protein compared to mitoxantrone, abiraterone acetate, apalutamide, enzalutamide, and flutamide. Ginkgetin exhibited the lowest binding energy of −9.10 kcal/mol with interacting Trp212 and Thr210. All the 17 compounds showed excellent binding energies than standard drugs for the modeled metalloreductase STEAP2 protein. These computational studies can be helpful to discover novel drug candidates.
CellRapeutics™ platform can support cellular therapy associated research by CAR cell in vitro assay services.
https://www.creative-biolabs.com/car-t/cellrapeutics-chimeric-antigen-receptor-car-technology.htm
This study investigated the association between the MMP-3 promoter polymorphism (-1171 5A->6A) and risk of oral submucous fibrosis (OSMF) and head and neck squamous cell carcinoma (HNSCC) in an Indian population. The authors genotyped 362 subjects, including 101 OSMF cases, 135 HNSCC cases, and 126 healthy controls. They found the 5A allele frequency was higher in OSMF (0.15) and HNSCC (0.13) compared to controls (0.07). Statistical analysis revealed a significant difference in 5A genotype frequency between OSMF and controls (p=0.01, OR=2.26) and between HNSCC
This document summarizes a study evaluating the toxicity and efficacy of novel targeted polymalic acid conjugate nanoparticles for treating triple-negative breast cancer. The nanoparticles were designed to target the epidermal growth factor receptor and laminin-411, a tumor vascular protein. In vitro and in vivo assays were conducted to evaluate the biocompatibility and toxicity of the polymalic acid platform and nanoparticle conjugates at low and high doses, as well as their efficacy in reducing tumor growth in mouse models over multiple treatment regimens. Blood analysis after intravenous administration in mice found no side effects, supporting the safety and potential of these nanoparticles for future cancer treatment in patients.
MTTlab is an expert testing service provider for the biomedical, biotechnological, pharmaceutical and food & beverage industries. They offer a wide range of in vitro and in vivo testing services to support drug development from early to mid-stage, including cytotoxicity testing, biomarker assays, preclinical drug evaluation, and toxicology studies in mice. MTTlab also assists with testing and certification requirements for food/beverage products. Their goal is to help clients fulfill their research and development needs through reliable, cost-effective testing services.
Molecular mechanisms of action and potential biomarkers of growth inhibition ...Enrique Moreno Gonzalez
Molecular targeted therapy has emerged as a promising treatment of Hepatocellular carcinoma (HCC). One potential target is the Src family Kinase (SFK). C-Src, a non-receptor tyrosine kinase is a critical link of multiple signal pathways that regulate proliferation, invasion, survival, metastasis, and angiogenesis. In this study, we evaluated the effects of a novel SFK inhibitor, dasatinib (BMS-354825), on SFK/FAK/p130CAS, PI3K/PTEN/Akt/mTOR, Ras/Raf/MAPK and Stats pathways in 9 HCC cell lines.
1. A study tested a combination of six phytochemicals (curcumin, resveratrol, genistein, quercetin, indole-3-carbinol, and C-phycocyanin) on breast cancer cells and mesenchymal stem cells.
2. When used individually, the phytochemicals were ineffective, but in combination they significantly suppressed breast cancer cell proliferation, inhibited migration and invasion, induced cell cycle arrest and apoptosis, resulting in 100% cell death, with no effects on mesenchymal stem cells.
3. Microarray analysis identified several differentially expressed genes involved in cell proliferation, survival, and metastasis that may underpin the combination's mode
1) Plumbagin (PL), derived from plants, inhibits prostate tumor development in mice lacking the tumor suppressor PTEN (Pten-KO mice), a model for prostate cancer.
2) PL treatment decreased expression of proteins involved in prostate cancer progression (PKCε, STAT3, AKT) and epithelial-to-mesenchymal transition (EMT).
3) Dietary PL inhibited primary prostate tumor growth and castration-resistant prostate cancer growth in Pten-KO mice, potentially by inhibiting PKCε, STAT3, AKT, and EMT markers.
This document evaluates the effects of tamoxifen and paclitaxel (taxol) in treating breast cancer. It begins with background on breast anatomy and cancer types. Tamoxifen is a selective estrogen receptor modulator that blocks estrogen's effects in breast tissue. Paclitaxel disrupts microtubule function, inhibiting cell division. The document reviews literature on paclitaxel's mechanism of action and structure. It will present results from a study on tamoxifen and paclitaxel's effects, and discuss methodology, interpretation and conclusions.
Current Perspective of Natural Alkaloid Carbazole and its Derivatives as Anti...Hanif Shaikh
This document summarizes research on natural and synthetic carbazole alkaloids and their potential as antitumor agents. Several studies are described that tested various carbazole derivatives for cytotoxicity against different cancer cell lines. Many of the carbazole compounds showed cytotoxicity in the low micromolar or nanomolar range against cell lines like MCF-7, A549, HCT-116, and others. The carbazoles are proposed to exert their antitumor effects through mechanisms like DNA intercalation or inhibition of DNA-dependent enzymes. The review concludes that natural carbazole alkaloids and their derivatives show promise as antitumor agents and warrant further investigation.
Propolis with its active component CAPE (Caffeic Acid Phenethyl Ester) stops breast cancer cell growth. These results of CAPE are present in the naturopathic formulation
of propolis, a widely available natural substance with an extended safety record, making it a naturally-occurring and readily available epigenetic agent with great potential in breast cancer and oncology in general. The ability to link the biological effects of a naturopathic remedy to the pharmacologic effects seen with an exciting class of drugs in the treatment of cancer opens the door to a host of new therapeutic opportunities for patients.
This document analyzes the kinetic and mechanical properties of interactions between MUC16 and PODXL (glycoproteins expressed on pancreatic cancer cells) and E- and L-selectins. Single molecule force spectroscopy was used to characterize the binding interactions and determine that MUC16- and PODXL-E-selectin interactions are mechanically stronger than the respective L-selectin interactions. Microfluidic assays were also used to study the interactions under flow, finding that the single molecule properties and contact time regulate rolling behavior on selectin surfaces under shear stress. Understanding the biophysics of these selectin-ligand interactions could aid in developing diagnostic tools and preventing cancer metastasis.
A New Platform for Combining the ‘Bottom-Up’ PBPK Paradigm and POPPK Data Ana...mjamei
The document discusses combining bottom-up physiologically-based pharmacokinetic (PBPK) modeling with top-down population pharmacokinetic (POPPK) data analysis. It describes current and previous Simcyp team members and grants received. It then contrasts top-down and bottom-up approaches, highlighting how the bottom-up approach can incorporate more physiological covariates compared to the empirical top-down approach. The document outlines Simcyp's parameter estimation module, which allows fitting simulations to clinical data to estimate population and drug parameters.
Activation of AMPK inhibits cervical cancer cell growth through AKT/FOXO3a/FO...Enrique Moreno Gonzalez
Although advanced-stage cervical cancer can benefit from current treatments, approximately 30% patients may fail after definitive treatment eventually. Therefore, exploring alternative molecular therapeutic approaches is imperatively needed for this disease. We have recently shown that activation of AMP-activated protein kinase (AMPK), a metabolic sensor, hampers cervical cancer cell growth through blocking the Wnt/β-catenin signaling activity. Here, we report that activated AMPK (p-AMPK) also inhibits cervical cancer cell growth by counteracting FOXM1 function.
Background and objectives: Iron is one of the major component
of hemoglobin, is required for transport of oxygen, ferritin is a
protein for storage iron, and transferring is a protein for iron
transport, all these it may be change in breast cancer.
The aim of present study was to measure the serum ferritin, iron,
total iron binding capacity, transferring, and C-reactive protein
concentration in breast tumors.
Material and method: A prospective study was carried out from
April 2013 to August 2014 by clinical biochemistry department in
College of Pharmacy-University of Sulaimani on (45) healthy
female individuals, (group 1) and (50) females with breast tumor
(group 2).
Results: The mean value of serum iron, transferring, total iron
binding capacity were significantly lower in females with breast
tumors (group 2), than that of healthy female individuals, (group
1), while serum ferritin was significantly higher in females with
breast tumors (group2), than that of healthy individuals (group
1).
Conclusion: Based on findings of the present study it can be
concluded that breast tumors can cause deficient of all iron
profile except ferritin will be increase in female breast cancers.
Key words- Serum iron, S.Ferritin, S. Total iron binding, S.
Transferrin, S.C-Reactive protein, breast tumors.
