transforming-growth-factor-beta and Triple-Negative-Breast-Neoplasms

The most effective way to control severity and mortality rate of the novel coronavirus disease (COVID-19) is through sensitive diagnostic approaches and an appropriate treatment protocol. We aimed to identify the effect of adding corticosteroid and Tocilizumab to a standard treatment protocol in treating COVID-19 patients with chronic disease through hematological and lab biomarkers.. This study was performed retrospectively on 68 COVID-19 patients with chronic disease who were treated by different therapeutic protocols. The patients were categorized into four groups: control group represented the patients' lab results at admission before treatment protocols were applied; group 1 included patients treated with anticoagulants, Hydroxychloroquine, and antibiotics; group 2 comprised patients treated with Dexamethasone; and group 3 included patients treated with Dexamethasone and Tocilizumab.. The study paves the way into the effectiveness of combining Dexamethasone with Tocilizumab in treatment COVID-19 patients with chronic diseases.. La forma más eficaz de controlar la gravedad y la tasa de mortalidad de la enfermedad del nuevo coronavirus (COVID-19) es mediante enfoques de diagnóstico sensibles y un protocolo de tratamiento adecuado. Nuestro objetivo fue identificar el efecto de agregar corticosteroides y tocilizumab a un protocolo de tratamiento estándar en el tratamiento de pacientes con COVID-19 con enfermedad crónica a través de biomarcadores hematológicos y de laboratorio.. Este estudio se realizó de forma retrospectiva en 68 pacientes COVID-19 con enfermedad crónica que fueron tratados por diferentes protocolos terapéuticos. Los pacientes se clasificaron en cuatro grupos: el grupo de control representaba los resultados de laboratorio de los pacientes en el momento de la admisión antes de que se aplicaran los protocolos de tratamiento; el grupo 1 incluyó a pacientes tratados con anticoagulantes, hidroxicloroquina y antibióticos; el grupo 2 estaba compuesto por pacientes tratados con dexametasona; y el grupo 3 incluyó a pacientes tratados con dexametasona y tocilizumab.. El estudio allana el camino hacia la eficacia de la combinación de dexametasona con tocilizumab en el tratamiento de pacientes con COVID-19 con enfermedades crónicas.. The Child-Mother Index constitutes a potential useful risk factor indicator for statistical analyses on data after birth. The value of the Child-Mother Index based on the estimated fetal weight before birth deserves evaluation.. Six ceria supports synthesized by various synthesis methodologies were used to deposit cobalt oxide. The catalysts were thoroughly characterized, and their catalytic activity for complete methane oxidation was studied. The supports synthesized by direct calcination and precipitation with ammonia exhibited the best textural and structural properties as well as the highest degree of oxidation. The remaining supports presented poorer textural properties to be employed as catalytic supports. The cobalt deposited over the first two supports presented a good dispersion at the external surface, which induced a significant redox effect that increased the number of Co. Some studies show that children with obesity are more likely to receive a diagnosis of depression, anxiety, or attention-deficit hyperactivity disorder (ADHD). But this does not necessarily mean obesity causes these conditions. Depression, anxiety, or ADHD could cause obesity. A child's environment, including family income or their parents' mental health, could also affect a child's weight and mental health. Understanding the nature of these relationships could help scientists develop better interventions for both obesity and mental health conditions. Genetic studies may help scientists better understand the role of the environment in these conditions, but it's important to consider both the child's and their parents’ genetics in these analyses. This is because parents and children share not only genes, but also environmental conditions. For example, families that carry genetic variants associated with higher body weight might also have lower incomes, if parents have been affected by biases against heavier people in society and the workplace. Children in these families could have worse mental health because of effects of their parent’s weight, rather than their own weight. Looking at both child and adult genetics can help disentangle these processes. Hughes et al. show that a child's own body mass index, a ratio of weight and height, is not strongly associated with the child’s mental health symptoms. They analysed genetic, weight, and health survey data from about 41,000 8-year-old children and their parents. The results suggest that a child's own BMI does not have a large effect on their anxiety symptoms. There was also no clear evidence that a child's BMI affected their symptoms of depression or ADHD. These results contradict previous studies, which did not account for parental genetics. Hughes et al. suggest that, at least for eight-year-olds, factors linked with adult weight and which differ between families may be more critical to a child's mental health than a child’s own weight. For older children and adolescents, this may not be the case, and the individual’s own weight may be more important. As a result, policies designed to reduce obesity in mid-childhood are unlikely to greatly improve the mental health of children. On the other hand, policies targeting the environmental or societal factors contributing to higher body weights, bias against people with higher weights, and poor child mental health directly may be more beneficial.. The development of an efficient photocatalyst for C2 product formation from CO. Оценка антиастенического эффекта последовательной терапии левокарнитином (ЛК) и ацетилкарнитином (АЛК) пациентов с артериальной гипертензией и/или ишемической болезнью сердца (ИБС) с астеническим синдромом (АС).. В открытое сравнительное исследование были включены 120 пациентов в возрасте 54—67 лет с артериальной гипертензией и/или ИБС с АС. Пациенты 1-й группы (. У больных 1-й группы отмечено статистически значимое уменьшение различных проявлений АС. Отличия носили достоверный характер по сравнению как с исходным уровнем, так и со 2-й группой. Установлено эндотелийпротективное действие ЛК и АЛК.. Полученные результаты свидетельствуют, что у таких коморбидных пациентов использование ЛК и АЛК уменьшает выраженность проявлений АС, а установленные эндотелиотропные свойства препаратов позволяют рекомендовать их в составе комплексной персонифицированной терапии пациентов с сердечно-сосудистыми заболеваниями.. Naproxen sodium 440 mg/diphenhydramine 50 mg combination demonstrated improvement in sleep maintenance (WASO) vs. naproxen sodium 550 mg and higher efficiency in average daily pain reduction compared with the comparison groups. The treatment was well tolerated There were no serious or unexpected adverse events reported in the study.. Сравнительный анализ эффективности и безопасности новой комбинации напроксена натрия и дифенгидрамина у пациентов с неспецифическим болевым синдромом в пояснично-крестцовом отделе спины (M54.5 «Боль внизу спины») и нарушением сна (G47.0 «Нарушения засыпания и поддержания сна [бессонница]»).. Проведено проспективное многоцентровое рандомизированное открытое сравнительное в параллельных группах клиническое исследование. Пациенты были рандомизированы в 3 группы. Больные 1-й группы получали напроксен натрия (440 мг) и дифенгидрамин (50 мг), 2-й — напроксен натрия (550 мг), 3-й — парацетамол (1000 мг) и дифенгидрамин (50 мг). Исследуемые препараты пациенты принимали однократно перед сном в течение 3 дней. Все пациенты также принимали 275 мг (1 таблетка) напроксена натрия в качестве препарата фоновой терапии. Первичным критерием эффективности было общее время бодрствования после наступления сна (WASO), измеряемое методом актиграфии. Также использовались критерии оценки продолжительности и качества сна и выраженности боли.. Анализ эффективности проведен для ITT популяции (. Применение комбинации напроксена натрия (440 мг) и дифенгидрамина (50 мг) характеризовалось более выраженным поддержанием сна по сравнению с напроксеном натрия 550 мг и более высокой эффективностью в отношении снижения интенсивности боли по сравнению со 2-й и 3-й группами. Отмечена хорошая переносимость препарата, серьезных нежелательных явлений зарегистрировано не было.

Topics: Acetaminophen; Acetylcarnitine; Acetylcholinesterase; Acids; Acinetobacter baumannii; Acinetobacter Infections; Adaptation, Psychological; Adolescent; Adsorption; Adult; Aged; Alcohol Drinking; Alzheimer Disease; Amikacin; Ammonia; Anaerobiosis; Animals; Anorexia; Anti-Bacterial Agents; Anti-Infective Agents; Anti-Inflammatory Agents; Anti-Inflammatory Agents, Non-Steroidal; Antineoplastic Agents; Anxiety; Aptamers, Nucleotide; Asthenia; Attention Deficit Disorder with Hyperactivity; Bacterial Proteins; Beryllium; beta-Lactamases; Biofuels; Biomass; Biosensing Techniques; Bismuth; Blister; Body Mass Index; Body Surface Area; Boronic Acids; Brain; Breast Neoplasms; Butyrylcholinesterase; Cannabis; Carbapenems; Carbonyl Cyanide m-Chlorophenyl Hydrazone; Carboxylic Acids; Carcinoma, Hepatocellular; Cardiovascular Diseases; Carnitine; Case-Control Studies; Catalysis; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; Child; China; Cholinesterase Inhibitors; Clarithromycin; Clostridioides; Clostridioides difficile; Clostridium Infections; Cohort Studies; Colistin; Colitis; Colon; Coloring Agents; Coronary Artery Bypass; Creatinine; Crystalloid Solutions; Cytokines; Depression; Dextran Sulfate; Dextrans; Diabetes Mellitus, Type 2; Diabetic Retinopathy; Diarrhea; Dietary Supplements; Diphenhydramine; Disease Models, Animal; Disease Outbreaks; Double-Blind Method; Doxorubicin; Drosophila; Drug Tapering; Dysbiosis; Electrons; Escherichia coli; Extracellular Vesicles; Fatigue; Female; Fermentation; gamma-Cyclodextrins; Gastrointestinal Microbiome; Glucose; Graft Survival; Graft vs Host Disease; Head and Neck Neoplasms; Heart Arrest, Induced; Hematopoietic Stem Cell Transplantation; High-Intensity Interval Training; Hippocampus; Humans; Hydrogen-Ion Concentration; Hypertension; Incidence; Interferon-gamma; Italy; Kinetics; Klebsiella Infections; Klebsiella pneumoniae; Lab-On-A-Chip Devices; Lactoferrin; Larva; Length of Stay; Lignin; Liver; Liver Neoplasms; Liver Transplantation; Living Donors; Low Back Pain; Lung; Lung Volume Measurements; Macrophages; Male; Melphalan; Men; Mendelian Randomization Analysis; Meropenem; Methane; Mice; Mice, Inbred C57BL; Microbial Sensitivity Tests; Mitochondrial Proteins; Molecular Docking Simulation; Molecular Structure; Mothers; Motivation; Mycoplasma; Mycoplasma hominis; Mycoplasma Infections; NAD; Nanocomposites; Nanoparticles; Nanotubes, Carbon; Naproxen; Neovascularization, Pathologic; Neurons; Nitrates; Nucleolin; Opuntia; Paratyphoid Fever; Phenotype; Phosphatidylinositol 3-Kinases; Phytochemicals; Plant Extracts; Pregnancy; Prevalence; Prospective Studies; Proto-Oncogene Proteins c-akt; Pulmonary Disease, Chronic Obstructive; Rats; Rats, Wistar; Resveratrol; Retrospective Studies; Rifampin; Risk Factors; RNA, Messenger; Selenium; Sleep; Social Behavior; Soil; Soil Pollutants; Squamous Cell Carcinoma of Head and Neck; Staphylococcus aureus; Structure-Activity Relationship; Suicidal Ideation; Suicide; Superoxide Dismutase-1; Surveys and Questionnaires; Swimming; Syndrome; Tannins; Temperature; Transforming Growth Factor beta; Transplantation Conditioning; Treatment Outcome; Triple Negative Breast Neoplasms; Troponin T; Tumor Microenvironment; United Kingdom; Ureaplasma; Ureaplasma urealyticum; Urinary Tract Infections; Viscum; Waste Disposal Facilities; Wastewater; Water; Water Pollutants, Chemical; Wolfiporia; Young Adult

We examined gene expression, germline variant, and somatic mutation features associated with pathologic response to neoadjuvant durvalumab plus chemotherapy in basal-like triple-negative breast cancer (bTNBC).. Germline and somatic whole-exome DNA and RNA sequencing, programmed death ligand 1 (PD-L1) IHC, and stromal tumor-infiltrating lymphocyte scoring were performed on 57 patients. We validated our results using 162 patients from the GeparNuevo randomized trial.. Gene set enrichment analysis showed that pathways involved in immunity (adaptive, humoral, innate), JAK-STAT signaling, cancer drivers, cell cycle, apoptosis, and DNA repair were enriched in cases with pathologic complete response (pCR), whereas epithelial-mesenchymal transition, extracellular matrix, and TGFβ pathways were enriched in cases with residual disease (RD). Immune-rich bTNBC with RD was enriched in CCL-3, -4, -5, -8, -23, CXCL-1, -3, -6, -10, and IL1, -23, -27, -34, and had higher expression of macrophage markers compared with immune-rich cancers with pCR that were enriched in IFNγ, IL2, -12, -21, chemokines CXCL-9, -13, CXCR5, and activated T- and B-cell markers (GZMB, CD79A). In the validation cohort, an immune-rich five-gene signature showed higher expression in pCR cases in the durvalumab arm (P = 0.040) but not in the placebo arm (P = 0.923) or in immune-poor cancers. Independent of immune markers, tumor mutation burden was higher, and PI3K, DNA damage repair, MAPK, and WNT/β-catenin signaling pathways were enriched in germline and somatic mutations in cases with pCR.. The TGFβ pathway is associated with immune-poor phenotype and RD in bTNBC. Among immune-rich bTNBC RD, macrophage/neutrophil chemoattractants dominate the cytokine milieu, and IFNγ and activated B cells and T cells dominate immune-rich cancers with pCR.