IPI-549 is a potent and selective inhibitor of PI3K-γ that has been shown to inhibit tumor growth in mouse models. IPI-549 blocks macrophage M2 polarization and migration in vitro. In mouse tumor models, IPI-549 treatment reduced tumor growth, decreased tumor-associated M2 macrophages and myeloid cells, and increased tumor-associated CD8+ T cells in a T cell-dependent manner. Combination of IPI-549 with checkpoint inhibitors showed enhanced anti-tumor effects over monotherapy in mouse models. These results suggest that targeting PI3K-γ with IPI-549 can alter the immune suppressive tumor microenvironment to support immune-mediated tumor growth inhibition.
This study evaluated the effects of Amgen 386, a novel angiopoietin inhibitor, alone and in combination with a c-MET inhibitor (Compound A) on metastasis and survival in a patient-derived xenograft model of clear cell renal cell carcinoma. Treatment with Amgen 386 alone and in combination with Compound A significantly reduced the number and size of lung metastases and prolonged survival compared to the control. The combination treatment was more effective at reducing metastases than either drug alone, suggesting potential clinical benefit from combination therapy for treating metastatic kidney cancer.
This document summarizes a study examining EphB4 as a potential therapeutic target in mesothelioma. The study found that EphB4 is overexpressed in 72% of human mesothelioma tissue samples, particularly in epithelioid subtypes. Treatment with an EphB4 inhibitor called sEphB4-HSA significantly inhibited tumor growth in mesothelioma xenograft models by inducing apoptosis and inhibiting PI3K and Src signaling. Combination treatment with sEphB4-HSA and an anti-VEGF antibody was more effective than either treatment alone and led to complete tumor regression in some cases. These results suggest that targeting EphB4, particularly with s
Nanoparticle (NP) Delivery of Chemotherapy
Drugs to Prostate Cancer Patients by Toluleke Oloruntobi Famuyiwa* and James Kwasi Kumi-Diaka in Integrative Journal of Conference Proceedings
The vitamin E phosphate nucleoside prodrug (VEPNP) platform is designed to bypass two major mechanisms of tumor resistance to nucleosides: nucleoside transport and kinase downregulation. Testing showed that VEPNPs NUC050 and NUC024 were relatively unaffected by the presence of dipyridamole, an inhibitor of nucleoside transport, while the activity of gemcitabine decreased significantly. Further experiments found that NUC050 was able to deliver gemcitabine-monophosphate intracellularly in deoxycytidine kinase-deficient cells, bypassing this mechanism of resistance. A pilot study in mice found that NUC050 had a half-life 13.9 times longer
The document summarizes the synthesis of new triazenoazaindole compounds with potential antitumor activity. It introduces that new chemotherapeutic agents are still needed to treat cancer. It describes that a new class of compounds bearing a triazenoazaindole scaffold were synthesized with the aim of identifying new antiproliferative agents. The antiproliferative activity of the compounds was evaluated in five human tumor cell lines and results were expressed as GI50 values, with dacarbazine as a reference, showing the compounds have cytotoxic potential and warrant further investigation.
1) The study investigated the signaling pathways associated with the reduction in mammary cancer burden from dietary common bean consumption.
2) The study found that common bean consumption reduced levels of factors that promote cancer cell growth and survival like cyclin D1 and increased apoptotic factors like Bax/Bcl-2 ratio.
3) Bean consumption also modified key metabolic signaling networks linked to cell growth and survival, especially the mTOR pathway, which was reduced in a dose-dependent manner corresponding to bean consumption levels.
Molecular Modeling of Metalloreductase STEAP2 Protein and Docking Interaction...BRNSS Publication Hub
This gene is an individual from the STEAP family and encodes a multipass film protein that confines to the Golgi complex, the plasma layer, and the vesicular cylindrical structures in the cytosol. A very comparative protein in mouse has both ferrireductase and cupric reductase action and invigorates the cell take-up of both iron and copper in vitro. Expanded transcriptional articulation of the human quality is related with prostate malignant growth movement. Substitute transcriptional graft variations, encoding distinctive isoforms, have been described. Therefore, in the present study, we generated a precise three-dimensional (3D) model of metalloreductase STEAP2 protein using MODELLER 9.21 and validated its structure using PROCHECK software. Modeled protein contains more than 94.5% of amino acids in core region. We interpreted the action of natural compounds docking against the modeled metalloreductase STEAP2 protein. Three compounds (ginkgetin, medicagenin, and erybraedin A) showed lower binding affinity values toward metalloreductase STEAP2 protein compared to mitoxantrone, abiraterone acetate, apalutamide, enzalutamide, and flutamide. Ginkgetin exhibited the lowest binding energy of −9.10 kcal/mol with interacting Trp212 and Thr210. All the 17 compounds showed excellent binding energies than standard drugs for the modeled metalloreductase STEAP2 protein. These computational studies can be helpful to discover novel drug candidates.
CellRapeutics™ platform can support cellular therapy associated research by CAR cell in vitro assay services.
https://www.creative-biolabs.com/car-t/cellrapeutics-chimeric-antigen-receptor-car-technology.htm
This study investigated the association between the MMP-3 promoter polymorphism (-1171 5A->6A) and risk of oral submucous fibrosis (OSMF) and head and neck squamous cell carcinoma (HNSCC) in an Indian population. The authors genotyped 362 subjects, including 101 OSMF cases, 135 HNSCC cases, and 126 healthy controls. They found the 5A allele frequency was higher in OSMF (0.15) and HNSCC (0.13) compared to controls (0.07). Statistical analysis revealed a significant difference in 5A genotype frequency between OSMF and controls (p=0.01, OR=2.26) and between HNSCC
This document summarizes a study evaluating the toxicity and efficacy of novel targeted polymalic acid conjugate nanoparticles for treating triple-negative breast cancer. The nanoparticles were designed to target the epidermal growth factor receptor and laminin-411, a tumor vascular protein. In vitro and in vivo assays were conducted to evaluate the biocompatibility and toxicity of the polymalic acid platform and nanoparticle conjugates at low and high doses, as well as their efficacy in reducing tumor growth in mouse models over multiple treatment regimens. Blood analysis after intravenous administration in mice found no side effects, supporting the safety and potential of these nanoparticles for future cancer treatment in patients.
MTTlab is an expert testing service provider for the biomedical, biotechnological, pharmaceutical and food & beverage industries. They offer a wide range of in vitro and in vivo testing services to support drug development from early to mid-stage, including cytotoxicity testing, biomarker assays, preclinical drug evaluation, and toxicology studies in mice. MTTlab also assists with testing and certification requirements for food/beverage products. Their goal is to help clients fulfill their research and development needs through reliable, cost-effective testing services.
Molecular mechanisms of action and potential biomarkers of growth inhibition ...Enrique Moreno Gonzalez
Molecular targeted therapy has emerged as a promising treatment of Hepatocellular carcinoma (HCC). One potential target is the Src family Kinase (SFK). C-Src, a non-receptor tyrosine kinase is a critical link of multiple signal pathways that regulate proliferation, invasion, survival, metastasis, and angiogenesis. In this study, we evaluated the effects of a novel SFK inhibitor, dasatinib (BMS-354825), on SFK/FAK/p130CAS, PI3K/PTEN/Akt/mTOR, Ras/Raf/MAPK and Stats pathways in 9 HCC cell lines.
1. A study tested a combination of six phytochemicals (curcumin, resveratrol, genistein, quercetin, indole-3-carbinol, and C-phycocyanin) on breast cancer cells and mesenchymal stem cells.
2. When used individually, the phytochemicals were ineffective, but in combination they significantly suppressed breast cancer cell proliferation, inhibited migration and invasion, induced cell cycle arrest and apoptosis, resulting in 100% cell death, with no effects on mesenchymal stem cells.
3. Microarray analysis identified several differentially expressed genes involved in cell proliferation, survival, and metastasis that may underpin the combination's mode
1) The study estimated the total phenol content and cytotoxic activity of ethanolic extracts of Momordica charantia against cervical and breast cancer cell lines.
2) Total phenol content decreased with increasing ethanol concentration from 50% to 100% in the extracts. The 50% ethanolic extract showed the highest cytotoxic activity.
3) In a time- and dose-dependent manner, the 50% ethanolic extract effectively inhibited the growth of cervical and breast cancer cell lines. The IC50 dose was lower for breast cancer cells, indicating it may be more potent against breast cancer.
Inhibition of glutathione by buthionine sulfoximine enhanced the anti-cancer ...Ashujit
Multiple myeloma (MM) is an incurable blood cancer. Melphalan is an alkylating agent given prior to stem cell transplantation to MM patients. Increased glutathione confers resistance to melphalan. This study investigate the effect of inhibition of glutathione by BSO in preclinical models of MM. Pretreatment with BSO enhanced the anti-cancer effect of melphalan in cell lines and animal models. BSO and melphalan combination was well tolerated by animals and enhanced the survival as compared to controls, BSO and melphalan alone. BSO enhanced depth and duration of responses induced by melphalan. In the combination group, majority of treated animals achieved complete response (CR) and more than 20% had maintained CR. Also, the survival of animals was doubled after combination treatment as compared to BSO or melphalan alone. Mechanistic investigation demonstrated that BSO enhanced melphalan induced DNA damage, caspase cleavage and apoptosis. The combination also achieved multi-logs of cells kills in nine human multiple myeloma cell lines and primary MM cells isolated from blood and bone marrows. Interestingly, the effect of BSO and melphalan combination was abolished when cells were treated with N-acetyl cysteine and sodium thiosulfate but not with vitamin C and vitamin E. This observation suggests that effect of BSO is primarily driven by its ability to deplete glutathione and therefore preventing melphalan detoxification. Together, this study provides framework for testing the combination in a Phase I trial.
Environment inside even a small tumor is characterized by total (anoxia) or partial oxygen deprivation, hypoxia. It has been shown that radiotherapy and some conventional chemotherapies may be less effective in hypoxia, and therefore it is important to investigate how different drugs act in different microenvironments. In this study we perform a large screening of the effects of 19 clinically used or experimental chemotherapeutic drugs on four different cell lines in conditions of normoxia, hypoxia and anoxia.
1) Prodigiosin, a bacterial metabolite, induces apoptosis in human breast cancer cells. Gene expression profiling found that prodigiosin strongly increased expression of the NAG-1 gene.
2) Experiments showed that prodigiosin triggers accumulation of the tumor suppressor protein p53, but induction of NAG-1 was independent of p53.
3) Prodigiosin causes inhibition of AKT and activation of glycogen synthase kinase-3B (GSK-3B). Induction of NAG-1 and apoptosis correlated with GSK-3B activation. Inhibiting GSK-3B reduced apoptosis, suggesting GSK-3B plays a key role in the proap
This document discusses the identification of TACC1, NOV, and PTTG1 as potential new biomarkers associated with endocrine therapy resistance in breast cancer. The study used two cell models, MVLN/CL6.7 and VP229/VP267, that were selected for resistance to tamoxifen and acquired cross-resistance to fulvestrant. 26 candidate genes were examined by RTQ-PCR and 8 genes were found overexpressed in breast cancer patient samples that relapsed after tamoxifen treatment. TACC1, NOV, and PTTG1 were identified as independent prognostic markers associated with shorter relapse-free survival. Aberrant mRNA and protein levels of these 3 genes were also observed in
This document discusses the identification of TACC1, NOV, and PTTG1 as potential new biomarkers associated with endocrine therapy resistance in breast cancer. Two cell models, MVLN/CL6.7 and VP229/VP267, were used that had acquired resistance to tamoxifen and cross-resistance to fulvestrant. 26 candidate genes related to cell proliferation, transformation, apoptosis and DNA repair were examined in these cell lines using RTQ-PCR. Genes with deregulated expression were further analyzed in samples from 48 ER-positive breast cancer patients, half of whom relapsed after tamoxifen treatment. TACC1, NOV, and PTTG1 were found to be over
This document discusses the identification of TACC1, NOV, and PTTG1 as potential new biomarkers associated with endocrine therapy resistance in breast cancer. Two cell models, MVLN/CL6.7 and VP229/VP267, were used that had acquired resistance to tamoxifen and cross-resistance to fulvestrant. 26 candidate genes related to cell proliferation, transformation, apoptosis and DNA repair were examined in these cell lines using RTQ-PCR. Genes with deregulated expression were further analyzed in samples from 48 ER-positive breast cancer patients, half of whom relapsed after tamoxifen treatment. Aberrant mRNA and protein levels of TACC1, NOV, and PTT
Recently, a phase II clinical trial in hepatocellular carcinoma (HCC) has suggested that the combination of sorafenib and 5-fluorouracil (5-FU) is feasible and side effects are manageable. However, preclinical experimental data explaining the interaction mechanism(s) are lacking. Our objective is to investigate the anticancer efficacy and mechanism of combined sorafenib and 5-FU therapy in vitro in HCC cell lines MHCC97H and SMMC-7721.
Cancer Cells and anticancer effect of some plants and materialsMaryam Yekefallah
The document discusses evaluation of anticancer agents and the anticancer effects of some materials and drugs. It provides an overview of the purpose and description of anticancer treatment, classifications of anticancer drugs, mechanisms of action, pre-clinical evaluation methods including in vitro and in vivo tests, and some modern treatments using plants and materials. Specifically, it summarizes the anticancer effects and molecular mechanisms of epigallocatechin-3-gallate, melatonin, and other compounds.
Odontogenic ameloblast-associated protein (ODAM) inhibits growth and migratio...Enrique Moreno Gonzalez
The Odontogenic Ameloblast-associated Protein (ODAM) is expressed in a wide range of
normal epithelial, and neoplastic tissues, and we have posited that ODAM serves as a novel
prognostic biomarker for breast cancer and melanoma. Transfection of ODAM into breast
cancer cells yields suppression of cellular growth, motility, and in vivo tumorigenicity.
Herein we have extended these studies to the effects of ODAM on cultured melanoma cell
lines.
1) Glyceollin I is a phytochemical that can reverse epithelial to mesenchymal transition (EMT) in letrozole-resistant breast cancer cells by inhibiting the zinc finger E-box binding homeobox 1 (ZEB1) transcription factor.
2) Treatment of letrozole-resistant breast cancer cells (LTLT-Ca cells) with glyceollin I led to decreased proliferation, invasion, and migration in vitro as well as weaker ZEB1 and N-cadherin staining and stronger E-cadherin staining in vivo.
3) Glyceollin I treatment of LTLT-Ca cells caused a 3.39-fold
This research paper examines the ability of the drug nelfinavir to overcome multidrug resistance in MCF-7/Dox breast cancer cells. The study finds that multiple exposures to physiologically achievable concentrations of nelfinavir can significantly decrease the doxorubicin IC50 in MCF-7/Dox cells by inhibiting P-glycoprotein expression and function, suppressing AKT signaling pathways, and inducing endoplasmic reticulum stress pathways. In mouse models carrying MCF-7/Dox tumor xenografts, combination treatment with nelfinavir and doxorubicin decreased tumor growth more than either drug alone. The results suggest that nelfinavir can overcome multiple drug resistance
Treatment of Ovarian Cancer First Line Chemotherapy or Targeted Therapy for R...ijtsrd
Ovarian cancer is the seventh most common gynecological cancer worldwide, ovarian cancer is the eighth leading cause of cancer death in women. In recent years, the number of ovarian cancer cases has been increasing in Japan, more than 9,000 women are diagnosed with ovarian cancer each year. The 5 year survival rate is 58 , the lowest among gynecological cancers, 4,758 ovarian cancer deaths in 2012. That is, it is reported that about one in two ovarian cancer patients has died. Because it is difficult to cure recurrent ovarian cancer, treatment is used to prolong life and improve quality of life. Because PARP inhibitors are oral targeted drugs that specifically act on cancer cells, they are expected to reduce the risk of disease progression and death while maintaining a good safety profile. In this way, the development of oral preparations has made it possible to avoid the burden on patients such as pain caused by conventional injections and the time constraints required for infusion. In this review, we discuss new treatments for ovarian cancer. Takuma Hayashi | Kaoru Abiko | Ken Yamaguchi | Junzo Hamanishi | Masaki Mandan | Ikuo Konishi "Treatment of Ovarian Cancer: First-Line Chemotherapy or Targeted Therapy for Recurrent Cases" Published in International Journal of Trend in Scientific Research and Development (ijtsrd), ISSN: 2456-6470, Volume-4 | Issue-3 , April 2020, URL: https://www.ijtsrd.com/papers/ijtsrd30470.pdf Paper Url :https://www.ijtsrd.com/medicine/other/30470/treatment-of-ovarian-cancer-firstline-chemotherapy-or-targeted-therapy-for-recurrent-cases/takuma-hayashi
EFFECT OF CITRUS LIMON JUICE AND TAMOXIFEN ON THE OXIDATIVE STRESS ACTIVITIES...oyepata
EFFECT OF CITRUS LIMON JUICE AND TAMOXIFEN ON THE
OXIDATIVE STRESS ACTIVITIES OF MCF-7 CELL INDUCED
BREAST CANCER IN SPRAWGUE DAWLEY RATS. JOSEPH OYEPATA SIMEON
Tumor Markers include a wide range of biomacromolecules orchestrated in abundance fixation by a wide assortment of neoplastic cells. The markers could be endogenous results of exceptionally dynamic metabolic threatening cells or the results of recently turned on qualities, which stayed unexpressed in early life or recently obtained antigens at cell and sub-cell levels. The presence of tumor marker and their focus are identified with the beginning and development of dangerous tumors in patients. A perfect tumor marker ought to be profoundly delicate, explicit, dependable with high prognostic worth, organ particularity and it should relate with tumor stages. Be that as it may, none of the tumor markers answered to date has every one of these attributes. Inspite of these impediments, numerous tumor markers have indicated incredible clinical significance in checking adequacy of various methods of treatments during whole course of sickness in malignant growth patients. Moreover, assurance of markers additionally helps in early discovery of malignant growth repeat and in anticipation.