Topics: Albumins; Antibodies, Monoclonal; Antineoplastic Combined Chemotherapy Protocols; Biomarkers, Tumor; Breast Neoplasms; Cyclophosphamide; Doxorubicin; Female; Humans; Neoadjuvant Therapy; Paclitaxel; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

The most effective way to control severity and mortality rate of the novel coronavirus disease (COVID-19) is through sensitive diagnostic approaches and an appropriate treatment protocol. We aimed to identify the effect of adding corticosteroid and Tocilizumab to a standard treatment protocol in treating COVID-19 patients with chronic disease through hematological and lab biomarkers.. This study was performed retrospectively on 68 COVID-19 patients with chronic disease who were treated by different therapeutic protocols. The patients were categorized into four groups: control group represented the patients' lab results at admission before treatment protocols were applied; group 1 included patients treated with anticoagulants, Hydroxychloroquine, and antibiotics; group 2 comprised patients treated with Dexamethasone; and group 3 included patients treated with Dexamethasone and Tocilizumab.. The study paves the way into the effectiveness of combining Dexamethasone with Tocilizumab in treatment COVID-19 patients with chronic diseases.. La forma más eficaz de controlar la gravedad y la tasa de mortalidad de la enfermedad del nuevo coronavirus (COVID-19) es mediante enfoques de diagnóstico sensibles y un protocolo de tratamiento adecuado. Nuestro objetivo fue identificar el efecto de agregar corticosteroides y tocilizumab a un protocolo de tratamiento estándar en el tratamiento de pacientes con COVID-19 con enfermedad crónica a través de biomarcadores hematológicos y de laboratorio.. Este estudio se realizó de forma retrospectiva en 68 pacientes COVID-19 con enfermedad crónica que fueron tratados por diferentes protocolos terapéuticos. Los pacientes se clasificaron en cuatro grupos: el grupo de control representaba los resultados de laboratorio de los pacientes en el momento de la admisión antes de que se aplicaran los protocolos de tratamiento; el grupo 1 incluyó a pacientes tratados con anticoagulantes, hidroxicloroquina y antibióticos; el grupo 2 estaba compuesto por pacientes tratados con dexametasona; y el grupo 3 incluyó a pacientes tratados con dexametasona y tocilizumab.. El estudio allana el camino hacia la eficacia de la combinación de dexametasona con tocilizumab en el tratamiento de pacientes con COVID-19 con enfermedades crónicas.. The Child-Mother Index constitutes a potential useful risk factor indicator for statistical analyses on data after birth. The value of the Child-Mother Index based on the estimated fetal weight before birth deserves evaluation.. Six ceria supports synthesized by various synthesis methodologies were used to deposit cobalt oxide. The catalysts were thoroughly characterized, and their catalytic activity for complete methane oxidation was studied. The supports synthesized by direct calcination and precipitation with ammonia exhibited the best textural and structural properties as well as the highest degree of oxidation. The remaining supports presented poorer textural properties to be employed as catalytic supports. The cobalt deposited over the first two supports presented a good dispersion at the external surface, which induced a significant redox effect that increased the number of Co. Some studies show that children with obesity are more likely to receive a diagnosis of depression, anxiety, or attention-deficit hyperactivity disorder (ADHD). But this does not necessarily mean obesity causes these conditions. Depression, anxiety, or ADHD could cause obesity. A child's environment, including family income or their parents' mental health, could also affect a child's weight and mental health. Understanding the nature of these relationships could help scientists develop better interventions for both obesity and mental health conditions. Genetic studies may help scientists better understand the role of the environment in these conditions, but it's important to consider both the child's and their parents’ genetics in these analyses. This is because parents and children share not only genes, but also environmental conditions. For example, families that carry genetic variants associated with higher body weight might also have lower incomes, if parents have been affected by biases against heavier people in society and the workplace. Children in these families could have worse mental health because of effects of their parent’s weight, rather than their own weight. Looking at both child and adult genetics can help disentangle these processes. Hughes et al. show that a child's own body mass index, a ratio of weight and height, is not strongly associated with the child’s mental health symptoms. They analysed genetic, weight, and health survey data from about 41,000 8-year-old children and their parents. The results suggest that a child's own BMI does not have a large effect on their anxiety symptoms. There was also no clear evidence that a child's BMI affected their symptoms of depression or ADHD. These results contradict previous studies, which did not account for parental genetics. Hughes et al. suggest that, at least for eight-year-olds, factors linked with adult weight and which differ between families may be more critical to a child's mental health than a child’s own weight. For older children and adolescents, this may not be the case, and the individual’s own weight may be more important. As a result, policies designed to reduce obesity in mid-childhood are unlikely to greatly improve the mental health of children. On the other hand, policies targeting the environmental or societal factors contributing to higher body weights, bias against people with higher weights, and poor child mental health directly may be more beneficial.. The development of an efficient photocatalyst for C2 product formation from CO. Оценка антиастенического эффекта последовательной терапии левокарнитином (ЛК) и ацетилкарнитином (АЛК) пациентов с артериальной гипертензией и/или ишемической болезнью сердца (ИБС) с астеническим синдромом (АС).. В открытое сравнительное исследование были включены 120 пациентов в возрасте 54—67 лет с артериальной гипертензией и/или ИБС с АС. Пациенты 1-й группы (. У больных 1-й группы отмечено статистически значимое уменьшение различных проявлений АС. Отличия носили достоверный характер по сравнению как с исходным уровнем, так и со 2-й группой. Установлено эндотелийпротективное действие ЛК и АЛК.. Полученные результаты свидетельствуют, что у таких коморбидных пациентов использование ЛК и АЛК уменьшает выраженность проявлений АС, а установленные эндотелиотропные свойства препаратов позволяют рекомендовать их в составе комплексной персонифицированной терапии пациентов с сердечно-сосудистыми заболеваниями.. Naproxen sodium 440 mg/diphenhydramine 50 mg combination demonstrated improvement in sleep maintenance (WASO) vs. naproxen sodium 550 mg and higher efficiency in average daily pain reduction compared with the comparison groups. The treatment was well tolerated There were no serious or unexpected adverse events reported in the study.. Сравнительный анализ эффективности и безопасности новой комбинации напроксена натрия и дифенгидрамина у пациентов с неспецифическим болевым синдромом в пояснично-крестцовом отделе спины (M54.5 «Боль внизу спины») и нарушением сна (G47.0 «Нарушения засыпания и поддержания сна [бессонница]»).. Проведено проспективное многоцентровое рандомизированное открытое сравнительное в параллельных группах клиническое исследование. Пациенты были рандомизированы в 3 группы. Больные 1-й группы получали напроксен натрия (440 мг) и дифенгидрамин (50 мг), 2-й — напроксен натрия (550 мг), 3-й — парацетамол (1000 мг) и дифенгидрамин (50 мг). Исследуемые препараты пациенты принимали однократно перед сном в течение 3 дней. Все пациенты также принимали 275 мг (1 таблетка) напроксена натрия в качестве препарата фоновой терапии. Первичным критерием эффективности было общее время бодрствования после наступления сна (WASO), измеряемое методом актиграфии. Также использовались критерии оценки продолжительности и качества сна и выраженности боли.. Анализ эффективности проведен для ITT популяции (. Применение комбинации напроксена натрия (440 мг) и дифенгидрамина (50 мг) характеризовалось более выраженным поддержанием сна по сравнению с напроксеном натрия 550 мг и более высокой эффективностью в отношении снижения интенсивности боли по сравнению со 2-й и 3-й группами. Отмечена хорошая переносимость препарата, серьезных нежелательных явлений зарегистрировано не было.

Topics: Acetaminophen; Acetylcarnitine; Acetylcholinesterase; Acids; Acinetobacter baumannii; Acinetobacter Infections; Adaptation, Psychological; Adolescent; Adsorption; Adult; Aged; Alcohol Drinking; Alzheimer Disease; Amikacin; Ammonia; Anaerobiosis; Animals; Anorexia; Anti-Bacterial Agents; Anti-Infective Agents; Anti-Inflammatory Agents; Anti-Inflammatory Agents, Non-Steroidal; Antineoplastic Agents; Anxiety; Aptamers, Nucleotide; Asthenia; Attention Deficit Disorder with Hyperactivity; Bacterial Proteins; Beryllium; beta-Lactamases; Biofuels; Biomass; Biosensing Techniques; Bismuth; Blister; Body Mass Index; Body Surface Area; Boronic Acids; Brain; Breast Neoplasms; Butyrylcholinesterase; Cannabis; Carbapenems; Carbonyl Cyanide m-Chlorophenyl Hydrazone; Carboxylic Acids; Carcinoma, Hepatocellular; Cardiovascular Diseases; Carnitine; Case-Control Studies; Catalysis; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; Child; China; Cholinesterase Inhibitors; Clarithromycin; Clostridioides; Clostridioides difficile; Clostridium Infections; Cohort Studies; Colistin; Colitis; Colon; Coloring Agents; Coronary Artery Bypass; Creatinine; Crystalloid Solutions; Cytokines; Depression; Dextran Sulfate; Dextrans; Diabetes Mellitus, Type 2; Diabetic Retinopathy; Diarrhea; Dietary Supplements; Diphenhydramine; Disease Models, Animal; Disease Outbreaks; Double-Blind Method; Doxorubicin; Drosophila; Drug Tapering; Dysbiosis; Electrons; Escherichia coli; Extracellular Vesicles; Fatigue; Female; Fermentation; gamma-Cyclodextrins; Gastrointestinal Microbiome; Glucose; Graft Survival; Graft vs Host Disease; Head and Neck Neoplasms; Heart Arrest, Induced; Hematopoietic Stem Cell Transplantation; High-Intensity Interval Training; Hippocampus; Humans; Hydrogen-Ion Concentration; Hypertension; Incidence; Interferon-gamma; Italy; Kinetics; Klebsiella Infections; Klebsiella pneumoniae; Lab-On-A-Chip Devices; Lactoferrin; Larva; Length of Stay; Lignin; Liver; Liver Neoplasms; Liver Transplantation; Living Donors; Low Back Pain; Lung; Lung Volume Measurements; Macrophages; Male; Melphalan; Men; Mendelian Randomization Analysis; Meropenem; Methane; Mice; Mice, Inbred C57BL; Microbial Sensitivity Tests; Mitochondrial Proteins; Molecular Docking Simulation; Molecular Structure; Mothers; Motivation; Mycoplasma; Mycoplasma hominis; Mycoplasma Infections; NAD; Nanocomposites; Nanoparticles; Nanotubes, Carbon; Naproxen; Neovascularization, Pathologic; Neurons; Nitrates; Nucleolin; Opuntia; Paratyphoid Fever; Phenotype; Phosphatidylinositol 3-Kinases; Phytochemicals; Plant Extracts; Pregnancy; Prevalence; Prospective Studies; Proto-Oncogene Proteins c-akt; Pulmonary Disease, Chronic Obstructive; Rats; Rats, Wistar; Resveratrol; Retrospective Studies; Rifampin; Risk Factors; RNA, Messenger; Selenium; Sleep; Social Behavior; Soil; Soil Pollutants; Squamous Cell Carcinoma of Head and Neck; Staphylococcus aureus; Structure-Activity Relationship; Suicidal Ideation; Suicide; Superoxide Dismutase-1; Surveys and Questionnaires; Swimming; Syndrome; Tannins; Temperature; Transforming Growth Factor beta; Transplantation Conditioning; Treatment Outcome; Triple Negative Breast Neoplasms; Troponin T; Tumor Microenvironment; United Kingdom; Ureaplasma; Ureaplasma urealyticum; Urinary Tract Infections; Viscum; Waste Disposal Facilities; Wastewater; Water; Water Pollutants, Chemical; Wolfiporia; Young Adult

The main mechanism underlying cancer dissemination is the epithelial to mesenchymal transition (EMT). This process is orchestrated by cytokines like TGFβ, involving "non-canonical" AKT- or STAT3-driven pathways. Recently, the alteration of copper homeostasis seems involved in the onset and progression of cancer.. We expose different breast cancer cell lines, including two triple negative (TNBC) ones, an HER2 enriched and one cell line representative of the Luminal A molecular subtype, to short- or long-term copper-chelation by triethylenetetramine (TRIEN). We analyse changes in the expression of EMT markers (E-cadherin, fibronectin, vimentin and αSMA), in the levels and activity of extracellular matrix components (LOXL2, fibronectin and MMP2/9) and of copper homeostasis markers by Western blot analyses, immunofluorescence, enzyme activity assays and RT-qPCR. Boyden Chamber and wound healing assays revealed the impact of copper chelation on cell migration. Additionally, we explored whether perturbation of copper homeostasis affects EMT prompted by TGFβ. Metabolomic and lipidomic analyses were applied to search the effects of copper chelation on the metabolism of breast cancer cells. Finally, bioinformatics analysis of data on breast cancer patients obtained from different databases was employed to correlate changes in kinases and copper markers with patients' survival.. Remarkably, only HER2 negative breast cancer cells differently responded to short- or long-term exposure to TRIEN, initially becoming more aggressive but, upon prolonged exposure, retrieving epithelial features, reducing their invasiveness. This phenomenon may be related to the different impact of the short and prolonged activation of the AKT kinase and to the repression of STAT3 signalling. Bioinformatics analyses confirmed the positive correlation of breast cancer patients' survival with AKT activation and up-regulation of CCS. Eventually, metabolomics studies demonstrate a prevalence of glycolysis over mitochondrial energetic metabolism and of lipidome changes in TNBC cells upon TRIEN treatment.. We provide evidence of a pivotal role of copper in AKT-driven EMT activation, acting independently of HER2 in TNBC cells and via a profound change in their metabolism. Our results support the use of copper-chelators as an adjuvant therapeutic strategy for TNBC.

Topics: Amino Acid Oxidoreductases; Biological Availability; Cell Line, Tumor; Cell Movement; Copper; Epithelial-Mesenchymal Transition; Fibronectins; Humans; Proto-Oncogene Proteins c-akt; Transforming Growth Factor beta; Trientine; Triple Negative Breast Neoplasms

Karyopherin-α3 (KPNA3), a karyopherin- α isoform, is intimately associated with metastatic progression via epithelial-mesenchymal transition (EMT). However, the molecular mechanism underlying how KPNA3 acts as an EMT inducer remains to be elucidated. In this report, we identified that KPNA3 was significantly upregulated in cancer cells, particularly in triple-negative breast cancer, and its knockdown resulted in the suppression of cell proliferation and metastasis. The comprehensive transcriptome analysis from KPNA3 knockdown cells indicated that KPNA3 is involved in the regulation of numerous EMTrelated genes, including the downregulation of GATA3 and E-cadherin and the up-regulation of HAS2. Moreover, it was found that KPNA3 EMT-mediated metastasis can be achieved by TGF-β or AKT signaling pathways; this suggests that the novel independent signaling pathways KPNA3-TGF-β-GATA3-HAS2/E-cadherin and KPNA3-AKT-HAS2/E-cadherin are involved in the EMT-mediated progress of TNBC MDA-MB-231 cells. These findings provide new insights into the divergent EMT inducibility of KPNA3 according to cell and cancer type. [BMB Reports 2023; 56(2): 120-125].

Topics: alpha Karyopherins; Breast Neoplasms; Cadherins; Cell Line, Tumor; Cell Movement; Cell Proliferation; Epithelial-Mesenchymal Transition; Female; Humans; Proto-Oncogene Proteins c-akt; Signal Transduction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

In triple-negative breast cancer (TNBC), the pleiotropic NDRG1 (N-Myc downstream regulated gene 1) promotes progression and worse survival, yet contradictory results were documented, and the mechanisms remain unknown. Phosphorylation and localization could drive NDRG1 pleiotropy, nonetheless, their role in TNBC progression and clinical outcome was not investigated. We found enhanced p-NDRG1 (Thr346) by TGFβ1 and explored whether it drives NDRG1 pleiotropy and TNBC progression. In tissue microarrays of 81 TNBC patients, we identified that staining and localization of NDRG1 and p-NDRG1 (Thr346) are biomarkers and risk factors associated with shorter overall survival. We found that TGFβ1 leads NDRG1, downstream of GSK3β, and upstream of NF-κB, to differentially regulate migration, invasion, epithelial-mesenchymal transition, tumor initiation, and maintenance of different populations of cancer stem cells (CSCs), depending on the progression stage of tumor cells, and the combination of TGFβ and GSK3β inhibitors impaired CSCs. The present study revealed the striking importance to assess both total NDRG1 and p-NDRG1 (Thr346) positiveness and subcellular localization to evaluate patient prognosis and their stratification. NDRG1 pleiotropy is driven by TGFβ to differentially promote metastasis and/or maintenance of CSCs at different stages of tumor progression, which could be abrogated by the inhibition of TGFβ and GSK3β.