A Review on Protein and Cancer ; Etiology, Metabolism and ManagementAbdulrahman Ragab
Altered metabolism is one of the hallmarks of cancer cells. Cell cycling and protein synthesis are both key
physiological tasks for cancer cells. In recent years, interest has been renewed as clear that many of the signaling
pathways that are affected by genetic mutations and the tumor microenvironment have a profound effect on core
metabolism of cancer cells. Metabolic alterations in cancer cells are numerous and include aerobic glycolysis,
reduced oxidative phosphorylation and the increased generation of biosynthetic intermediates needed for cell
growth and proliferation. Furthermore, accelerated protein turnover seen in many cancer patients and whole body
protein turnover is increased with advancing stage of disease. Cancer cells alter their consumption and the way
they process sugars, fats, amino acids and other energy sources to satisfy the demands of continuous proliferation.
The possible effects of specific amino acid, methionine, asparagine, arginine, tyrosine and glutamine, etc. on
protein cancer metabolism are discussed. Evidences confirm a contribution of proteins in all cancer stages and
describe metabolism of protein in cancer and how amino acids can be targeted to management or initially prevent
different types of cancer. Several studies suggest that people who eat more red meat have higher risk for
developing colorectal cancer than those who eat less red meat, but avoiding processed meats is even more
important for cancer prevention. In this review we summarize the role of proteins in cancer etiology, metabolism,
its complication, prevention and treatments.
Similar to Equol enhances tamoxifen's anti-tumor activity by induction of caspase-mediated apoptosis in MCF-7 breast cancer cells (18)
Incidence of pneumonia and risk factors among patients with head and neck can...Enrique Moreno Gonzalez
This study investigated the incidence and patient- and treatment-related risk factors related to pneumonia acquired during radiotherapy (PNRT) in head and neck cancer (HNC) patients.
Gene expression analysis of a Helicobacter pyloriinfected and high-salt diet-...Enrique Moreno Gonzalez
Helicobacter pylori (H. pylori) infection and excessive salt intake are known as important risk factors for stomach cancer in humans. However, interactions of these two factors with gene expression profiles during gastric carcinogenesis remain unclear. In the present study, we investigated the global gene expression associated with stomach carcinogenesis and prognosis of human gastric cancer using a mouse model.
Acute myeloid leukemia (AML) is a hematopoietic malignancy with a dismal outcome in the majority of cases. A detailed understanding of the genetic alterations and gene expression changes that contribute to its pathogenesis is important to improve prognostication, disease monitoring, and therapy. In this context, leukemia-associated misexpression of microRNAs (miRNAs) has been studied, but no coherent picture has emerged yet, thus warranting further investigations.
Differences in microRNA expression during tumor development in the transition...Enrique Moreno Gonzalez
The prostate is divided into three glandular zones, the peripheral zone (PZ), the transition zone (TZ), and the central zone. Most prostate tumors arise in the peripheral zone (70-75%) and in the transition zone (20-25%) while only 10% arise in the central zone. The aim of this study was to investigate if differences in miRNA expression could be a possible explanation for the difference in propensity of tumors in the zones of the prostate.
Multicentric and multifocal versus unifocal breast cancer: differences in the...Enrique Moreno Gonzalez
This study compared the expression of E-cadherin, β-catenin, and MUC1 in multicentric/multifocal breast cancers versus unifocal breast cancers of identical tumor size and grade. The study found significantly downregulated expression of E-cadherin in multicentric/multifocal cancers compared to unifocal cancers. In contrast, no significant differences were seen in β-catenin expression between the two groups. Within the unifocal group, E-cadherin and β-catenin expression were positively correlated, but this was not seen in the multicentric/multifocal group. The results suggest multicentric/multifocal and unifocal breast cancers differ in E-
The life in sight application study (LISA): design of a randomized controlled...Enrique Moreno Gonzalez
It is widely recognized that spiritual care plays an important role in physical and psychosocial well-being of cancer patients, but there is little evidence based research on the effects of spiritual care. We will conduct a randomized controlled trial on spiritual care using a brief structured interview scheme supported by an e-application. The aim is to examine whether an assisted reflection on life events and ultimate life goals can improve quality of life of cancer patients.
Clinical and experimental studies regarding the expression and diagnostic val...Enrique Moreno Gonzalez
Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) is a multifunctional Ig-like cell adhesion molecule that has a wide range of biological functions. According to previous reports, serum CEACAM1 is dysregulated in different malignant tumours and associated with tumour progression. However, the serum CEACAM1 expression in nonsmall-cell lung carcinomas (NSCLC) is unclear. The different expression ratio of CEACAM1-S and CEACAM1-L isoform has seldom been investigated in NSCLC. This research is intended to study the serum CEACAM1 and the ratio of CEACAM1-S/L isoforms in NSCLC.
Assessment of preoperative exercise capacity in hepatocellular carcinoma pati...Enrique Moreno Gonzalez
Cardiopulmonary exercise testing measures oxygen uptake at increasing levels of work and predicts cardiopulmonary performance under conditions of stress, such as after abdominal surgery. Dynamic assessment of preoperative exercise capacity may be a useful predictor of postoperative prognosis. This study examined the relationship between preoperative exercise capacity and event-free survival in hepatocellular carcinoma (HCC) patients with chronic liver injury who underwent hepatectomy.
Overexpression of YAP 1 contributes to progressive features and poor prognosi...Enrique Moreno Gonzalez
Yes-associated protein 1 (YAP 1), the nuclear effector of the Hippo pathway, is a key regulator of organ size and a candidate human oncogene in multiple tumors. However, the expression dynamics of YAP 1 in urothelial carcinoma of the bladder (UCB) and its clinical/prognostic significance are unclear.
CXCR7 is induced by hypoxia and mediates glioma cell migration towards SDF-1a...Enrique Moreno Gonzalez
Glioblastomas, the most common and malignant brain tumors of the central nervous system, exhibit high invasive capacity, which hinders effective therapy. Therefore, intense efforts aimed at improved therapeutics are ongoing to delineate the molecular mechanisms governing glioma cell migration and invasion.
Abnormal expression of Pygopus 2 correlates with a malignant phenotype in hum...Enrique Moreno Gonzalez
Pygopus 2 (Pygo2) is a Pygo family member and an important component of the Wnt signaling transcriptional complex. Despite this data, no clinical studies investigating Pygo2 expression in lung cancer have yet been reported.
Differentiation of irradiation and cetuximab induced skin reactions in patien...Enrique Moreno Gonzalez
In order to improve the clinical outcome of patients with locally advanced squamous cell carcinoma of the head and neck (LASCCHN) not being capable to receive platinum-based chemoradiation, radiotherapy can be intensified by addition of cetuximab, a monoclonal antibody that blocks the epidermal growth factor receptor (EGFR). The radioimmunotherapy with cetuximab is a feasible treatment option showing a favourable toxicity profile. The most frequent side effect of radiotherapy is radiation dermatitis, the most common side effect of treatment with cetuximab is acneiform rash. Incidence and severity of these frequent, often overlapping and sometimes limiting skin reactions, however, are not well explored. A clinical and molecular differentiation between radiogenic skin reactions and skin reactions caused by cetuximab which may correlate with outcome, have never been described before.
Cholestasis induces reversible accumulation of periplakin in mouse liverEnrique Moreno Gonzalez
Periplakin (PPL) is a rod-shaped cytolinker protein thought to connect cellular adhesion junctional complexes to cytoskeletal filaments. PPL serves as a structural component of the cornified envelope in the skin and interacts with various types of proteins in cultured cells; its level decreases dramatically during tumorigenic progression in human epithelial tissues. Despite these intriguing observations, the physiological roles of PPL, especially in noncutaneous tissues, are still largely unknown. Because we observed a marked fluctuation of PPL expression in mouse liver in association with the bile acid receptor farnesoid X receptor (FXR) and cholestasis, we sought to characterize the role of PPL in the liver and determine its contributions to the etiology and pathogenesis of cholestasis.
Functional p53 is required for rapid restoration of daunorubicin-induced lesi...Enrique Moreno Gonzalez
This document summarizes a research article that studied the role of p53 in daunorubicin (DNR)-induced lesions in the spleen. The key findings were:
1) DNR treatment caused more rapid cell death and weight loss in the spleens of wild type mice compared to p53-null mice.
2) While wild type mouse spleens recovered normal morphology 8 days after DNR treatment, p53-null mouse spleens still had large necrotic lesions.
3) DNR treatment increased p21 levels in wild type mice but not p53-null mice, indicating p53 is required for p21 induction.
4) The results suggest p53
Post-diagnosis hemoglobin change associates with overall survival of multiple...Enrique Moreno Gonzalez
Anemia refers to low hemoglobin (Hb) level and is a risk factor of cancer patient survival. The National Comprehensive Cancer Network recently suggested that post-diagnosis Hb change, regardless of baseline Hb level, indicates the potential presence of anemia. However, there is no epidemiological study evaluating whether Hb change has direct prognostic values for cancer patients at the population level.
Cost-effectiveness of MRI for breast cancer screening in BRCA1/2 mutation car...Enrique Moreno Gonzalez
Women with mutations in BRCA1 or BRCA2 are at high risk of developing breast cancer and, in British Columbia, Canada, are offered screening with both magnetic resonance imaging (MRI) and mammography to facilitate early detection. MRI is more sensitive than mammography but is more costly and produces more false positive results. The purpose of this study was to calculate the cost-effectiveness of MRI screening for breast cancer in BRCA1/2 mutation carriers in a Canadian setting.
Impaired mitochondrial beta-oxidation in patients with chronic hepatitis C: r...Enrique Moreno Gonzalez
Hepatic steatosis is often seen in patients with chronic hepatitis C (CH-C). It is still unclear whether these patients have an impaired mitochondrial β-oxidation. In this study we assessed mitochondrial β-oxidation in CH-C patients by investigating ketogenesis during fasting.
Intraepithelial lymphocyte distribution differs between the bulb and the seco...Enrique Moreno Gonzalez
Evaluation of intraepithelial duodenal lymphocytosis (IDL) is important in celiac disease (CD). There is no established cut-off value for increased number of IELs in the bulb. We therefore investigated the relation between IEL counts in the bulb and duodenal specimens in non-celiac subjects.
Sticky siRNAs targeting survivin and cyclin B1 exert an antitumoral effect on...Enrique Moreno Gonzalez
Melanoma represents one of the most aggressive and therapeutically challenging malignancies as it often gives rise to metastases and develops resistance to classical chemotherapeutic agents. Although diverse therapies have been generated, no major improvement of the patient prognosis has been noticed. One promising alternative to the conventional therapeutic approaches currently available is the inactivation of proteins essential for survival and/or progression of melanomas by means of RNA interference. Survivin and cyclin B1, both involved in cell survival and proliferation and frequently deregulated in human cancers, are good candidate target genes for siRNA mediated therapeutics.
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The biomechanics of running involves the study of the mechanical principles underlying running movements. It includes the analysis of the running gait cycle, which consists of the stance phase (foot contact to push-off) and the swing phase (foot lift-off to next contact). Key aspects include kinematics (joint angles and movements, stride length and frequency) and kinetics (forces involved in running, including ground reaction and muscle forces). Understanding these factors helps in improving running performance, optimizing technique, and preventing injuries.
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Travel Clinic Cardiff: Health Advice for International TravelersNX Healthcare
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Discuss and demonstrate the approaches with array and NGS genotyping methods for star allele calling to prep for downstream analysis.
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2. Equol enhances tamoxifen’s anti-tumor activity by
induction of caspase-mediated apoptosis in MCF-7
breast cancer cells
Christiana Charalambous1
Email: ccharala@ucy.ac.cy
Chara A Pitta1
Email: pittachara@gmail.com
Andreas I Constantinou1*
*
Corresponding author
Email: andreasc@ucy.ac.cy
1
Department of Biological Sciences, University of Cyprus, 75 Kallipoleos str, PO
box 20537, Lefkosia 1678, Cyprus
Abstract
Background
Soy phytoestrogens, such as daidzein and its metabolite equol, have been proposed to be
responsible for the low breast cancer rate in Asian women. Since the majority of estrogen
receptor positive breast cancer patients are treated with tamoxifen, the basic objective of this
study is to determine whether equol enhances tamoxifen’s anti-tumor effect, and to identify
the molecular mechanisms involved.
Methods
For this purpose, we examined the individual and combined effects of equol and tamoxifen
on the estrogen-dependent MCF-7 breast cancer cells using viability assays, annexin-V/PI
staining, cell cycle and western blot analysis.
Results
We found that equol (>50 µM) and 4-hydroxy-tamoxifen (4-OHT; >100 nΜ) significantly
reduced the MCF-7 cell viability. Furthermore, the combination of equol (100 µΜ) and 4-
OHT (10 µΜ) induced apoptosis more effectively than each compound alone. Subsequent
treatment of MCF-7 cells with the pan-caspase inhibitor Z-VAD-FMK inhibited equol- and
4-OHT-mediated apoptosis, which was accompanied by PARP and α-fodrin cleavage,
indicating that apoptosis is mainly caspase-mediated. These compounds also induced a
marked reduction in the bcl-2:bax ratio, which was accompanied by caspase-9 and caspase-7
activation and cytochrome-c release to the cytosol. Taken together, these data support the
notion that the combination of equol and tamoxifen activates the intrinsic apoptotic pathway
more efficiently than each compound alone.
3. Conclusions
Consequently, equol may be used therapeutically in combination treatments and clinical
studies to enhance tamoxifen’s effect by providing additional protection against estrogen-
responsive breast cancers.
Keywords
Apoptosis, Breast cancer, Caspases, Equol, Tamoxifen
Background
Evidence from epidemiological studies suggest that nutrition plays an important role in the
development of breast cancer, which remains the most common malignancy and the second
most lethal cancer in women worldwide [1-4]. It was observed that the incidence of breast
cancer is much lower in Asian women compared to Western women, and this was attributed
to the daily consumption of soy products by Asian women, which contain phytoestrogens [5].
Equol (7-hydroxy-3-(4′-hydroxyphenyl)-chroman) is the bioactive metabolite of daidzein, a
major phytoestrogen found in soy products. Recent studies suggest that equol has the greatest
in vitro bioactivity and anti-oxidant activity when compared to soy isoflavones [6-8]. As
known, 30-50% of the adult population cannot metabolize daidzein to equol and,
interestingly, clinical response is usually limited to people who are “equol producers” [9,10].
Equol is reported to bind to both estrogen receptors ERα and ERβ, with a higher binding
affinity for ERβ, which has been implicated in the inhibition of proliferation and induction of
apoptosis in breast cancer cells [8,11-13]. Previous studies suggest that equol induces
apoptosis in the ER negative breast cancer cells [14,15], while it seems to have a biphasic
effect in ER-positive breast cancer cells enhancing cell proliferation at low concentrations (<
10 µΜ) [15-18] and possibly exerting an inhibitory effect at high concentrations (50–100
µΜ) [14]. As the role of equol in relation to breast cancer remains unclear, this study was
designed to delineate the effect of equol on estrogen-dependent breast cancer cells using
MCF-7 cells as a model system. This is particularly important as the controversy of results
obtained in the soy isoflavone human intervention studies and the inability to establish the
beneficial effects of soy isoflavones could be attributed to the failure to distinguish between
“equol producers” and “non-equol producers” [10,19]. Therefore, the significance of
evaluating the therapeutic potential of equol becomes more evident and may facilitate the
design and implementation of future equol intervention studies for cancer.
Several reports suggest that equol and daidzein induce cell cycle arrest and apoptosis in
breast cancer cells [2,8,14,20-25]. More specifically, it has been recently shown that daidzein
induces MCF-7 breast cancer cell apoptosis via the intrinsic (mitochondrial) caspase-
dependent apoptotic pathway [2]. However, the biological effects of equol have not been
investigated as well as those of daidzein. Therefore, the aim of this study is to thoroughly
explore the mechanism of equol-mediated apoptosis.