Topics: Cell Cycle Proteins; Cell Line, Tumor; Gene Expression Regulation, Neoplastic; Glycogen Synthase Kinase 3 beta; Humans; Intracellular Signaling Peptides and Proteins; NF-kappa B; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Among breast cancer patients, metastases are the leading cause of death. Despite decades of effort, little progress has been made to improve the treatment of breast cancer metastases, especially triple-negative breast cancer (TNBC). The extracellular matrix plays an important role in tumour growth and metastasis by causing its deposition, remodelling, and signalling. As we know, the process of fibrosis results in excessive amounts of extracellular matrix being deposited within the cells. So, it will be interesting to study if the use of anti-fibrotic drugs in combination with conventional chemotherapy drugs can produce synergistic antitumor effects. In this study, we assessed the efficacy of Pirfenidone (PFD), an FDA-approved medication for the treatment of idiopathic pulmonary fibrosis, on TNBC cells as well as its anti-tumour effects in xenograft tumour model. PFD inhibited in a dose-dependent manner breast cancer cell proliferation, migration, and invasion, while promoted their apoptosis in vitro. PFD also suppressed TGF-β-induced activation of Smad signalling pathway and expression level of EMT-inducing transcription factors (e.g. SNAI2, TWIST1, ZEB1) as well as the mesenchymal genes such as VIMENTIN and N-Cadherin. On the contrary, the expression level of epithelial marker gene E-Cadherin was up-regulated in the presence of PFD. In vivo, PFD alone exerted a milder but significant anti-tumour effect than the chemotherapy drug nanoparticle albumin-bound paclitaxel (nab-PTX) did in the breast cancer xenograft mouse model. Interestingly, PFD synergistically boosted the cancer-killing effect of nab-PTX. Furthermore, Our data suggest that PFD suppressed breast cancer metastasis by inhibiting the activity of the TGFβ/SMAD pathway.

Topics: Animals; Cell Line, Tumor; Cell Movement; Epithelial-Mesenchymal Transition; Humans; Mice; Signal Transduction; Smad Proteins; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Triple-negative breast cancer (TNBC) is the most challenging breast cancer subtype to treat due to its aggressive characteristics and low response to the existing clinical therapies. Distant metastasis is the main cause of death of TNBC patients. Better understanding of the mechanisms underlying TNBC metastasis may lead to new strategies of early diagnosis and more efficient treatment. In our study, we uncovered that the autophagy receptor optineurin (OPTN) plays an unexpected role in TNBC metastasis. Data mining of publicly available data bases revealed that the mRNA level of OPTN in TNBC patients positively correlates with relapse free and distance metastasis free survival. Importantly, in vitro and in vivo models demonstrated that OPTN suppresses TNBC metastasis. Mechanistically, OPTN inhibited the pro-oncogenic transforming growth factor-β (TGFβ) signaling in TNBC cells by interacting with TGFβ type I receptor (TβRI) and promoting its ubiquitination for degradation. Consistent with our experimental findings, the clinical TNBC samples displayed a negative correlation between OPTN mRNA expression and TGFβ gene response signature and expression of proto-typic TGFβ target genes. Altogether, our study demonstrates that OPTN is a negative regulator for TGFβ receptor/SMAD signaling and suppresses metastasis in TNBC.

Topics: Cell Cycle Proteins; Cell Line, Tumor; Humans; Membrane Transport Proteins; Neoplasm Metastasis; Neoplasm Recurrence, Local; RNA, Messenger; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Triple-negative breast cancer (TNBC) has a poor prognosis due to the lack of specific and highly effective therapeutic agents. Cancer stem cells (CSCs) are one of the main factors contributing to TNBC relapse and metastasis. Therefore, targeting CSCs selectively with small molecules is a novel strategy for drug development. In this study, the natural product harmine (HM) was identified as a hit compound from 2632 natural product monomers based on phenotypic screening of a 2D assay and patient-derived organoid (PDO) model that was established from a patient who had multiple drug resistance and various visceral and contralateral breast metastases. Next, harmine was further modified and optimized to obtain a lead compound (YH677) with a tetrahydro-β-carboline scaffold. YH677 showed potent antiproliferative and antimigratory activities against several TNBC cell lines in vitro. In addition, YH677 inhibited epithelial mesenchymal transition (EMT) and stem cell marker expression in a dose-dependent manner. More importantly, YH677 suppressed breast cancer growth and metastasis in orthotopic, metastatic xenograft and patient-derived xenograft (PDX) models in vivo. Mechanistic studies showed that YH677 inhibits the expansion of CSCs by regulating the TGFβ/Smad signaling pathway. These preclinical data provide a basis for the development of YH677 as a lead compound for TNBC treatment.

Topics: Animals; Cell Line, Tumor; Cell Proliferation; Disease Models, Animal; Epithelial-Mesenchymal Transition; Harmine; Humans; Neoplasm Recurrence, Local; Neoplastic Stem Cells; Signal Transduction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

The poor permeability of therapeutic drugs, limited T-cell infiltration, and strong immunosuppressive tumor microenvironment of triple-negative breast cancer (TNBC) acts as a prominent barrier to the delivery of drugs and immunotherapy including programmed cell death ligand-1 antibody (anti-PD-L1). Transforming growth factor (TGF)-β, an important cytokine produced by cancer-associated fibroblasts (CAFs) and tumor cells contributes to the pathological vasculature, dense tumor stroma and strong immunosuppressive tumor microenvironment (TME). Herein, a nanomedicine platform (HA-LSL/siTGF-β) employing dual-targeting, alongside hyaluronidase (HAase) and glutathione (GSH) triggered release was elaborately constructed to efficiently deliver TGF-β small interference RNA (siTGF-β). It was determined that this system was able to improve the efficacy of anti-PD-L1. The siTGF-β nanosystem efficiently silenced TGF-β-related signaling pathways in both activated NIH 3T3 cells and 4T1 cells in vitro and in vivo. This occurred firstly, through CD44-mediated uptake, followed by rapid escape mediated by HAase in endo/lysosomes and release of siRNA mediated by high GSH concentrations in the cytoplasm. By simultaneous silencing of TGF-β in stromal and tumor cells, HA-LSL/siTGF-β dramatically reduced stroma deposition, promoted the penetration of nanomedicines for deep remodeling of the TME, improved oxygenation, T cells infiltration and subsequent anti-PD-L1 deep penetration. The double suppression of TGF-β has been demonstrated to promote blood vessel normalization, inhibit an epithelial-to-mesenchymal transition (EMT), and further modify the immunosuppressive TME, which was supported by an overall increase in the proportion of dendritic cells and cytotoxic T cells. Further, a reduction in the proportion of immunosuppression cells such as regulatory T cells and myeloid-derived suppressor cells was also observed in the TME. Based on the comprehensive remodeling of the tumor microenvironment by this nanosystem, subsequent anti-PD-L1 therapy elicited robust antitumor immunity. Specifically, this system was able to suppress the growth of both primary and distant tumor while preventing tumor metastasis to the lung. Therefore, the combination of the dual-targeted siTGF-β nanosystem, alongside anti-PD-L1 may serve as a novel method to enhance antitumor immunotherapy against stroma-rich TNBC.

Topics: Animals; Cell Line, Tumor; Humans; Immunotherapy; Mice; RNA, Small Interfering; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Tumor Microenvironment

The activation of RIG-I-like receptor (RLR) signaling in cancer cells is widely recognized as a critical cancer therapy method. The expected mechanism of RLR ligand-mediated cancer therapy involves the promotion of cancer cell death and strong induction of interferon (IFN)-β that affects the tumor microenvironment. We have recently shown that activation of RLR signaling in triple-negative breast cancer cells (TNBC) attenuates transforming growth factor-β (TGF-β) signaling, which partly contributes to the promotion of cancer cell pyroptosis. However, the consequences of suppression of TGF-β signaling by RLR ligands with respect to IFN-β-mediated tumor suppression are not well characterized. This study showed that transfection of a typical RLR ligand polyI:C in cancer cells produces significant levels of IFN-β, which inhibits the growth of the surrounding cancer cells. In addition, IFN-β-induced cell cycle arrest in surrounding cancer cells was inhibited by the expression of constitutively active Smad3. Constitutively active Smad3 suppresses IFN-β expression through the alleviation of IFN regulatory factor 3 binding to the canonical target genes, as suggested by ChIP sequencing analysis. Based on these findings, a new facet of the protumorigenic function of TGF-β that suppresses IFN-β expression is suggested when RLR-mediated cancer treatment is used in TNBC.

Topics: Cell Death; Cell Line, Tumor; Cell Proliferation; Gene Expression Regulation, Neoplastic; Humans; Interferon Regulatory Factor-3; Interferon-beta; Poly I-C; Signal Transduction; Smad3 Protein; Transfection; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Tumor Microenvironment

Triple-negative breast cancer (TNBC) represents the most aggressive subtype of breast cancer that is highly resistant to current therapeutic options. According to the public databases Oncomine and KM plotter, the CLK4 expression is correlated with poor patient survival in TNBC, especially in mesenchymal-like TNBC (MES-TNBC) that has strong metastatic potential. Therefore, we investigated the potential involvement of CLK4 in the metastasis and progression of MES-TNBC. In the MES-TNBC cell lines, the CLK4 expression was elevated. Notably, the RNAi-mediated silencing of CLK4 reduced the expression of multiple epithelial-mesenchymal transition (EMT) genes that mediate metastasis. Furthermore, CLK4 silencing reduced both the invasive behaviors of the cultured cells and tumor metastasis in the mouse xenograft model. It is also noteworthy that CLK4 silencing repressed the invasive and cancer stem cell (CSC) properties that are induced by the TGF-β signaling. Importantly, the pharmacological inhibition of CLK4 potently repressed the invasion and proliferation of MES-TNBC cell lines and patient-derived cells, which demonstrates its clinical applicability. Collectively, our results suggest that CLK4 plays a crucial role in invasion and proliferation of MES-TNBC, especially in the processes that are induced by TGF-β. Also, this study characterizes CLK4 as a novel therapeutic target in breast cancer.

Topics: Animals; Cell Line, Tumor; Cell Movement; Cell Proliferation; Epithelial-Mesenchymal Transition; Humans; Mice; Neoplastic Stem Cells; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Although chemotherapy and recently approved immunotherapies have improved treatment of triple-negative breast cancer (TNBC), the clinical outcome for this deadly disease remains unsatisfactory. We found that both cluster of differentiation 73 (CD73) and transforming growth factor (TGF)β were elevated in TNBC and correlated with the epithelial-mesenchymal transition (EMT), fibrotic stroma, an immune-tolerant tumor environment, and poor prognosis. To explore the efficacy of CD73-TGFβ dual-blockade, we generated a bifunctional anti-CD73-TGFβ construct consisting of the CD73 antibody MEDI9447 fused with the TGFβRII extracellular-domain (termed MEDI-TGFβR). MEDI-TGFβR retained full and simultaneous blocking efficiency for CD73 and TGFβ. Compared with MEDI9447 activity alone, MEDI-TGFβR demonstrated superior inhibitory activity against CD73-dependent cell migration and the EMT in CD73-high TNBC cells and effectively reduced lung metastasis in a syngeneic mouse model of TNBC. Mechanistically, the CD73-TGFβ dual-blockade reverted the EMT and stromal fibrosis and induced tumor cell death, which was accompanied by the accumulation of M1-macrophages and production of tumor necrosis factor α (TNFα). The CD73-TGFβ dual-blockade promoted a multifaceted inflammatory tumor microenvironment, as shown by the diminished levels of myeloid-derived suppressor cells (MDSCs) and M2-macrophages, and substantially increased levels of activated dendritic cells, cytotoxic T cells, and B cells. Collectively, our results have highlighted a novel strategy for TNBC treatment.

Topics: 5'-Nucleotidase; Animals; Antigens, Differentiation, B-Lymphocyte; Cell Line, Tumor; Cell Movement; Epithelial-Mesenchymal Transition; Histocompatibility Antigens Class II; Mice; Signal Transduction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Tumor Microenvironment

Diacylglycerol kinase ζ (DGKZ) is a diacylglycerol kinase that metabolizes diacylglycerol to yield phosphatidic acid, and its function in breast cancer progression remains unclear. In this study, via screening of a CRISPR-Cas9 knockout library containing lipid metabolic genes, DGKZ was identified as a potential prometastatic gene. We first confirmed that high DGKZ expression correlated with tumor progression and poor prognosis in patients. Next, knockout of DGKZ in triple-negative breast cancer cell lines were found to significantly inhibit metastatic behaviors in vitro and in vivo, whereas its overexpression increased the metastatic potential of cell lines. Mechanistic studies based on RNA sequencing and bioinformatic analysis indicated that DGKZ might regulate cell metastasis by promoting epithelial-mesenchymal transition via the transforming growth factor β (TGFβ) signaling pathway. Furthermore, we found that overexpression of DGKZ activated the TGFβ/TGFβR2/Smad3 signaling pathway by inhibiting the degradation of TGFβR2 through suppression of caveolin/lipid raft-dependent endocytosis. Moreover, the caveolin/lipid raft-dependent endocytosis of TGFβR2 was regulated by the metabolite phosphatidic acid, which might alter TGFβR2 partitioning in lipid rafts and nonlipid rafts by affecting the fluidity of the plasma membrane. These findings suggested that DGKZ is a novel promoter of metastasis and that it could be a potential prognostic indicator in patients with triple-negative breast cancer.

Topics: Caveolins; Cell Line, Tumor; Cell Proliferation; Diacylglycerol Kinase; Endocytosis; Humans; Membrane Microdomains; Phosphatidic Acids; Receptor, Transforming Growth Factor-beta Type II; Signal Transduction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Triple-negative breast cancer is the most aggressive subtype of breast cancer and the incidence of depression in breast cancer patients is high, which leading to worse survival and increased risk of recurrence. The effect of antidepressants on breast cancer patients remains contradictory, which might be due to variations in antidepression targets. Therefore, there is significant value to explore the antitumor potential of antidepressants and discover new therapeutic targets for breast patients. The authors screen antidepressant-related oncogenes or suppressors by using siRNAs. After combining functional experiments with online database analysis, 5-hydroxytryptamine receptor 1A (HTR1A is selected with antitumor potential in breast cancer cells in vivo and in vitro. RNA-seq analysis and coimmunoprecipitation assays indicate that HTR1A interacts with TRIM21 and PSMD7 to inhibit the degradation of TβRII through the ubiquitin-proteasome pathway, thereby inhibiting the transforming growth factor-β (TGF-β) canonical and noncanonical pathway. In addition, HTR1A is an independent predictive factor for breast cancer patients. The combined treatment of HTR1A agonists with demethylation drugs may significantly improve patient survival. It is of great significance to clarify the function and mechanism of the depression-related gene HTR1A in breast cancer, which might provide a new approach for triple-negative breast cancer patients.

Topics: Humans; Prognosis; Receptor, Serotonin, 5-HT1A; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

The development of drug resistance remains to be a major cause of therapeutic failure in breast cancer patients. How drug-sensitive cells first evade drug inhibition to proliferate remains to be fully investigated.. Here we characterized the early transcriptional evolution in response to TGF-β in the human triple-negative breast cells through bioinformatical analysis using a published RNA-seq dataset, for which MCF10A cells were treated with 5 ng/ml TGF-β1 for 0 h, 24 h, 48 h and 72 h, and the RNA-seq were performed in biological duplicates. The protein-protein interaction networks of the differentially expressed genes were constructed. KEGG enrichment analysis, cis-regulatory sequence analysis and Kaplan-Meier analysis were also performed to analyze the cellular reprograming induced by TGF-β and its contribution to the survival probability decline of breast cancer patients.. Transcriptomic analysis revealed that cell growth was severely suppressed by TGF-β in the first 24 h but this anti-proliferate impact attenuated between 48 h and 72 h. The oncogenic actions of TGF-β happened within the same time frame with its anti-proliferative effects. In addition, sustained high expression of several drug resistance markers was observed after TGF-β treatment. We also identified 17 TGF-β induced genes that were highly correlated with the survival probability decline of breast cancer patients.. Together, TGF-β plays an important role in tumorigenesis and the development of drug resistance, which implies potential therapeutic strategies targeting the early-stage TGF-β signaling activities.

Topics: Cell Line, Tumor; Cell Proliferation; Drug Resistance, Neoplasm; Female; Gene Expression Profiling; Humans; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Triple-negative breast cancer (TNBC) is the most severe type of breast cancer owing to its high heterogeneity, aggressiveness and lack of treatment. Studies have reported that uncarboxylated osteocalcin (GluOC) promotes the development of prostate and other cancers. Studies have also found elevated levels of serum osteocalcin in breast cancer patients with bone metastasis, and serum osteocalcin can be a marker of bone metastasis. However, whether GluOC promotes the development of TNBC and the related mechanisms need to be further clarified.. Our results revealed that GluOC is associated with the proliferation and metastasis of MDA-MB-231 cells. GluOC increased the viability and proliferation of MDA-MB-231 cells. In addition, GluOC enhanced the metastatic ability of MDA-MB-231 cells by promoting the expression of matrix metalloproteinase-2 (MMP2), matrix metalloproteinase-13 (MMP13), and vascular endothelial growth factor (VEGF) and inducing epithelial-mesenchymal transition (EMT). We also found that GluOC upregulated the expression of interleukin-8 (IL-8) and parathyroid hormone-related protein (PTHrP) genes in MDA-MB-231 breast cancer cells. Moreover, the promoting effect of GluOC was reversed in MDA-MB-231 breast cancer cells treated with specific inhibitor of SMAD3 (SIS3), a SMAD3 phosphorylation inhibitor.. Our research proved for the first time that GluOC facilitates the proliferation and metastasis of MDA-MB-231 cells by accelerating the transforming growth factor-β (TGF-β)/SMAD3 signaling pathway. Moreover, GluOC also promotes the gene expression of IL-8 and PTHrP. Both IL-8 and PTHrP can act as osteolytic factors in breast cancer cells. This study indicates that GluOC may be a useful target for preventing TNBC bone metastasis.

Topics: Cell Proliferation; Humans; Interleukin-8; Matrix Metalloproteinase 2; Osteocalcin; Parathyroid Hormone-Related Protein; Signal Transduction; Smad3 Protein; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Vascular Endothelial Growth Factor A

Despite the remarkable success of immunotherapies over the past decade, their effectiveness against triple-negative breast cancer (TNBC) is limited to a small subset of patients, mainly due to the low immunogenicity and unfavorable tumor microenvironment. In this study, we successfully constructed a programmed site-specific delivery nanosystem for the combined delivery of transforming growth factor beta (TGF-β) receptor inhibitor LY3200882 (LY) and PD-L1 siRNA (siPD-L1) to boost anti-tumor immunotherapy. As expected, LY in the outer layer of the nanosystem was released by stimulation of MMP2, and dramatically down-regulated the expression of extracellular matrix (ECM) in the tumor-associated fibroblasts (TAFs), and thus promoted the infiltration of effector T cells and penetration of nanomedicines. Simultaneously, the blockade of TGF-β by LY also triggered immunogenic cell death (ICD) of tumor cells and induced the maturation of dendritic cells. Moreover, the programmed design provided the siPD-L1/protamine cationic inner core with easier access to tumor cells and TAFs after MMP2-stimulated breakup of the outer layer, down-regulating the expression of PD-L1 in both types of cells. Notably, the synergistic effect of LY and siPD-L1 remarkably enhanced the tumor antigen presentation and immunosuppressive microenvironment remodeling, thus efficiently inhibiting the TNBC growth, metastasis, and recurrence. Therefore, the programmed site-specific delivery nanosystem is a promising drug delivery platform for boosting anti-tumor immunotherapy efficacy for TNBC.

Topics: Antineoplastic Agents; B7-H1 Antigen; Cell Line, Tumor; Humans; Immunologic Factors; Immunotherapy; Matrix Metalloproteinase 2; RNA, Small Interfering; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Tumor Microenvironment

Triple-negative breast cancer (TNBC) is a highly aggressive and invasive type of breast cancer. In addition, type 2 diabetes mellitus (T2DM) is recognized as a risk factor for cancer metastasis, which is associated with mortality in patients with breast cancer. Cancer-associated fibroblasts (CAFs) generated from adipose tissue-derived mesenchymal stem cells (AT-MSCs) play a vital role in the progression of TNBC. However, to date, whether T2DM affects the ability of AT-MSCs to differentiate into CAFs is still unclear. In this study, we found that in coculture with TNBC cells [breast cancer cells (BCCs)] under hypoxic conditions, AT-MSCs derived from T2DM donors (dAT-MSCs) were facilitated to differentiate into CAFs, which showed fibroblastic morphology and the induced expression of fibroblastic markers, such as fibroblast activation protein, fibroblast-specific protein, and vimentin. This was involved in the higher expression of transforming growth factor beta receptor 2 (TGFβR2) and the phosphorylation of Smad2/3. Furthermore, T2DM affected the fate and functions of CAFs derived from dAT-MSCs. While CAFs derived from AT-MSCs of healthy donors (AT-CAFs) exhibited the markers of inflammatory CAFs, those derived from dAT-MSCs (dAT-CAFs) showed the markers of myofibroblastic CAFs. Of note, in comparison with AT-CAFs, dAT-CAFs showed a higher ability to induce the proliferation and in vivo metastasis of BCCs, which was involved in the activation of the transforming growth factor beta (TGFβ)-Smad2/3 signaling pathway. Collectively, our study suggests that T2DM contributes to metastasis of BCCs by inducing the myofibroblastic CAFs differentiation of dAT-MSCs. In addition, targeting the TGFβ-Smad2/3 signaling pathway in dAT-MSCs may be useful in cancer therapy for TNBC patients with T2DM.

Topics: Breast Neoplasms; Cancer-Associated Fibroblasts; Cell Line, Tumor; Diabetes Mellitus, Type 2; Female; Fibroblasts; Humans; Mesenchymal Stem Cells; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Bone-related events caused by breast cancer bone metastasis substantially compromise the survival and quality of life of patients. Because triple-negative breast cancer (TNBC) lacks hormone receptors and Her2-targeted therapeutic options, progress in the treatment of TNBC bone metastasis has been very slow. Intercellular adhesion molecule 1 (ICAM1) is highly expressed in various cancers and plays an important role in tumorigenesis and metastasis. However, the effect and mechanism of ICAM1 in TNBC bone metastasis are still unknown. We found that ICAM1 was highly expressed in TNBC and correlated with prognosis in TNBC patients. Cell lines with high expression of ICAM1 exhibited enhanced bone metastasis in tumor-bearing mice, and silencing ICAM1 expression significantly inhibited bone metastasis in mice. ICAM1 interacted with integrins to activate the epithelial-to-mesenchymal transition program through TGF-β/SMAD signaling, ultimately enhancing cell invasiveness. Therefore, the findings of the present study provide a strong rationale for the application of ICAM1-targeted therapy in TNBC patients with bone metastasis.

Topics: Animals; Cell Line, Tumor; Cell Movement; Cell Proliferation; Epithelial-Mesenchymal Transition; Humans; Integrins; Intercellular Adhesion Molecule-1; Mice; Neoplasm Metastasis; Quality of Life; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

The chemotherapy effect of docetaxel (DTX) against triple-negative breast cancer (TNBC) remains mediocre and limited when encapsulated in conventional cholesterol liposomes, mainly ascribed to poor penetration and immunosuppressive tumor microenvironment (TME) caused by tumor stroma cells, especially cancer-associated fibroblasts (CAFs). Many studies have attempted to address these problems but trapped into the common dilemma of excessively complicated formulation strategies at the expense of druggability as well as clinical translational feasibility. To better address the discrepancy, ginsenoside Rg3 was utilized to substitute cholesterol to develop a multifunctional DTX-loaded Rg3 liposome (Rg3-Lp/DTX). The obtained Rg3-Lp/DTX was proved to be preferentially uptake by 4T1 cells and accumulate more at tumor site via the interaction between the glycosyl moiety of Rg3 exposed on liposome surface and glucose transporter1 (Glut1) overexpressed on tumor cells. After reaching tumor site, Rg3 was shown to reverse the activated CAFs to the resting stage and attenuate the dense stroma barrier by suppressing secretion of TGF-β from tumor cells and regulating TGF-β/Smad signaling. Therefore, reduced levels of CAFs and collagens were found in TME after incorporation of Rg3, inducing enhanced penetration of Rg3-Lp/DTX in the tumor and reversed immune system which can detect and neutralize tumor cells. Compared with wooden cholesterol liposomes, the smart and versatile Rg3-Lp/DTX could significantly improve the anti-tumor effect of DTX, providing a promising approach for TNBC therapy with excellent therapeutic efficacy and simple preparation process.

Topics: Docetaxel; Ginsenosides; Glucose; Glucose Transporter Type 1; Humans; Liposomes; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Tumor Microenvironment

The transcription factor SNAI1 mediates epithelial-mesenchymal transition, fibroblast activation and controls inter-tissue migration. High SNAI1 expression characterizes metastatic triple-negative breast carcinomas, and its knockout by CRISPR/Cas9 uncovered an epithelio-mesenchymal phenotype accompanied by reduced signaling by the cytokine TGFβ. The SNAI1 knockout cells exhibited plasticity in differentiation, drifting towards the luminal phenotype, gained stemness potential and could differentiate into acinar mammospheres in 3D culture. Loss of SNAI1 de-repressed the transcription factor FOXA1, a pioneering factor of mammary luminal progenitors. FOXA1 induced a specific gene program, including the androgen receptor (AR). Inhibiting AR via a specific antagonist regenerated the basal phenotype and blocked acinar differentiation. Thus, loss of SNAI1 in the context of triple-negative breast carcinoma cells promotes an intermediary luminal progenitor phenotype that gains differentiation plasticity based on the dual transcriptional action of FOXA1 and AR. This function of SNAI1 provides means to separate cell invasiveness from progenitor cell de-differentiation as independent cellular programs.

Topics: Breast Neoplasms; Cell Line, Tumor; Cell Plasticity; Epithelial-Mesenchymal Transition; Female; Humans; Receptors, Androgen; Snail Family Transcription Factors; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Cyclic GMP-AMP synthase (cGAS) is an endogenous DNA sensor that synthesizes cyclic guanosine monophosphate-adenosine monophosphate (2'3'-cGAMP) from ATP and GTP. 2'3'-cGAMP activates the stimulator of interferon genes (STING) pathway, resulting in the production of interferons and pro-inflammatory cytokines. Ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1) is the phosphodiesterase that negatively regulates the STING pathway by hydrolyzing 2'3'-cGAMP. It has been established that the cGAS-STING pathway plays a major role in inhibiting tumor growth by upregulating T cell response. Herein, we demonstrate that AVA-NP-695, a selective and highly potent ENPP1 inhibitor, apart from the immunomodulatory effect also modulates cancer metastasis by negatively regulating epithelial-mesenchymal transition (EMT). We established that the combined addition of 2'3'-cGAMP and AVA-NP-695 significantly abrogated the transforming growth factor beta (TGF-ꞵ)-induced EMT in MDA-MB-231 cells. Finally, results from the in vivo study showed superior tumor growth inhibition and impact on tumor metastasis of AVA-NP-695 compared to Olaparib and PD-1 in a syngeneic 4T1 breast cancer mouse model. The translation of efficacy from in vitro to in vivo 4T1 tumor model provides a strong rationale for the therapeutic potential of AVA-NP-695 against triple-negative breast cancer (TNBC) as an immunomodulatory and anti-metastatic agent.

Topics: Adenosine Triphosphate; Animals; DNA; Guanosine Triphosphate; Humans; Interferons; Membrane Proteins; Mice; Nucleotidyltransferases; Phosphoric Diester Hydrolases; Programmed Cell Death 1 Receptor; Pyrophosphatases; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Agents blocking programmed cell death protein 1/programmed death-ligand 1 (PD-1/PD-L1) have been approved for triple-negative breast cancer (TNBC). However, the response rate of anti-PD-1/PD-L1 is still unsatisfactory, partly due to immunosuppressive factors such as transforming growth factor-beta (TGF-β). In our previous pilot study, the bispecific antibody targeting TGF-β and murine PD-L1 (termed YM101) showed potent antitumor effect. In this work, we constructed a bispecific antibody targeting TGF-β and human PD-L1 (termed BiTP) and explored the antitumor effect of BiTP in TNBC.. BiTP was developed using Check-BODY. BiTP exhibited high binding affinity to dual targets. In vitro experiments verified that BiTP effectively counteracted TGF-β-Smad and PD-L1-PD-1-NFAT signaling. In vivo animal experiments demonstrated that BiTP had superior antitumor activity relative to anti-PD-L1 and anti-TGF-β monotherapy. Mechanistically, BiTP decreased collagen deposition, enhanced CD8. BiTP retains parent antibodies' binding affinity and bioactivity, with superior antitumor activity to parent antibodies in TNBC. Our data suggest that BiTP might be a promising agent for TNBC treatment.

Topics: Animals; Antibodies, Bispecific; Antibodies, Neoplasm; Humans; Mice; Pilot Projects; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Tumor Microenvironment

Co-enzyme nicotinamide adenine dinucleotide NAD(H) regulates hundreds of biochemical reactions within the cell. We previously reported that NAD(H) redox status may have prognostic value for predicting breast cancer metastasis. However, the mechanisms of NAD(H) involvement in metastasis remain elusive. Given the important roles of TGFβ signalling in metastatic processes, such as promoting the epithelial-to-mesenchymal transition, we aimed to investigate the involvement of the mitochondrial NAD(H) redox status in TGFβ receptor signalling. Here we present the initial evidence that NAD(H) redox status is responsive to TGFβ receptor signalling in triple-negative breast cancer cells in culture. The mitochondrial NAD(H) redox status was determined by the optical redox imaging (ORI) technique. Cultured HCC1806 (less aggressive) and MDA-MB-231 (more aggressive) cells were subjected to ORI after treatment with exogenous TGFβ1 or LY2109761, which stimulates or inhibits TGFβ receptor signalling, respectively. Cell migration was determined with the transwell migration assay. Global averaging quantification of the ORI images showed that 1) TGFβ1 stimulation resulted in differential responses between HCC1806 and MDA-MB-231 lines, with HCC1806 cells having a significant change in the mitochondrial redox status, corresponding to a larger increase in cell migration; 2) HCC1806 cells acutely treated with LY2109761 yielded immediate increases in ORI signals. These preliminary data are the first evidence that suggests the existence of a cell line-dependent shift of the mitochondrial NAD(H) redox status in the TGFβ receptor signalling induced migratory process of breast cancer cells. Further research should be conducted to confirm these results as improved understanding of the underlying mechanisms of metastatic process may contribute to the identification of prognostic biomarkers and therapeutic targets.