Tamoxifen, on the other hand, is an ΕRα antagonist classified as a non-steroidal selective
estrogen receptor modulator (SERM), widely used in cancer chemoprevention and
chemotherapy to prevent primary breast tumors or the development of recurrences,
respectively [26-28]. Tamoxifen, and its bioactive metabolite 4-hydroxy-tamoxifen (4-OHT),
4. inhibit proliferation and induce apoptosis in several types of ER-positive and ER-negative
breast cancer cells, rat mammary tumors and other cancer types [29-34]. However, the anti-
tumor mechanism of tamoxifen is not yet completely understood.
Accumulating experimental evidence from in vivo studies is beginning to support the
possibility that soy components may enhance tamoxifen’s anti-tumor effect, by providing
stronger protection against mammary carcinogenesis than tamoxifen alone [35,36].
Moreover, we have previously identified daidzein as the soy ingredient enhancing
tamoxifen’s ability to prevent rat mammary tumor formation [37]. Since equol is the
bioactive metabolite of daidzein [38,39], these findings support the premise that equol may
potentiate tamoxifen’s efficacy against mammary carcinogenesis. We are reporting here the
mechanism by which this daidzein metabolite enhances tamoxifen’s anti-tumor activity in ER
positive breast cancer cells.
Methods
Cell culture
MCF-7 breast cancer cell line (obtained from ATCC) was cultured in MEM supplemented
with 10% fetal bovine serum (FBS), 1% antibiotic-antimycotic, 1 mΜ sodium pyruvate, 1%
non-essential aminoacids (MEM-NEAA), 2 mM L-glutamine (Gibco, Life Technologies,
Paisley, UK) and 0.06 µg/ml insulin (Sigma, St. Louis, MI, USA). They were incubated at 37
°C in a humidified incubator with 5% CO2. For estrogen deprivation, three days before
treatment with equol or tamoxifen, cells were cultured in phenol-red free MEM supplemented
with 10% dextran-coated charcoal (DCC) - treated FBS, 1% antibiotic-antimycotic, 1% non-
essential aminoacids, 2 mM L-glutamine, 1 mM sodium pyruvate and 0.06 µg/ml insulin
[40].
Antibodies and reagents
Equol and 4-OHT were purchased from LC laboratories (Woburn, MA, USA) and Alexis
Biochemicals (Enzo Life Sciences, Lausen, Switzerland), respectively. Reagents also
included the pan-caspase inhibitor Z-VAD-FMK (Calbiochem, Nottingham, UK) and the
MTT reagent (Sigma, St. Louis, MI, USA). The bcl-2, bax, glyceraldehyde 3-phosphate
dehydrogenase (GAPDH) and cyclo-oxygenase-4 (COX-4) antibodies were purchased from
Santa Cruz Biotechnology (Heidelberg, Germany) whereas the poly-(ADP ribose)-
polymerase-1 (PARP-1), α-fodrin, caspase-9, caspase-8, caspase-7, caspase-6, cytochrome-c,
and α-tubulin antibodies were purchased from Cell Signaling Technology (Danvers, MA,
USA).
MTT assay
The effect of equol, 4-OHT and their combination on MCF-7 viability was examined using
the MTT (monotetrazolium) assay [41]. The cells were plated in 96-well plates (3×103
cells/well) and treated with different concentrations of equol and 4-OHT for 24, 48 and 72 h.
The MTT reagent was subsequently added (1:10 dilution) for 4 h at 37°C. The media were
then removed and DMSO (150 µL/well) was added for 20 min. The absorbance, measured at
570 nm, was proportional to the number of viable cells per well.
5. Mitochondrial/cytosolic extract preparation
Cells were cultured in 150-mm Petri dishes and treated for 48 h with vehicle control (DMSO
and ethanol), equol (100 µΜ), 4-OHT (10 µΜ) or their combination. Mitochondrial and
cytosolic extracts were prepared using the mitochondrial/cytosol fractionation kit (Abcam,
UK).
Western blot analysis
MCF-7 cells were treated with equol (100 µM), 4-OHT (10 µΜ) and their combination for 48
h, with or without Z-VAD-FMK (20 µΜ), and whole cell or mitochondrial/cytosolic extracts
were prepared as previously described [42]. Protein content in the extracts was quantified
using a bicinchoninic acid (BCA) protein assay kit (Pierce, Germany). Equal amount of
proteins (40 µg/lane) were separated on SDS-PAGE and electrotransferred to 0.45 µm
nitrocellulose membranes. The membranes were then blocked with 5% non-fat dry milk in
TBST (Tris buffered saline supplemented with 0.1% Tween-20) and probed with antibodies
against PARP-1 (1:1000 dilution), α-fodrin (1:500 dilution), caspase-9 (1:500 dilution),
caspase-8 (1:500 dilution), caspase-7 (1:500 dilution), caspase-6 (1:500 dilution), GAPDH;
1:1000 dilution), bcl-2 (1:500 dilution), bax (1:500 dilution), COX-4, (1:250 dilution),
cytochrome-c (1:250 dilution), and α-tubulin (1:1000 dilution) followed by HRP-conjugated
anti-rabbit or anti-mouse immunoglobulin-G (IgG; 1:2000 dilution). Protein bands were
detected by chemiluminescence using the Luminol substrate (Santa Cruz) according to the
manufacturer’s protocol and analyzed using the UVP bioimaging system (Cambridge, UK).
Cell death ELISA (Enzyme-linked immunosorbent assay)
MCF-7 cells were plated in 96-well plates in quadruplicates at a concentration of 3 × 104
cells/ml (100 µl/well). Cells were treated with equol (100 µΜ), 4-ΟΗΤ (10 µΜ) and their
combination and lysed after 72 h. Lysates were analyzed for the presence of nucleosomes
using the Cell Death Detection ELISA Plus kit (Roche Diagnostics, Mannheim, Germany).
Absorbance, measured at 405 nm, was proportional to cell death.
Tali™
apoptosis kit
Cells were plated in 60-mm plates and treated with equol (100 µΜ), 4-OHT (10 µΜ) and
their combination, with or without Z-VAD-FMK (20 µΜ). Cells were harvested 72 h post-
treatment and stained using annexin-V Alexa Fluor® 488/PI (propidium iodide), as described
by the Tali™
apoptosis kit (Life Technologies). Cell viability, death and apoptosis were
evaluated using the Tali™
Image-based Cytometer (Life Technologies). The annexin-V
positive/PI negative cells were recognized as apoptotic cells by the cytometer software
whereas the annexin V positive/PI positive cells were identified as dead cells. Similarly, the
annexin V-negative/PI negative cells were identified as viable cells.
Cell cycle analysis
Cells were plated in 100-mm plates and treated with equol (100 µΜ), 4-OHT (10 µΜ), and
their combination for 6, 12, 24, 48 and 72 h. They were harvested, fixed in 70% ethanol,
incubated with the PI staining solution (containing 1 mg/ml PI and 100 µg/ml RNase) for 15
6. min at 37 °C and analyzed for DNA content using the Guava EasyCyte™
flow cytometer and
the GuavaSoft analysis software (Millipore, Watford,UK).
Statistical analysis
Values are presented as the mean ± SEM. Statistical significance was evaluated using
student’s t-test for paired comparison. P<0.05 was considered statistically significant. Data
are representative of three individual experiments. Each experimental group was repeated in
triplicates or quadruplicates, as described in the Figure Legends section.
Results
Equol and 4-OHT reduce MCF-7 viability
To examine the ability of equol and 4-OHT to inhibit MCF-7 cell growth, their individual and
combined effects on cell viability were observed. Equol (> 50 µΜ) and 4-OHT (>100 nM)
provoked a marked reduction in MCF-7 viability in a dose- and time-dependent manner
(Figure 1A-C). In contrast, lower concentrations of equol (1 nΜ- 1 µΜ) did not exert a
significant effect on cell growth (data not shown). Futhermore, the combination of equol (100
µΜ) and 4-OHT (10 µΜ) reduced cell viability in an additive manner (72 h; Figure 1A),
suggesting that equol enhances tamoxifen’s anti-proliferative effect in MCF-7 cells.