Topics: Breast Neoplasms; Cell Line, Tumor; Epithelial-Mesenchymal Transition; Female; Humans; Mitochondria; NAD; Optical Imaging; Oxidation-Reduction; Receptors, Transforming Growth Factor beta; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Cyclin-dependent kinase 7 (CDK7) is a master regulatory kinase that drives cell cycle progression and stimulates expression of oncogenes in a myriad of cancers. Inhibitors of CDK7 (CDK7i) are currently in clinical trials; however, as with many cancer therapies, patients will most likely experience recurrent disease due to acquired resistance. Identifying targets underlying CDK7i resistance will facilitate prospective development of new therapies that can circumvent such resistance. Here we utilized triple-negative breast cancer as a model to discern mechanisms of resistance as it has been previously shown to be highly responsive to CDK7 inhibitors. After generating cell lines with acquired resistance, high-throughput RNA sequencing revealed significant upregulation of genes associated with efflux pumps and transforming growth factor-beta (TGF-β) signaling pathways. Genetic silencing or pharmacological inhibition of ABCG2, an efflux pump associated with multidrug resistance, resensitized resistant cells to CDK7i, indicating a reliance on these transporters. Expression of activin A (INHBA), a member of the TGF-β family of ligands, was also induced, whereas its intrinsic inhibitor, follistatin (FST), was repressed. In resistant cells, increased phosphorylation of SMAD3, a downstream mediator, confirmed an increase in activin signaling, and phosphorylated SMAD3 directly bound the ABCG2 promoter regulatory region. Finally, pharmacological inhibition of TGF-β/activin receptors or genetic silencing of SMAD4, a transcriptional partner of SMAD3, reversed the upregulation of ABCG2 in resistant cells and phenocopied ABCG2 inhibition. This study reveals that inhibiting the TGF-β/Activin-ABCG2 pathway is a potential avenue for preventing or overcoming resistance to CDK7 inhibitors.

Topics: ATP Binding Cassette Transporter, Subfamily G, Member 2; Cell Line, Tumor; Cyclin-Dependent Kinase-Activating Kinase; Cyclin-Dependent Kinases; Drug Resistance, Neoplasm; Female; Gene Expression Regulation, Neoplastic; Humans; Inhibin-beta Subunits; Neoplasm Proteins; Protein Kinase Inhibitors; Signal Transduction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Up-Regulation

Development of innovative therapeutic modalities would address an unmet clinical need in the treatment of triple negative breast cancer (TNBC). Activation of retinoic acid-inducible gene-I (RIG-I)-like receptors (RLRs) such as melanoma differentiation-associated gene 5 (MDA5) and RIG-I in cancer cells is suggested to suppress tumor progression by inducing cell death. Transfection of polyI:C, a conventionally used synthetic double-stranded RNA (dsRNA) analogue that activates RLRs, has been evaluated in clinical trials. However, detailed mechanisms of tumor suppression by RLRs, especially interactions with other signaling pathways, remain elusive. Here, we showed that transfection of polyI:C suppressed transforming growth factor-β (TGF-β) signaling in a MDA5- and RIG-I-dependent manner. We found that suppression of TGF-β signaling by polyI:C promoted cancer cell death, which was attenuated by forced expression of constitutively active Smad3. More detailed analysis suggested that cell death by polyI:C transfection exhibited characteristics of pyroptosis, which is distinct from apoptosis. Therapeutic efficacy of polyI:C transfection was also demonstrated using a mouse model. These results indicated that intratumor administration of polyI:C and related dsRNA analogues may be promising treatments for TNBC through inhibition of the anti-pyroptotic function of TGF-β.

Topics: Animals; Down-Regulation; Female; Humans; Mice; Mice, Inbred BALB C; Poly I-C; Pyroptosis; RNA, Double-Stranded; Signal Transduction; THP-1 Cells; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

Cancer immunotherapy shows limited efficacy against many solid tumors that originate from epithelial tissues, including triple-negative breast cancer (TNBC). We identify the SOX4 transcription factor as an important resistance mechanism to T cell-mediated cytotoxicity for TNBC cells. Mechanistic studies demonstrate that inactivation of SOX4 in tumor cells increases the expression of genes in a number of innate and adaptive immune pathways important for protective tumor immunity. Expression of SOX4 is regulated by the integrin αvβ6 receptor on the surface of tumor cells, which activates TGFβ from a latent precursor. An integrin αvβ6/8-blocking monoclonal antibody (mAb) inhibits SOX4 expression and sensitizes TNBC cells to cytotoxic T cells. This integrin mAb induces a substantial survival benefit in highly metastatic murine TNBC models poorly responsive to PD-1 blockade. Targeting of the integrin αvβ6-TGFβ-SOX4 pathway therefore provides therapeutic opportunities for TNBC and other highly aggressive human cancers of epithelial origin.

Topics: Animals; Antibodies, Monoclonal; Antigens, Neoplasm; Antineoplastic Agents, Immunological; Cell Line, Tumor; Drug Resistance, Neoplasm; Female; Gene Expression Regulation, Neoplastic; Humans; Integrins; Mice; Neoplasm Transplantation; Sequence Analysis, RNA; Signal Transduction; SOXC Transcription Factors; T-Lymphocytes, Cytotoxic; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Tumor Escape; Xenograft Model Antitumor Assays

Triple-negative breast cancer (TNBC) is the most aggressive histological subtype of breast cancer and is characterized by poor outcomes and a lack of specific-targeted therapies. Transforming growth factor-β (TGF-β) acts as the key cytokine in the epithelial-mesenchymal transition (EMT) and the metastasis of TNBC. However, the regulatory mechanisms of the TGF-β signaling pathway remain largely unknown. In this study, we identified that the USP1/WDR48 complex could effectively enhance TGF-β-mediated EMT and migration of TNBC cells. Furthermore, lower phosphorylation of Smad2/3, Erk, Jnk, and p38 was noted on the suppression of the expression of endogenous USP1 or WDR48. Moreover, the USP1-WDR48 complex was found to downregulate the polyubiquitination of TAK1 and mediate its in vitro stability. Therefore, our findings have shed a light on the novel role of the USP1/WDR48 complex in promoting TGF-β-induced EMT and migration in TNBC via in vitro stabilization of TAK1.

Topics: Cell Line, Tumor; Cell Movement; Epithelial-Mesenchymal Transition; Female; Humans; Intracellular Signaling Peptides and Proteins; MAP Kinase Kinase Kinases; Protein Stability; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Ubiquitin-Specific Proteases

Metastasis is the main cause of breast cancer mortality. Recent studies have proved that lipid metabolic reprogramming plays critical roles in breast cancer carcinogenesis and metastasis. We aim to identify critical lipid metabolism genes in breast cancer metastasis.. We designed and cloned a CRISPR pooled library containing lipid metabolic gene guide RNAs and performed a genetic screen in vivo. Transwell assay and animal experiments were used to evaluate cell metastatic ability in vitro or in vivo, respectively. We performed immunohistochemistry with breast cancer tissue microarray to study the clinical significance of NSDHL.. We identified a cholesterol metabolic enzyme, NSDHL, as a potential metastatic driver in triple-negative breast cancer. NSDHL was highly expressed in breast cancer tissues and predicted a poor prognosis. NSDHL knockdown significantly suppressed cell proliferation and migration. Mechanistically, NSDHL activated the TGFβ signaling pathway by inhibiting the endosomal degradation of TGFβR2. In addition, blocking the upstream metabolism of NSDHL with ketoconazole rescued cancer metastasis and TGFβR2 degradation. However, the inactivation of NSDHL (Y151X) did not rescue the migration ability and the TGFβR2 protein expression.. Taken together, our findings established that NSDHL serves as a metastatic driver, and its function depends on its enzyme activity in cholesterol biosynthesis and is mediated by the NSDHL-TGFβR2 signal pathway. Our study indicated that NSDHL and steroid biosynthesis may serve as new drug targets for patients with advanced breast cancer.

Topics: 3-Hydroxysteroid Dehydrogenases; Animals; Breast Neoplasms; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cholesterol; Female; Humans; Neoplasm Metastasis; Signal Transduction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Tumor associated neutrophils (TANs) are frequently detected in triple-negative breast cancer (TNBC). Recent studies also reveal the importance of neutrophils in promoting tumor progression and metastasis during breast cancer. However, the mechanisms regulating neutrophil trafficking to breast tumors are less clear. We sought to determine whether neutrophil trafficking to breast tumors is determined directly by the malignant potential of cancer cells. We found that tumor conditioned media (TCM) harvested from highly aggressive, metastatic TNBC cells induced a polarized morphology and robust neutrophil migration, while TCM derived from poorly aggressive estrogen receptor positive (ER+) breast cancer cells had no activity. In a three-dimensional (3D) type-I collagen matrix, neutrophils migrated toward TCM from aggressive breast cancer cells with increased velocity and directionality. Moreover, in a neutrophil-tumor spheroid co-culture system, neutrophils migrated with increased directionality towards spheroids generated from TNBC cells compared to ER+ cells. Based on these findings, we next sought to characterize the active factors secreted by TNBC cell lines. We found that TCM-induced neutrophil migration is dependent on tumor-derived chemokines, and screening TCM elution fractions based on their ability to induce polarized neutrophil morphology revealed the molecular weight of the active factors to be around 12 kDa. TCM from TNBC cell lines contained copious amounts of GRO (CXCL1/2/3) chemokines and TGF-β cytokines compared to ER+ cell-derived TCM. TCM activity was inhibited by simultaneously blocking receptors specific to GRO chemokines and TGF-β, while the activity remained intact in the presence of either single receptor inhibitor. Together, our findings establish a direct link between the malignant potential of breast cancer cells and their ability to induce neutrophil migration. Our study also uncovers a novel coordinated function of TGF-β and GRO chemokines responsible for guiding neutrophil trafficking to the breast tumor.

Topics: Cell Line; Cell Line, Tumor; Cell Movement; Cells, Cultured; Chemokines; Culture Media, Conditioned; Cytokines; Female; Humans; Ligands; MCF-7 Cells; Neutrophil Infiltration; Neutrophils; Receptors, Interleukin-8B; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Transforming growth factor beta (TGF-β) is the main cytokine responsible for the induction of the epithelial-mesenchymal transition of breast cancer cells, which is a hallmark of tumor transformation to the metastatic phenotype. Recently, research demonstrated that the chemokine CCL2 gene expression level directly correlates with the TGF-β activity in breast cancer patients. CCL2 attracts tumor-associated macrophages and is, therefore, considered as an important inductor of breast cancer progression; however, the precise mechanisms underlying its regulation by TGF-β are unknown. Here, we studied the behavior of the CCL2 gene in MDA-MB-231 and HCC1937 breast cancer cells representing mesenchymal-like phenotype activated by TGF-β. Using bioinformatics, deletion screening and point mutagenesis, we identified binding sites in the CCL2 promoter and candidate transcription factors responsible for its regulation by TGF-β. Among these factors, only the knock-down of EGR1 and RXRA made CCL2 promoter activity independent of TGF-β. These factors also demonstrated binding to the CCL2 promoter in a TGF-β-dependent manner in a chromatin immunoprecipitation assay, and point mutations in the EGR1 and RXRA binding sites totally abolished the effect of TGF-β. Our results highlight the key role of EGR1 and RXRA transcription factors in the regulation of CCL2 gene in response to TGF-β pathway.

Topics: Base Sequence; Binding Sites; Cell Line, Tumor; Chemokine CCL2; Early Growth Response Protein 1; Female; Gene Expression Regulation, Neoplastic; Humans; Models, Biological; Point Mutation; Promoter Regions, Genetic; Protein Binding; Retinoid X Receptor alpha; Signal Transduction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

TGF-β signaling pathway plays a key role in breast cancer metastasis. Recent studies suggest that TGF-β regulates tumor progression and invasion not only via transcriptional regulation, but also via translational regulation. Using both bioinformatics and experimental tools, we identified a micropeptide CIP2A-BP encoded by LINC00665, whose translation was downregulated by TGF-β in breast cancer cell lines. Using TNBC cell lines, we showed that TGF-β-activated Smad signaling pathway induced the expression of translation inhibitory protein 4E-BP1, which inhibited eukaryote translation initiation factor elF4E, leading to reduced translation of CIP2A-BP from LINC00665. CIP2A-BP directly binds tumor oncogene CIP2A to replace PP2A's B56γ subunit, thus releasing PP2A activity, which inhibits PI3K/AKT/NFκB pathway, resulting in decreased expression levels of MMP-2, MMP-9, and Snail. Downregulation of CIP2A-BP in TNBC patients was significantly associated with metastasis and poor overall survival. In the MMTV-PyMT model, either introducing CIP2A-BP gene or direct injection of CIP2A-BP micropeptide significantly reduced lung metastases and improved overall survival. In conclusion, we provide evidence that CIP2A-BP is both a prognostic marker and a novel therapeutic target for TNBC.

Topics: Adaptor Proteins, Signal Transducing; Animals; Apoptosis; Autoantigens; Biomarkers, Tumor; Cell Cycle Proteins; Cell Proliferation; Disease Progression; Female; Gene Expression Regulation, Neoplastic; Humans; Intracellular Signaling Peptides and Proteins; Membrane Proteins; Mice; Mice, Nude; Neoplasm Invasiveness; Peptide Fragments; Phosphatidylinositol 3-Kinases; Prognosis; Protein Phosphatase 2; Proto-Oncogene Proteins c-akt; RNA, Long Noncoding; Signal Transduction; Smad4 Protein; Survival Rate; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

Therapies directed to specific molecular targets are still unmet for patients with triple-negative breast cancer (TNBC). Deubiquitinases (DUB) are emerging drug targets. The identification of highly active DUBs in TNBC may lead to novel therapies.. Using DUB activity probes, we profiled global DUB activities in 52 breast cancer cell lines and 52 patients' tumor tissues. To validate our findings. Two DUB activity profiling approaches identified UCHL1 as being highly active in TNBC cell lines and aggressive tumors. Functionally, UCHL1 promoted metastasis in zebrafish and murine breast cancer xenograft models. Mechanistically, UCHL1 facilitates TGFβ signaling-induced metastasis by protecting TGFβ type I receptor and SMAD2 from ubiquitination. We found that these responses are potently suppressed by the specific UCHL1 inhibitor, 6RK73. Furthermore, UCHL1 levels were significantly increased in sera of patients with TNBC, and highly enriched in sera exosomes as well as TNBC cell-conditioned media. UCHL1-enriched exosomes stimulated breast cancer migration and extravasation, suggesting that UCHL1 may act in a paracrine manner to promote tumor progression.. Our DUB activity profiling identified UCHL1 as a candidate oncoprotein that promotes TGFβ-induced breast cancer metastasis and may provide a potential target for TNBC treatment.

Topics: Animals; Biomarkers, Tumor; Cell Line, Tumor; Cell Proliferation; Deubiquitinating Enzymes; Female; Humans; Mice; Mice, Inbred BALB C; Mice, Nude; Neoplasm Metastasis; Oncogene Proteins; Signal Transduction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Ubiquitin Thiolesterase; Xenograft Model Antitumor Assays; Zebrafish

Breast cancer (BC) is one of the most ordinary fatal cancers. Recent studies have identified the vital role of genes in the development and progression of Tri-negative breast cancer (TNBC). In this research, DGCR8 was studied to identify how it functioned in the metastasis of TNBC.. DGCR8 expression of tissues was detected by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) in 50 TNBC patients. Wound healing assay and transwell assay were used to observe the changes in the biological behaviors of TNBC cells through knockdown or overexpression of DGCR8. In addition, qRT-PCR and Western blot assay were performed to discover the potential target protein of DGCR8 in TNBC.. DGCR8 expression level in TNBC samples was higher than that of adjacent ones. Besides, the migration ability and invasion ability of TNBC cells were inhibited after DGCR8 was silenced, while they were promoted after DGCR8 was overexpressed. In addition, TGF-β was downregulated after silencing of DGCR8 in TNBC cells, while TGF-β was upregulated after overexpression of DGCR8 in TNBC cells. Furthermore, TGF-β was upregulated in TNBC tissues, which was positively associated with DGCR8.. Our study uncovers a new oncogene in TNBC and suggests that DGCR8 can enhance TNBC cell migration and invasion via targeting TGF-β, which provides a novel therapeutic target for TNBC patients.

Topics: Cells, Cultured; Epigenesis, Genetic; Humans; RNA-Binding Proteins; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Tumor progression and metastasis are the major causes of death among cancer associated mortality. Metastatic cells acquire features of migration and invasion and usually undergo epithelia-mesenchymal transition (EMT). Acquirement of these various hallmarks rely on different cellular pathways, including TGF-β and Wnt signaling. Recently, we reported that WW domain-containing oxidoreductase (WWOX) acts as a tumor suppressor and has anti-metastatic activities involving regulation of several key microRNAs (miRNAs) in triple-negative breast cancer (TNBC). Here, we report that WWOX restoration in highly metastatic MDA-MB435S cancer cells alters mRNA expression profiles; further, WWOX interacts with various proteins to exert its tumor suppressor function. Careful alignment and analysis of gene and miRNA expression in these cells revealed profound changes in cellular pathways mediating adhesion, invasion and motility. We further demonstrate that WWOX, through regulation of miR-146a levels, regulates SMAD3, which is a member of the TGF-β signaling pathway. Moreover, proteomic analysis of WWOX partners revealed regulation of the Wnt-signaling activation through physical interaction with Disheveled. Altogether, these findings underscore a significant role for WWOX in antagonizing metastasis, further highlighting its role and therapeutic potential in suppressing tumor progression.

Topics: Cell Movement; Cell Proliferation; Dishevelled Proteins; Epithelial-Mesenchymal Transition; Female; Gene Expression Regulation, Neoplastic; Genetic Pleiotropy; Humans; MicroRNAs; Neoplasm Metastasis; Protein Interaction Maps; Signal Transduction; Smad3 Protein; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Tumor Suppressor Proteins; Wnt Signaling Pathway; WW Domain-Containing Oxidoreductase

Topics: 5'-Nucleotidase; Adenosine; Adult; Aged; Animals; Breast; Cancer-Associated Fibroblasts; Carcinogenesis; Datasets as Topic; Extracellular Matrix; Female; Gene Expression Profiling; Humans; Kaplan-Meier Estimate; Mammary Glands, Animal; Mammary Neoplasms, Experimental; Mice; Mice, Transgenic; Middle Aged; Myeloid Cells; Receptor, Adenosine A2B; Signal Transduction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Altered signaling pathways typify breast cancer and serve as direct inputs to steroid hormone receptor sensors. We previously reported that phospho-Ser134-GR (pS134-GR) species are elevated in triple-negative breast cancer (TNBC) and cooperate with hypoxia-inducible factors, providing a novel avenue for activation of GR in response to local or cellular stress.. We probed GR regulation by factors (cytokines, growth factors) that are rich within the tumor microenvironment (TME). TNBC cells harboring endogenous wild-type (wt) or S134A-GR species were created by CRISPR/Cas knock-in and subjected to transwell migration, invasion, soft-agar colony formation, and tumorsphere assays. RNA-seq was employed to identify pS134-GR target genes that are regulated both basally (intrinsic) or by TGFβ1 in the absence of exogenously added GR ligands. Regulation of selected basal and TGFβ1-induced pS134-GR target genes was validated by qRT-PCR and chromatin immunoprecipitation assays. Bioinformatics tools were used to probe public data sets for expression of pS134-GR 24-gene signatures.. In the absence of GR ligands, GR is transcriptionally activated via p38-dependent phosphorylation of Ser134 as a mechanism of homeostatic stress-sensing and regulated upon exposure of TNBC cells to TME-derived agents. The ligand-independent pS134-GR transcriptome encompasses TGFβ1 and MAPK signaling gene sets associated with TNBC cell survival and migration/invasion. Accordingly, pS134-GR was essential for TNBC cell anchorage-independent growth in soft-agar, migration, invasion, and tumorsphere formation, an in vitro readout of cancer stemness properties. Both pS134-GR and expression of the MAPK-scaffolding molecule 14-3-3ζ were essential for a functionally intact p38 MAPK signaling pathway downstream of MAP3K5/ASK1, indicative of a feedforward signaling loop wherein self-perpetuated GR phosphorylation enables cancer cell autonomy. A 24-gene pS134-GR-dependent signature induced by TGFβ1 predicts shortened overall survival in breast cancer patients.. Phospho-S134-GR is a critical downstream effector of p38 MAPK signaling and TNBC migration/invasion, survival, and stemness properties. Our studies define a ligand-independent role for GR as a homeostatic "sensor" of intrinsic stimuli as well as extrinsic factors rich within the TME (TGFβ1) that enable potent activation of the p38 MAPK stress-sensing pathway and nominate pS134-GR as a therapeutic target in aggressive TNBC.

Topics: Biomarkers, Tumor; Cell Line, Tumor; Cell Movement; Extracellular Matrix Proteins; Female; Gene Editing; Humans; Neoplasm Staging; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Receptors, Glucocorticoid; Transcriptome; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Tumor Microenvironment

Cancer/testis (CT) antigens are proteins whose expression is normally restricted to germ cells yet aberrantly activated in tumors, where their functions remain relatively cryptic. Here we report that ZNF165, a CT antigen frequently expressed in triple-negative breast cancer (TNBC), associates with SMAD3 to modulate transcription of transforming growth factor β (TGFβ)-dependent genes and thereby promote growth and survival of human TNBC cells. In addition, we identify the KRAB zinc finger protein, ZNF446, and its associated tripartite motif protein, TRIM27, as obligate components of the ZNF165-SMAD3 complex that also support tumor cell viability. Importantly, we find that TRIM27 alone is necessary for ZNF165 transcriptional activity and is required for TNBC tumor growth in vivo using an orthotopic xenograft model in immunocompromised mice. Our findings indicate that aberrant expression of a testis-specific transcription factor is sufficient to co-opt somatic transcriptional machinery to drive a pro-tumorigenic gene expression program in TNBC.

Topics: Animals; Cell Line, Tumor; DNA-Binding Proteins; Female; Gene Expression Regulation, Neoplastic; Humans; Male; Mice; Signal Transduction; Smad3 Protein; Testis; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

The tumour microenvironment is rich in multiple cells that influence cancer development. Among them, macrophages are the most abundant immune cells, which secrete factors involved in carcinogenesis. Since protein-bound polysaccharides (PBP) from the. 4T1 cells were cultured in conditioned media (CM) collected after: stimulation of the macrophages with PBP (CM-PBP) or incubation of non-treated macrophages (CM-NT). A co-cultured model of both cell lines was also employed to investigate the crosstalk between the cells. Cell viability was measured using the MTT assay. The levels of cytokines and chemokines were determined by ELISA methods. Commercial assay kits were used to assess the activity of both arginase 1 and inducible nitric oxide synthase (iNOS) and the level of cell migration.. The results revealed that CM-NT promotes proliferation and migration of 4T1 cells, and increases the secretion of pro-angiogenic factors (VEGF, MCP-1) by cancer cells. In contrast, CM-PBP inhibits 4T1 cell growth and migration, decreases the secretion of pro-angiogenic factors (VEGF, MCP-1) and upregulates the production of pro-inflammatory mediators (IL-6, TNF-α) with certain anti-tumoral properties Moreover, PBP-treated CM significantly decreases the level of M2 macrophage markers (arginase 1 activity, IL-10 and TGF-β concentrations), but upregulates iNOS activity and IL-6 and TNF-α production, which are M1 cell markers.. The results suggest that PBP suppress the favourable tumour microenvironment by inhibiting the crosstalk between 4T1 cells and macrophages through the regulation of production of angiogenic and inflammatory mediators, and modulating the M1/M2 macrophage subtype.

Topics: Animals; Arginase; Cell Movement; Cell Proliferation; Cell Survival; Chemokine CCL2; Chemokines; Coculture Techniques; Culture Media, Conditioned; Inflammation; Interleukin-10; Interleukin-6; Mice; Neovascularization, Pathologic; Nitric Oxide Synthase Type II; Polyporaceae; Polysaccharides; RAW 264.7 Cells; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Tumor Microenvironment; Tumor Necrosis Factor-alpha; Tumor-Associated Macrophages; Vascular Endothelial Growth Factor A

Triple-Negative Breast Cancer (TNBC) is an aggressive and complex subtype of breast cancer. The current biomarkers used in the context of breast cancer treatment are highly dependent on the targeting of oestrogen receptor, progesterone receptor, or HER2, resulting in treatment failure and disease recurrence and creating clinical challenges. Thus, there is still a crucial need for the improvement of TNBC treatment; the discovery of effective biomarkers that can be easily translated to the clinics is essential.. We report an approach for the discovery of biomarkers that can predict tumour relapse and pathologic complete response (pCR) in TNBC on the basis of mRNA expression quantified using the NanoString nCounter Immunology Panel. To overcome the limited sample size, prediction models based on random Forest were constructed using the differentially expressed genes (DEGs) as selected features. We also evaluated the differences between pre- and post-treatment groups aiming for the combinatorial assessment of pCR and relapse using additive models in edgeR.. We identify nine and 13 DEGs strongly associated with pCR and relapse, respectively, from 579 immune genes in a small number of samples (n = 55) using edgeR. An additive model for the comparison of pre- and post-treatment groups via the adjustment of the independent subject in the relapse group revealed associations for 41 genes. Comprehensive analysis indicated that our prediction models outperformed those constructed using features extracted from the existing feature selection model Elastic Net in terms of accuracy. The prediction models were assessed using a randomization test to validate the robustness (empirical P for the model of pCR = 0.015 and empirical P for the model of relapse = 0.018). Furthermore, three DEGs (FCER1A, EDNRB, and TGFBI) in the model of relapse showed prognostic significance for predicting the survival of patients with cancer through Cox proportional hazards regression model-based survival analysis.. Gene expression quantified via the NanoString nCounter Immunology Panel can be seamlessly analysed using edgeR, even considering small sample sizes. Our approach provides a scalable framework that can easily be applied for the discovery of biomarkers based on the NanoString nCounter Immunology Panel.. The source code will be available from github at https://github.com/sungheep/nanostring .

Topics: Biomarkers, Tumor; Extracellular Matrix Proteins; Female; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Humans; Immunity; Models, Genetic; Prognosis; Proportional Hazards Models; Receptor, Endothelin B; Receptors, IgE; Survival Analysis; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

The transforming growth factor β (TGFβ) and bone morphogenetic protein (BMP) signaling pathways are both constitutively activated in triple-negative breast cancer (TNBC). We are interested in isolating the naturally-derived small-molecule inhibitor that could simultaneously targeting TGFβ/BMP pathways and further studying its anti-proliferative/-metastatic effects as well as the underlying mechanisms in multiple tumor models.. Multiple in vitro cell-based assays are used to examine the compound's inhibitory efficacy on TNBC cell growth, stemness, epithelial-mesenchymal transition (EMT), invasion and migration by targeting TGFβ/BMP signaling pathways. Transgenic breast cancer mouse model (MMTV-PyMT), subcutaneous xenograft and bone metastasis models are used to examine ZL170's effects on TNBC growth and metastasis potentials in vivo.. ZL170 dose-dependently inhibits cell proliferation, EMT, stemness, invasion and migration in vitro via specifically targeting canonical TGFβ/BMP-SMADs pathways in TNBC cells. The compound significantly hinders osteolytic bone metastasis and xenograft tumor growth without inflicting toxicity on vital organs of tumor-bearing nude mice. ZL170 strongly inhibits primary tumor growth and lung metastases in MMTV-PyMT transgenic mice. ZL170-treated tumors exhibit impaired TGFβ/BMP signaling pathways in both epithelial and stromal compartments, thereby creating a suppressive tumor microenvironment characterized by reduced extracellular matrix deposition and decreased infiltration of stromal cells.. ZL170 inhibits tumor EMT, stemness and metastasis and could be further developed as a potent anti-metastatic agent used in combination with cytotoxic drugs for treatment of TNBC and other advanced metastatic cancers.

Topics: Animals; Bone Morphogenetic Proteins; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cell Survival; Epithelial-Mesenchymal Transition; Female; Humans; Mice; Mice, Nude; Mice, Transgenic; Neoplastic Stem Cells; Oxindoles; Signal Transduction; Small Molecule Libraries; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Xenograft Model Antitumor Assays

The invasive and metastatic phenotypes of breast cancer correlate with high recurrence rates and poor survival outcomes. Transforming growth factor-β (TGFβ) promotes tumor progression and metastasis in aggressive breast cancer. Here, we identified the kisspeptin KiSS1 as a downstream target of canonical TGFβ/Smad2 pathway in triple negative breast cancer cells. We also found KiSS1 expression to be required for TGFβ-induced cancer cell invasion. Indeed, knockdown expression of KiSS1 blocked TGFβ-mediated cancer cell invasion as well as metalloproteinase (MMP9) expression and activity. Interestingly, Kisspeptin-10 (KP-10), the smallest active form of kisspeptin also stimulates cancer cell invasive behavior through activation of MAPK/Erk pathway. We described a positive feedback loop between KiSS1 and p21 downstream of TGFβ, further contributing to TGFβ-induced cancer cell invasion. Lastly, we explored both the clinical utility of KiSS1 as a lymph node involvement predictive tool and its potential as a therapeutic target. We found KiSS1 high expression to correlate with lymph node positive status. Furthermore, blocking KiSS1 using a specific small peptide antagonist (p234) impaired TGFβ-mediated cell invasion and MMP9 induction. Together, our results define an essential role of KiSS1 in regulating TGFβ pro-invasive effects and define KiSS1 as a therapeutic new target for triple negative breast cancer.