Figure 1 Comparison of the effect of equol and 4-OHT on MCF-7 cell viability. (A) Cells
(3x103
/well) were plated in 96-well plates and treated with different concentrations of equol
and 4-OHT, individually or combined. After 24, 48 and 72 h, cell viability was evaluated
using the MTT assay. The OD reading at 570 nm was proportional to cell viability. * P <
0.05, ** P < 0.005 and *** P <0.0005. PEquol (100 µΜ) vs control= 0.003; P4-OHT(10 µΜ) vs control =
0.002; P[Equol (100 µΜ)+4-ΟΗΤ(10 µΜ)] vs control = 0.0003. Dose response curves for equol (B) or 4-
OHT (C) effect on MCF-7 cell viability. Cells (3 × 103
/well) were seeded in 96-well plates
and treated with different concentrations of equol (B) or 4-OHT (C). After 72 h, cell viability
was evaluated using the MTT assay. The data are expressed as percentage change in viability
in comparison to the vehicle treated control group. Each experimental group was repeated in
quadruplicates and data are representative of three individual experiments. Bars correspond to
the standard error of mean (SEM).
Equol and 4-OHT induce MCF-7 cell death via apoptosis
We began evaluating the mechanism implicated in the reduction of MCF-7 cell viability by
determining cell death following treatment with equol and 4-OHT. These compounds induced
MCF-7 death after 72 h of treatment (Figure 2A). Interestingly, their combination enhanced
cell death in an additive manner (P [Equol+4-OHT] vs. 4-OHT = 0.028; P [Equol+4-OHT] vs. Equol =0.023).
Figure 2 Effect of equol and 4-OHT on cell death (A), apoptosis (B) and cell cycle
distribution (C). For the determination of cell death (A), MCF-7 cells were seeded in 96-
well plates (3 × 103
cells/well). Upon attachment cells were treated with equol (100 µΜ)
and/or 4-OHT (100 µΜ). After 72 hours, cell death was evaluated using the Cell Death
ELISA. The OD reading at 405 nm was proportional to the number of nucleosomes released
in the cell lysates of the cells. The data are expressed as OD (405 nm) in comparison to the
vehicle- treated control group. Each group was repeated in quadruplicates.* PEquol vs control =
7. 0.023; ** P4-OHT vs control = 0.032; *** P[Equol+4-OHT] vs control = 0.016. (B) Effect of equol and 4-
OHT on MCF-7 cell apoptosis using annexin-V Alexa Fluor® 488/PI staining. Cells were
plated in 60-mm plates and treated with equol (100 µΜ) and 4-OHT (10 µΜ) for 72 h. Cell
viability, death and apoptosis were evaluated using the TaliTM
apoptosis kit and the TaliTM
Image-based Cytometer. Each experimental group was repeated in triplicate. Bars correspond
to the standard error of mean (SEM). * PEquol vs control =0.032; ** P4-OHT vs control =0.011; ***
P[Equol + 4-OHT] vs control = 0.013. (C) Effect of equol and 4-OHT on cell cycle distribution using
PI staining. MCF-7 cells were treated with equol (100 µΜ) and 4-OHT (10 µΜ) for 72 h. Cell
cycle distribution was evaluated using PI staining for 15 min at 37 °C. Sample analysis was
performed using the Guava EasyCyteTM
flow cytometer and the GuavaSoft analysis software.
Each experimental group was repeated in triplicate. Bars correspond to the standard error of
mean (SEM). * PEquol vs control = 0.0025; ** P4-OHT vs control = 0.026; *** P[Equol+4-OHT] vs control =
0.0037.
To examine whether cell death was mediated through apoptosis, cells were stained with
annexin-V/PI following treatment with equol and 4-OHT. Each compound produced a
substantial increase in the percentage of apoptotic cells (Figure 2B). The combination of
equol and 4-OHT had an additive effect on cell apoptosis (P [Equol+4-OHT] vs. 4-OHT=0.028; P
[Equol+4-OHT] vs. Equol = 0.018).
The effects of equol and tamoxifen on cell cycle progression were also determined using flow
cytometry. Even though no substantial changes were evident in cell cycle distribution from
6–48 h of treatment (data not shown), significant increase in the sub-G1 phase, which is
indicative of apoptosis, was observed at 72 h, accompanied by a marked reduction in the
percentage of cells in the G0/G1, S and G2/M phases (Figure 2C). These results show that
68.9±3.6% of the cells treated with the equol/4-OHT combination were in the sub-G1 phase,
which is significantly higher than the corresponding percentage of equol-treated cells
(32.1±0.5%), 4-OHT-treated cells (52.1±4.2%) or vehicle control treated cells (7.8±1.1%) (P
= 0.0037; Figure 2C). Taken together, these results indicate that these agents do not induce
cell cycle arrest, and that their combination is more effective in activating apoptosis than each
compound alone. This is consistent with our previous data, demonstrating that equol and 4-
OHT do not increase p53 and p21 expression, which is up-regulated in cells undergoing G1
arrest (data not shown).
Z-VAD-FMK inhibits equol and 4-OHT mediated apoptosis
To elucidate the precise pathways involved in equol- and 4-OHT-induced apoptosis, cells
were treated with the pan-caspase inhibitor Z-VAD-FMK in combination with equol and/or
4-OHT and apoptosis was evaluated using annexin-V/PI staining. Z-VAD-FMK significantly
inhibited equol- and 4-OHT-induced apoptosis, indicating activation of the caspase-
dependent pathway by these compounds (Figure 3). However, the inhibition was not
complete, suggesting that caspase-independent mechanisms may be implicated in addition to
the caspase dependent mechanisms.
Figure 3 Effect of the pan-caspase inhibitor Z-VAD-FMK on equol and 4-OHT induced
MCF-7 cell apoptosis. Cells were plated in 60-mm plates and treated with equol (100 µΜ)
and 4-OHT (10 µΜ) for 72 h. Cell apoptosis was evaluated using annexin-V Alexa Fluor®
488/PI staining and the Tali™
Image-based Cytometer. Each experimental group was repeated
in triplicates and data are representative of three individual experiments. The bars correspond
8. to the SEM. * P Equol vs [Z-VAD-FMK+Equol] = 0.014; ** P4-OHT vs [ZVAD-FMK+4-OHT] =0.012; ***
P[Equol+4OHT] vs [Z-VAD-FMK+Equol+4-OHT] =0.017.
Equol and 4-OHT induce PARP and α-fodrin proteolysis
The apoptotic mechanisms involved in the death response to equol and 4-OHT were further
characterized by monitoring PARP and α-fodrin expression using western blotting. PARP
and α-fodrin are known substrates cleaved by the effector caspases-3 and −7, which are
activated in apoptotic cells [43,44]. PARP and α-fodrin proteolysis was evident with equol or
4-OHT treatment and was significantly enhanced by their combination (Figure 4A). This
effect was prevented to a large extent by Z-VAD-FMK (Figure 4A), reconfirming that equol-
and 4-OHT activate caspase-mediated apoptosis.
Figure 4 Effect of equol and 4-OHT on pro-apoptotic protein (A) and caspase (B)
expression. MCF-7 were treated for 48 h with equol (100 µΜ) and 4-OHT (10 µΜ) with and
without Z-VAD-FMK (20 µΜ) (A), or without Z-VAD-FMK (B), and whole cell extracts
were prepared. Protein expression was then analyzed by western blot. Data are representative
of three individual experiments. C, vehicle control; E, Equol (100 µΜ); T, 4-OHT (10 µΜ), E
+ T, Equol (100 µΜ) + 4-OHT (10 µΜ).
Equol and 4-OHT induce apoptosis via the intrinsic pathway
Based on our previous results suggesting activation of caspase-dependent apoptosis by equol
and 4-OHT, we examined their effect on caspase expression and activation. To distinguish
between the intrinsic and the extrinsic apoptotic pathways, we investigated the effect of equol
and 4-OHT on the initiator caspases −8 and −9 and the effector caspases −6 and −7. Equol
and 4-OHT induced a pronounced pro-caspase-7 and pro-caspase-9 cleavage and activation,
which was greatly enhanced by their combination (Figure 4B). In contrast, caspase-8 and
caspase-6 remained unaffected by these treatments (Figure 4B), indicating that these
compounds act mainly through the intrinsic apoptotic pathway.
The combination of equol and 4-OHT promotes cytochrome-c release and
reduction of bcl-2 expression
The key event causing caspase-9 cleavage, and thus activation of the intrinsic apoptotic
pathway, is cytochrome-c release from the mitochondria to the cytosol [45]. Therefore, we
explored the effect of equol and tamoxifen on cytochrome-c expression and localization. The
combination of equol and 4-OHT induced a substantial cytochrome-c release from the
mitochondria to the cytosol of MCF-7 cells (Figure 5) which was not detected in cells treated
with equol or 4-OHT alone, thus confirming the activation of the intrinsic apoptotic pathway.