Topics: Adenocarcinoma; Antineoplastic Agents; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin-Dependent Kinase Inhibitor p21; Extracellular Signal-Regulated MAP Kinases; Feedback, Physiological; Female; Gene Expression Regulation, Neoplastic; Humans; Kisspeptins; Lymphatic Metastasis; Matrix Metalloproteinase 9; MCF-7 Cells; Mitogen-Activated Protein Kinases; Protein Isoforms; Signal Transduction; Smad2 Protein; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Topics: Antibiotics, Antineoplastic; Apoptosis; Cell Line, Tumor; Drug Resistance, Neoplasm; Epirubicin; Epithelial-Mesenchymal Transition; Female; Humans; Neoplastic Stem Cells; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Cadmium (Cd) has been linked to a variety of cancers, including breast cancer; however, the molecular mechanism of its carcinogenic activity is not fully understood. To this end, the present study investigated the roles of ferroportin (FPN), a prognostic marker of breast cancer, in Cd-induced stimulation of cell proliferation and cell migration. Triple-negative MDA-MB-231 cells were treated with 1-3 μM Cd. The cells exhibited significant reduction in FPN expression and concomitant increase in iron concentration. Cells treated with Cd for 8 weeks displayed elevated proliferative and migratory activities which were inversely related with FPN expression. Reduced FPN expression also resulted in EMT as indicated by an increase in the expression of E-cadherin, and a decrease in the expression of N-cadherin, Twist and Slug. Further investigation revealed that Cd suppressed FPN expression at least partially by activating TGF-β, a known regulator of FPN expression. Taken together, these results indicate that Cd-induced stimulation of MDA-MB-231 cell proliferation, EMT, and migration is brought about by suppression of FPN expression and associated disruption of iron homeostasis.

Topics: Antigens, CD; Cadherins; Cadmium; Cation Transport Proteins; Cell Line, Tumor; Cell Movement; Cell Proliferation; Epithelial-Mesenchymal Transition; Female; Gene Expression Regulation, Neoplastic; Homeostasis; Humans; Iron; RNA, Small Interfering; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Triple-negative breast cancer (TNBC) is often aggressive and metastatic. Transforming growth factor-β acts as a tumor-promoter in TNBC. Smad3, a major downstream effector protein in the TGF-β signaling pathway, is regulated by phosphorylation at several sites. The functional significance of the phosphorylation of the linker region in Smad3 is poorly understood for TNBC. Among the four sites in the Smad3 linker region, threonine-179 (T179) appears to be unique as it serves as the binding site for multiple WW-domain-containing proteins upon phosphorylation, suggesting that this phosphorylation is a key for Smad3 to engage other pathways. Using genome editing, we introduced for the first time a knock-in (KI) mutation in the endogenous Smad3 gene in IV2, a lung-tropic subline of the human MDA-MB-231 TNBC cell line. In the resulting cell line, the Smad3 T179 phosphorylation site is replaced by non-phosphorylatable valine (T179V) with the mutation in both alleles. The T179V KI reduced cell growth rate and mammosphere formation. These phenomena were accompanied by a significant upregulation of p21

Topics: Animals; Binding Sites; Cell Line, Tumor; Cell Proliferation; Female; Gene Editing; Gene Knock-In Techniques; Humans; Lung Neoplasms; Mice; Mice, SCID; Phosphorylation; Point Mutation; Signal Transduction; Smad3 Protein; Threonine; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Valine; Xenograft Model Antitumor Assays

Triple-negative breast cancer (TNBC) accounts for a disproportionately high number of deaths due to a lack of targeted therapies and an increased likelihood of distant recurrence. Estrogen receptor beta (ERβ), a well-characterized tumor suppressor, is expressed in 30% of TNBCs, and its expression is associated with improved patient outcomes. We demonstrate that therapeutic activation of ERβ elicits potent anticancer effects in TNBC through the induction of a family of secreted proteins known as the cystatins, which function to inhibit canonical TGFβ signaling and suppress metastatic phenotypes both in vitro and in vivo. These data reveal the involvement of cystatins in suppressing breast cancer progression and highlight the value of ERβ-targeted therapies for the treatment of TNBC patients.

Topics: Animals; Cell Line, Tumor; Cystatins; Estrogen Receptor beta; Female; Humans; Mice; Signal Transduction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Tumor Suppressor Proteins

In the era of immunotherapy and personalized medicine, there is an urgent need for advancing the knowledge of immune evasion in different cancer types and identifying reliable biomarkers that guide both therapy selection and patient inclusion in clinical trials. Given the differential immune responses and evasion mechanisms in breast cancer, we expect to identify different breast cancer groups based on their expression of immune-related genes. For that, we used the sequential biclustering method on The Cancer Genome Atlas RNA-seq breast cancer data and identified 7 clusters. We found that 77.4% of the clustered tumor specimens evade through transforming growth factor-beta (TGF-β) immunosuppression, 57.7% through decoy receptor 3 (DcR3) counterattack, 48.0% through cytotoxic T-lymphocyte-associated protein 4 (CTLA4), and 34.3% through programmed cell death-1 (PD-1). TGF-β and DcR3 are potential novel drug targets for breast cancer immunotherapy. Targeting TGF-β and DcR3 may provide a powerful approach for treating breast cancer because 57.7% of patients overexpressed these two molecules. Furthermore, triple-negative breast cancer (TNBC) patients clustered equally into two subgroups: one with impaired antigen presentation and another with high leukocyte recruitment but four different evasion mechanisms. Thus, different TNBC patients may be treated with different immunotherapy approaches. We identified biomarkers to cluster patients into subgroups based on immune evasion mechanisms and guide the choice of immunotherapy. These findings provide a better understanding of patients' response to immunotherapies and shed light on the rational design of novel combination therapies.

Topics: B7-H1 Antigen; Biomarkers, Tumor; Breast Neoplasms; CTLA-4 Antigen; Female; Gene Expression; Humans; Immune Evasion; Immunotherapy; Programmed Cell Death 1 Receptor; Receptors, Tumor Necrosis Factor, Member 6b; Sequence Analysis, RNA; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Glycopeptide-preferring polypeptide N-acetylgalactosamine transferase (ppGalNAc‑T) is a key enzyme that initiates the formation of the first GalNAc monosaccharide to polypeptides at Thr/Ser residues by O‑linked glycosylation. In order to investigate the effects of ppGalNAc‑T1 and ppGalNAc‑T2 on the initiation of O‑glycosylation, siRNA‑ppGalNAc‑T1 (si‑T1) and siRNA‑ppGalNAc‑T2 (si‑T2) were transfected into highly‑invasive estrogen receptor‑negative MDA‑MB‑231 cells to inhibit O‑glycosylation. Downregulation of ppGalNAc‑T1 demonstrated a significant reduction in the number of terminal α2,3 sialic acids, when compared to cells transfected with si‑T2 or si‑T1/T2. This downregulation led to a decrease in the invasion capabilities of the breast carcinoma cells, as well as enhanced chemosensitivity, which was the result antineoplastic drug effects. In addition, immunoprecipitation assays demonstrated that downregulation of ppGalNAc‑T1 led to a reduction in the number of terminal α2,3 sialic acids on O‑linked glycans of the matrix metalloproteinase‑14 (MMP14) glycoprotein. Furthermore, MMP14 and vascular endothelial growth factor were downregulated in the si‑T1 groups when compared with the si‑T2 and si‑T1/T2 groups. In conclusion, the results of the present study suggest that ppGalNAc‑T1 may serve a pivotal role in the initiation of O‑glycosylation, which may lead to a low density of α2,3 sialic acids on O‑linked glycans of MMP14 when downregulated. Glycosylation serves a significant role in regulating the sensitivity of MMP14 to self‑proteolysis, which ultimately decreases the invasion capabilities of breast cancer cells. The results of the present study may be useful in establishing the function of ppGalNAc‑T1 during breast cancer invasion and metastasis.

Topics: Antineoplastic Agents; Cell Movement; Cell Proliferation; Drug Resistance, Neoplasm; Gene Expression Regulation, Neoplastic; Gene Silencing; Glycosylation; Humans; Matrix Metalloproteinase 14; N-Acetylgalactosaminyltransferases; Polypeptide N-acetylgalactosaminyltransferase; RNA Interference; RNA, Small Interfering; Sialic Acids; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Vascular Endothelial Growth Factor A

Transforming growth factor β (TGF-β)-induced migration of triple-negative breast cancer (TNBC) cells is dependent on nuclear export of the orphan receptor NR4A1, which plays a role in proteasome-dependent degradation of SMAD7. In this study, we show that TGF-β induces p38α (mitogen-activated protein kinase 14 [MAPK14]), which in turn phosphorylates NR4A1, resulting in nuclear export of the receptor. TGF-β/p38α and NR4A1 also play essential roles in the induction of epithelial-to-mesenchymal transition (EMT) and induction of β-catenin in TNBC cells, and these TGF-β-induced responses and nuclear export of NR4A1 are blocked by NR4A1 antagonists, the p38 inhibitor SB202190, and kinase-dead [p38(KD)] and dominant-negative [p38(DN)] forms of p38α. Inhibition of NR4A1 nuclear export results in nuclear export of TGF-β-induced β-catenin, which then undergoes proteasome-dependent degradation. TGF-β-induced β-catenin also regulates NR4A1 expression through formation of the β-catenin-TCF-3/TCF-4/LEF-1 complex on the NR4A1 promoter. Thus, TGF-β-induced nuclear export of NR4A1 in TNBC cells plays an essential role in cell migration, SMAD7 degradation, EMT, and induction of β-catenin, and all of these pathways are inhibited by bis-indole-derived NR4A1 antagonists that inhibit nuclear export of the receptor and thereby block TGF-β-induced migration and EMT.

Topics: Active Transport, Cell Nucleus; Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; beta Catenin; Cell Line, Tumor; Cell Movement; Epithelial-Mesenchymal Transition; Female; Humans; Imidazoles; Lymphoid Enhancer-Binding Factor 1; Mitogen-Activated Protein Kinase 14; Multiprotein Complexes; Nuclear Receptor Subfamily 4, Group A, Member 1; Phosphorylation; Pyridines; RNA Interference; RNA, Small Interfering; Smad7 Protein; Transcription Factor 4; Transcription Factor 7-Like 1 Protein; Transcription Factors; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Triple negative breast cancer (TNBC) is a highly aggressive breast cancer subtype that lacks effective targeted therapies. Although TNBC is not defined by specific therapeutic targets, a subset of patients have tumors that overexpress cyclins. High cyclin D/E expression catalyzes CDK4/2 activity. In turn, CDK4/2 can non-canonically phosphorylate Smad3, a key TGFβ signaling intermediate, and this phosphorylation has been associated with the shift from tumor-suppressive to oncogenic TGFβ pathway action in breast oncogenesis. Additionally, CDK-mediated Smad3 phosphorylation facilitates an interaction between Smad3 and Pin1, a cis-trans isomerase that is also overexpressed in aggressive breast cancers. Treatment with CYC065, a CDK2/9 inhibitor, decreased non-canonical Smad3 phosphorylation and inhibited the Pin1-Smad3 interaction. We hypothesized that the interaction of Pin1 and Smad3, facilitated by CDK-mediated Smad3 phosphorylation, promotes TNBC cell aggressiveness. Inhibition of the Pin1-Smad3 interaction in TNBC cell lines, through depletion of Pin1 or CYC065 treatment, resulted in decreased cell migration/invasion and impeded the EMT program. Inhibition of CDK-mediated phosphorylation of Smad3 by mutagenesis also decreased cell migration, underscoring the importance of non-canonical CDK2 phosphorylation of Smad3 to enable cell motility. Pin1 depletion restored Smad3 protein levels and tumor-suppressive activity, suggesting that the Pin1-Smad3 interaction has a negative impact on canonical Smad3 action. Collectively, the data show that the Pin1-Smad3 interaction, facilitated by CDK-mediated Smad3 phosphorylation, is associated with oncogenic TGFβ signaling and breast cancer progression. Inhibition of this interaction with CYC065 treatment may provide an important therapeutic option for TNBC patients.

Topics: Adenosine; Cell Line, Tumor; Cell Movement; Cyclin-Dependent Kinase 2; Cyclin-Dependent Kinase 4; Female; Humans; NIMA-Interacting Peptidylprolyl Isomerase; Phosphorylation; Protein Binding; Signal Transduction; Smad3 Protein; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Triple negative breast cancer (TNBC), an aggressive subtype of breast cancer, display poor prognosis and exhibit resistance to conventional therapies, partly due to an enrichment in breast cancer stem cells (BCSCs). Here, we investigated the role of the cyclooxygenase-2 (COX-2), a downstream target of TGFβ, in regulating BCSCs in TNBC. Bioinformatics analysis revealed that COX-2 is highly expressed in TNBC and that its expression correlated with poor survival outcome in basal subtype of breast cancer. We also found TGFβ-mediated COX-2 expression to be Smad3-dependent and to be required for BCSC self-renewal and expansion in TNBCs. Knocking down COX-2 expression strikingly blocked TGFβ-induced tumorsphere formation and TGFβ-induced enrichment of the two stem-like cell populations, CD24

Topics: Biomarkers, Tumor; CD24 Antigen; Cell Line, Tumor; Cyclooxygenase 2; Female; Fibronectins; Gene Expression Regulation, Neoplastic; Humans; Hyaluronan Receptors; Kaplan-Meier Estimate; Neoplastic Stem Cells; Smad3 Protein; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

"Triple negative breast cancer" (TNBC) is associated with a higher rate and earlier time of recurrence and worse prognosis after recurrence. In this study, we aimed to examine the crosstalk between fibroblasts and TNBC cells. The fibroblasts were isolated from TNBC patients' tissue in tumor burden zones, distal normal zones and interface zones. The fibroblasts were indicated as cancer-associated fibroblasts (CAFs), normal zone fibroblasts (NFs) and interface zone fibroblasts (INFs). Our study found that INFs grew significantly faster than NFs and CAFs in vitro. The epithelial BT20 cells cultured with the conditioned medium of INFs (INFs-CM) and CAFs (CAFs-CM) showed more spindle-like shape and cell scattering than cultured with the conditioned medium of NFs (NFs-CM). These results indicated that factors secreted by INFs-CM or CAFs-CM could induce the epithelial-mesenchymal transition (EMT) phenotype in BT20 cells. Using an in vitro co-culture model, INFs or CAFs induced EMT and promoted cancer cell migration in BT20 cells. Interestingly, we found that emodin inhibited INFs-CM or CAFs-CM-induced EMT programming and phenotype in BT20 cells. Previous studies reported that CAFs and INFs-secreted TGF-β promoted human breast cancer cell proliferation, here; our results indicated that TGF-β initiated EMT in BT20 cells. Pretreatment with emodin significantly suppressed the TGF-β-induced EMT and cell migration in BT20 cells. These results suggest that emodin may be used as a novel agent for the treatment of TNBC.