Figure 5 Effect of equol and 4-OHT on cytochrome-c expression. MCF-7 were treated for
48 h with equol (100 µΜ) and 4-OHT (10 µΜ) and mitochondrial and cytosolic extracts were
prepared. Protein expression was then analyzed by western blot. Data are representative of
three individual experiments. C, vehicle control; E, Equol (100 µΜ); T, 4-OHT (10 µΜ), E +
T, Equol (100 µΜ) + 4-OHT (10 µΜ).
To complete the picture, we investigated the effect of the two compounds and their
combination on the expression of the anti-apoptotic protein bcl-2 and the pro-apoptotic
9. protein bax [46]. Bcl-2 and bax are proteins that can prevent or facilitate cytochrome-c
release from the mitochondria respectively, thus inhibiting or promoting apoptosis [47]. The
bcl-2:bax ratio is important in determining whether a cell will undergo apoptosis or survive
[47]. We found that equol and 4-OHT induced a time-dependent reduction in the total levels
of bcl-2 in MCF-7 cells, whereas they did not affect bax expression (Figure 6). The
combination of equol and tamoxifen had an additive effect in the reduction of bcl-2
expression, which was more evident at 72 h (Figure 6). Equol and 4-OHT did not affect bcl-2
or bax expression at 24 h of treatment (data not shown). Therefore, equol and 4-OHT induce
a time-dependent reduction of the bcl-2:bax ratio, promoting in this way cytochrome-c
release and activation of the intrinsic apoptotic pathway.
Figure 6 Effect of equol and 4-OHT on bcl-2 and bax expression. MCF-7 were treated for
48 and 72 h with equol (100 µΜ) and 4-OHT (10 µΜ) and whole cell extracts were prepared.
Protein expression was then analyzed by western blot using anti-bcl-2 and anti-bax
polyclonal antibodies. Data are representative of three individual experiments. C, vehicle
control; E, Equol (100 µΜ); T, 4-OHT (10 µΜ), E + T, Equol (100 µΜ) + 4-OHT (10 µΜ).
Discussion
In this study, we evaluated the individual and combined effects of equol and 4-OHT, the
bioactive metabolite of tamoxifen, in the ER positive MCF-7 breast cancer cells. Our findings
show for the first time that equol not only does not abolish the anti-tumor effects of
tamoxifen, but instead it induces apoptosis and significantly enhances tamoxifen’s pro-
apoptotic effects in these cells (Figure 1A-C and Figure 2A-C). Moreover, the pan-caspase
inhibitor Z-VAD-FMK significantly inhibited equol- and tamoxifen- induced apoptosis
(Figure 3), suggesting that these compounds activate the caspase-mediated apoptotic
pathway. However, the inhibition was not complete, suggesting that caspase-independent
mechanisms may also be involved in equol and tamoxifen induced apoptosis. Previous
studies support our findings showing that equol inhibits MCF-7 proliferation and induces
caspase-mediated apoptosis in ER negative breast cancer cells and rat mammary tumors
[8,48,49]. With respect to tamoxifen, previous studies provide evidence that tamoxifen
induces caspase-dependent apoptosis in MCF-7 and other types of cancer cells [30,32,50-53].
Even though high concentrations of equol (100 µΜ) were required to activate MCF-7
apoptosis, which are not physiologically achievable in human plasma due to metabolic
conversion of the active aglycone equol to the inactive conjugated form [54], our results may
find applications in targeted immunotherapies, which may enable maximal delivery of equol
into the cancer cells. This strategy was previously used successfully for genistein, which was
immunoconjugated with a monoclonal antibody and targeted to a B cell-specific receptor for
treatment of an animal model of B-cell precursor leukemia [55].
To fully explore the apoptotic pathway activated by equol and tamoxifen, we investigated
their effects on key proteins involved in apoptosis, such PARP, α-fodrin and caspases −6, -7,
-8 and −9. Caspase-9 is part of the intrinsic (mitochondrial) apoptotic pathway and is
activated by cytochrome-c release from the mitochondria, whereas caspase-8 is part of the
extrinsic apoptotic pathway activated by external signals through the death receptors [45].
Active caspase −9 and caspase-8 in turn induce cleavage and activation of the effector
caspases −3, -6 and −7 [43,45,56,57], which subsequently cleave nuclear and cytosolic
targets, such as PARP and α-fodrin, resulting in cell destruction [43,44]. Since MCF-7 cells
are deficient of functional caspase-3, the effector caspase-7 is responsible for apoptosis in
10. these cells [58-60]. Our experiments show that equol and 4-OHT induce PARP and α-fodrin
proteolysis, which was significantly enhanced by their combination and partially inhibited by
the pan-caspase inhibitor Z-VAD-FMK (Figure 4A), suggesting that additional proteases
besides caspases may be involved in equol- and tamoxifen-induced apoptosis. Furthermore,
the combination of equol and tamoxifen induced a pronounced caspase-9 and caspase-7
cleavage accompanied with cytochrome-c release into the cytosol, without affecting
caspases-6 and −8 (Figure 4B and Figure 5). Treatment with either equol or tamoxifen, on the
other hand, had a lesser effect on caspase-9 and caspase-7 cleavage associated with a trivial
effect on cytochrome-c release from the mitochondria into the cytosol. Consequently, the
combination of equol and tamoxifen is significantly more potent in inducing MCF-7 cell
apoptosis than each compound alone. Therefore, our data suggest that equol and tamoxifen
activate the intrinsic apoptotic pathway. Previous studies support our findings as they have
shown activation of the intrinsic apoptotic pathway in MCF-7 cells by tamoxifen and
daidzein [2,29-31,51,61,62]. Moreover, equol and tamoxifen induced a time-dependent
reduction in blc-2 expression and hence the bcl-2:bax ratio, which was further reduced by the
combination of the two compounds (Figure 6). Decreased bcl-2 expression was observed in
several cancer cell types treated with tamoxifen and daidzein [14,63,64] and in equol-induced
apoptosis in mammary carcinomas [14,48].
Conclusions
In conclusion, this study suggests that equol induces MCF-7 cell apoptosis and enhances
tamoxifen’s pro-apoptotic effect via activation of the intrinsic apoptotic pathway. The
significance of our findings is that women with ER-positive early-stage breast cancer,
undergoing tamoxifen adjuvant treatment, may be further benefitted by co-treatment with
pharmacological doses of equol. Our results also suggest that “equol producers” may be at
lower risk of developing breast cancer due to the apoptotic action of equol against ER
positive breast cancer cells. Future clinical trials designed to determine the safety and
efficacy of equol in adjuvant hormonal therapy against breast cancer are warranted.
Abbreviations
BCA, Bicinchronic acid; COX-4, Cyclo-oxygenase-4; DMBA, 6,12 -
imethylbenz[a]anthracene; ELISA, Enzyme-linked immunosorbent assay; ER, Estrogen
receptor; FBS, Fetal bovine serum; GAPDH, Glyceraldehyde 3-phosphate dehydrogenase;
IgG, Immunoglobulin G; MTT, Monotetrazolium; 4-OHT, 4-hydroxy-tamoxifen; PARP,
Poly (ADP ribose) polymerase; PI, Propidium iodide; SERM, Selective estrogen receptor
modulator.
Competing interests
The authors declare that they have no competing interests.
Authors’ contributions
CC carried out all the experiments included in this manuscript and participated in the design,
data acquisition, analysis and interpretation. CAP provided assistance with some of the
experiments and valuable feedback. AIC participated in the experimental design, data
11. analysis and interpretation. Both CC and AIC participated in drafting and critically revising
the manuscript. All authors read and approved the final manuscript.
Acknowledgements
The authors wish to thank Dr Paul Costeas, Dr Laoura Koumas and Dr Carsten Lederer for
their help with flow cytometric analysis. This work was supported by the Cyprus Research
Promotion Foundation grant YGEIA/TROFH/0308(BE).
Study design
The role of equol, tamoxifen and their combination in breast cancer treatment.
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20. A
PARP (116 kDa)
cleaved PARP (89 kDa)
α-fodrin (240 kDa)
cleaved fodrin (150
kDa)
GAPDH
Vehicle control Z-VAD-FMK (20 µΜ)
C E T E+T C E T E+T
pro-caspase-8
cleaved caspase-8
GAPDH
pro-caspase-9
cleaved caspase-9
pro-caspase-7
cleaved caspase-7
pro-caspase-6
cleaved caspase-6
C E T E+T
B
Figure 4
21. C E T E+T
cytochrome-c
COX-4
C E T E+T
Mitochondrial
extract
Cytosolic
extract
cytochrome-c
α-tubulinFigure 5
22. 48h 72h
C E T E+T C E T E+T
GAPDH
bcl-2
bax
Figure 6