Topics: Cancer-Associated Fibroblasts; Cell Line, Tumor; Emodin; Epithelial-Mesenchymal Transition; Female; Fibroblasts; Humans; Phenotype; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Breast cancer is one of the most invasive cancer types in female population. The functional activity of Transforming growth factor β-activated kinase 1 (TAK1) in breast cancer progression increasingly attracts attention as it provides a potential target for antibreast cancer drug development. However, the fundamental role of TAK1 for triple-negative breast cancer (TNBC) progression and the effect of potential anti-TAK1 drug candidate needs to be further evaluated. Herein, we focused on the role of TAK1 in human breast cancer cells, and we hypothesized that the inhibition of TAK1 activation can repress the growth of human TNBC cells. We found that the TAK1 is robustly activated within cancer cell population of clinic-derived TNBC samples and the human breast cancer cell lines in culture. Furthermore, we determined the effect of 5Z-7-oxozeaenol (5Z-O), a TAK1-specific small molecule inhibitor, on proliferation of human TNBC cell line. 5Z-O treatment significantly suppressed the proliferation of human TNBC cells. Collectively, these demonstrate the role of TAK1 in human breast cancer and the antiproliferate effect of TAK1 inhibitor. Our study sets the stage for further research on TAK1 as a promising target for development of anti-TNBC drugs and therapeutic strategies.

Topics: Antineoplastic Agents; Cell Line, Tumor; Cell Proliferation; ErbB Receptors; Female; Humans; MAP Kinase Kinase Kinases; Middle Aged; Phosphorylation; Protein Kinase Inhibitors; Signal Transduction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Zearalenone

Triple-negative breast cancer (TNBC) is a highly aggressive tumor subtype lacking effective prognostic indicators or therapeutic targets. Mitochondrial function is dysregulated frequently in cancer cells to allow for adaptation to a harsh tumor microenvironment. Targeting mitochondrial biogenesis and bioenergetics is, therefore, an attractive therapeutic strategy. In this study, we performed quantitative proteomic analyses in human parental and metastatic breast cancer cell lines to identify mitochondrial proteins involved in TNBC metastasis. We found that single-strand DNA-binding protein 1 (SSBP1) was downregulated in highly metastatic breast cancer cells. Moreover, SSBP1 downregulation promoted TNBC cell metastasis in vitro and in vivo. Mechanistically, SSBP1 loss decreased mitochondrial DNA copy number, thereby potentiating calcineurin-mediated mitochondrial retrograde signaling that induced c-Rel/p50 nuclear localization, activated TGFβ promoter activity, and TGFβ-driven epithelial-to-mesenchymal transition. Low SSBP1 expression correlated with tumor progression and poor prognosis in patients. Collectively, our findings identified SSBP1 as a novel metastasis suppressor and elucidated the mechanisms by which dysregulated mitochondrial signaling contributes to metastatic potential, providing potential new prognostic indicators for patients with TNBC.

Topics: Cell Line, Tumor; Cell Proliferation; DNA-Binding Proteins; Epithelial-Mesenchymal Transition; Female; Humans; Mitochondria; Mitochondrial Proteins; Neoplasm Metastasis; Signal Transduction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Triple negative breast cancer is exceptionally difficult to treat due to the lack of distinguishing biomarkers for drug targeting. An alternative approach based on recent data indicates that these cells may be more susceptible to mechanical influences, such as alterations in the tumor stroma. Three dimensional collagen gels containing co-cultures of mesenchymal cells and MDA-MB-231 cancer cells were utilized to explore the effects of multi-walled nanotubes (MWNT) on cell contraction, invasion, viability, MMP-9 expression, and migration of breast cancer cells. MWNT were able to restrict each of these features for the cancer cells without impeding the associated mesenchymal cells. MWNT-collagen gels are useful tools for cellular reprogramming of cancer cells and should be considered in greater detail as a potential agent for therapeutic treatment of triple-negative breast cancer.. Breast cancer is still a leading cause of death for women worldwide. One subtype of this cancer which is very aggressive is the triple negative breast cancer. The behavior of tumors may be affected by the tumor stromal environment. In this study, the authors investigated the effects of multi-walled nanotubes (MWNT) on tumor cell biology. The positive findings may point a new way in using this modality for treatment of triple-negative breast cancer in the future.

Topics: Cell Line, Tumor; Cell Movement; Cell Proliferation; Cellular Reprogramming; Coculture Techniques; Collagen; Female; Gene Expression Regulation, Neoplastic; Humans; Mesenchymal Stem Cells; Nanotubes, Carbon; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Mesenchymal stem-like/claudin-low (MSL/CL) breast cancers are highly aggressive, express low cell-cell adhesion cluster containing claudins (CLDN3/CLDN4/CLDN7) with enrichment of epithelial-to-mesenchymal transition (EMT), immunomodulatory, and transforming growth factor-β (TGF-β) genes. We examined the biological, molecular and prognostic impact of TGF-β upregulation and/or inhibition using in vivo and in vitro methods. Using publically available breast cancer gene expression databases, we show that upregulation and enrichment of a TGF-β gene signature is most frequent in MSL/CL breast cancers and is associated with a worse outcome. Using several MSL/CL breast cancer cell lines, we show that TGF-β elicits significant increases in cellular proliferation, migration, invasion, and motility, whereas these effects can be abrogated by a specific inhibitor against TGF-β receptor I and the anti-diabetic agent metformin, alone or in combination. Prior reports from our lab show that TNBC is exquisitely sensitive to metformin treatment. Mechanistically, metformin blocks endogenous activation of Smad2 and Smad3 and dampens TGF-β-mediated activation of Smad2, Smad3, and ID1 both at the transcriptional and translational level. We report the use of ID1 and ID3 as clinical surrogate markers, where high expression of these TGF-β target genes was correlated to poor prognosis in claudin-low patients. Given TGF-β's role in tumorigenesis and immunomodulation, blockade of this pathway using direct kinase inhibitors or more broadly acting inhibitors may dampen or abolish pro-carcinogenic and metastatic signaling in patients with MCL/CL TNBC. Metformin therapy (with or without other agents) may be a heretofore unrecognized approach to reduce the oncogenic activities associated with TGF-β mediated oncogenesis.

Topics: Biomarkers, Tumor; Carcinogenesis; Cell Line, Tumor; Cell Proliferation; Claudins; Disease-Free Survival; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Gene Knock-In Techniques; Humans; Mesenchymal Stem Cells; Metformin; Neoplasm Invasiveness; Prognosis; Protein Kinase Inhibitors; RNA, Messenger; Signal Transduction; Smad Proteins; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Up-Regulation

P-element-induced wimpy testis (PIWI) proteins bind to PIWI-interacting RNAs and play key roles in the biogenesis and functions of PIWI-interacting RNAs. It has been reported that PIWI proteins are essential for stem cell self-renewal and germline development in diverse organisms and that they are ectopically expressed in multiple forms of cancer. However, the role of PIWI in cancer remains elusive. Here we report that one of the four PIWI proteins in humans, PIWIL4, is highly expressed in both breast cancer tissues and the cytoplasm of MDA-MB-231 cells derived from breast cancer. Reducing PIWIL4 expression drastically impairs the migration ability of MDA-MB-231 cells, significantly increases their apoptosis, and mildly affects their proliferation. Our transcriptome and proteome analysis reveal that these functions are at least partially achieved via the PIWIL4 regulation of TGF-β and FGF signaling pathways and MHC class II proteins. These findings suggest that PIWIL4 may serve as a potential therapeutic target for breast cancer.

Topics: Apoptosis; Argonaute Proteins; Breast Neoplasms; Cell Line, Tumor; Cell Movement; Cell Proliferation; Female; Fibroblast Growth Factors; Gene Knockdown Techniques; Humans; RNA-Binding Proteins; RNA, Messenger; RNA, Neoplasm; Signal Transduction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

We previously reported that constitutive c-Abl activity (CST-Abl) abrogates the tumorigenicity of triple-negative breast cancer cells through the combined actions of two cellular events: downregulated matrix metalloproteinase (MMP) and upregulated p21Waf1/Cip1 expression. We now find decreased c-Abl expression to be significantly associated with diminished relapse-fee survival in breast cancer patients, particularly those exhibiting invasive and basal phenotypes. Moreover, CST-Abl expression enabled 4T1 cells to persist innocuously in the mammary glands of mice, doing so by exhausting their supply of cancer stem cells. Restoring MMP-9 expression and activity in CST-Abl-expressing 4T1 cells failed to rescue their malignant phenotypes; however, rendering these same cells deficient in p21 expression not only delayed their acquisition of senescent phenotypes, but also partially restored their tumorigenicity in mice. Although 4T1 cells lacked detectable expression of p53, those engineered to express CST-Abl exhibited robust production and secretion of TGF-β1 that engendered the reactivated expression of p53. Mechanistically, TGF-β-mediated p53 expression transpired through the combined actions of Smad1/5/8 and Smad2, leading to the dramatic upregulation of p21 and its stimulation of TNBC senescence. Collectively, we identified a novel c-Abl:p53:p21 signaling axis that functions as a powerful suppressor of mammary tumorigenesis and metastatic progression.

Topics: Animals; Breast Neoplasms; Cell Line, Tumor; Cell Transformation, Neoplastic; Cyclin-Dependent Kinase Inhibitor p21; Disease Models, Animal; Disease Progression; Female; Gene Expression Regulation, Neoplastic; Humans; Mammary Neoplasms, Animal; Matrix Metalloproteinase 9; Mice; Models, Biological; Proto-Oncogene Proteins c-abl; Smad Proteins, Receptor-Regulated; Transcriptional Activation; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Tumor Suppressor Protein p53

Increased collagen expression in tumors is associated with increased risk of metastasis, and triple-negative breast cancer (TNBC) has the highest propensity to develop distant metastases when there is evidence of central fibrosis. Transforming growth factor-β (TGF-β) ligands regulated by cancer-associated fibroblasts (CAFs) promote accumulation of fibrosis and cancer progression. In the present study, we have evaluated TNBC tumors with enhanced collagen to determine whether we can reduce metastasis by targeting the CAFs with Pirfenidone (PFD), an anti-fibrotic agent as well as a TGF-β antagonist. In patient-derived xenograft models, TNBC tumors exhibited accumulated collagen and activated TGF-β signaling, and developed lung metastasis. Next, primary CAFs were established from 4T1 TNBC homograft tumors, TNBC xenograft tumors and tumor specimens of breast cancer patients. CAFs promoted primary tumor growth with more fibrosis and TGF-β activation and lung metastasis in 4T1 mouse model. We then examined the effects of PFD in vitro and in vivo. We found that PFD had inhibitory effects on cell viability and collagen production of CAFs in 2D culture. Furthermore, CAFs enhanced tumor growth and PFD inhibited the tumor growth induced by CAFs by causing apoptosis in the 3D co-culture assay of 4T1 tumor cells and CAFs. In vivo, PFD alone inhibited tumor fibrosis and TGF-β signaling but did not inhibit tumor growth and lung metastasis. However, PFD inhibited tumor growth and lung metastasis synergistically in combination with doxorubicin. Thus, PFD has great potential for a novel clinically applicable TNBC therapy that targets tumor-stromal interaction.

Topics: Animals; Antineoplastic Agents; Cancer-Associated Fibroblasts; Cell Line, Tumor; Cell Proliferation; Collagen; Dose-Response Relationship, Drug; Female; Humans; Lung Neoplasms; Mice, Inbred BALB C; Molecular Targeted Therapy; Pyridones; Signal Transduction; Time Factors; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Tumor Burden; Tumor Cells, Cultured; Tumor Microenvironment; Xenograft Model Antitumor Assays

Metastatic bone disease has a major impact on morbidity of breast cancer (BC) patients. Alterations in mTOR signaling are involved both in cancer progression and in osteoclast differentiation. The purpose of this study was to assess the role of mTOR inhibitor Everolimus (Eve) on osteoclastogenesis induced by triple negative BC cells. To this aim, we developed an in vitro human model of osteoclastogenesis from peripheral blood monocytes co-cultured with the triple negative SCP2 and the hormonal receptor positive MCF7 cell lines. Osteoclastogenesis was evaluated by TRAP staining, evaluation of F actin rings and Calcitonin Receptor expression. Eve significantly reduced differentiation induced by cancer cells and resulted more effective when evaluated in combination with Denosumab and Zoledronic Acid (Zol). Combination with Zol showed a total abrogation of osteoclast differentiation induced by the triple negative cell line, not by MCF7. Finally, we observed that Eve was active in the inhibition of the crosstalk between cancer cells and osteoclasts reproduced by our model, highlighting a new therapeutic choice for the subsetting of triple negative BC patients. We observed a difference in the response to bone-targeted therapy with respect to BC subtypes. Our model may represent a valid platform for preclinical trials on bone-targeted drugs and for the study of the interplay of BC with bone stromal cells.

Topics: Antineoplastic Agents; Biomarkers; Blotting, Western; Bone Neoplasms; Cell Differentiation; Cell Line, Tumor; Cells, Cultured; Coculture Techniques; Denosumab; Diphosphonates; Everolimus; Gene Expression; Humans; Imidazoles; Intercellular Adhesion Molecule-1; Interleukin-6; MCF-7 Cells; Models, Biological; Osteoclasts; Osteonectin; RANK Ligand; Receptors, CXCR4; Reverse Transcriptase Polymerase Chain Reaction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Zoledronic Acid

The contemporary use of nanomedicines for cancer treatment has been largely limited to serving as carriers for existing therapeutic agents. Here, we provide definitive evidence that, the metallofullerenol nanomaterial Gd@C82(OH)22, while essentially not toxic to normal mammary epithelial cells, possesses intrinsic inhibitory activity against triple-negative breast cancer cells. Gd@C82(OH)22 blocks epithelial-to-mesenchymal transition with resultant efficient elimination of breast cancer stem cells (CSCs) resulting in abrogation of tumour initiation and metastasis. In normoxic conditions, Gd@C82(OH)22 mediates these effects by blocking TGF-β signalling. Moreover, under hypoxic conditions found in the tumour microenvironment, cellular uptake of Gd@C82(OH)22 is facilitated where it functions as a bi-potent inhibitor of HIF-1α and TGF-β activities, enhancing CSC elimination. These studies indicate that nanomaterials can be engineered to directly target CSCs. Thus, Gd-metallofullerenol is identified as a kind of non-toxic CSC specific inhibitors with significant therapeutic potential.

Topics: Animals; Apoptosis; Cell Adhesion; Cell Line, Tumor; Cell Movement; Cell Proliferation; Enzyme-Linked Immunosorbent Assay; Epithelial-Mesenchymal Transition; Female; Fullerenes; Gadolinium; Humans; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; Mammary Neoplasms, Experimental; Mice; Mice, Inbred BALB C; Nanomedicine; Nanostructures; Neoplasm Invasiveness; Neoplasm Metastasis; Neoplasm Transplantation; Neoplastic Stem Cells; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Some mutations in cancer cells can be exploited for therapeutic intervention. However, for many cancer subtypes, including triple-negative breast cancer (TNBC), no frequently recurring aberrations could be identified to make such an approach clinically feasible. Characterized by a highly heterogeneous mutational landscape with few common features, many TNBCs cluster together based on their 'basal-like' transcriptional profiles. We therefore hypothesized that targeting TNBC cells on a systems level by exploiting the transcriptional cell state might be a viable strategy to find novel therapies for this highly aggressive disease. We performed a large-scale chemical genetic screen and identified a group of compounds related to the drug PKC412 (midostaurin). PKC412 induced apoptosis in a subset of TNBC cells enriched for the basal-like subtype and inhibited tumor growth in vivo. We employed a multi-omics approach and computational modeling to address the mechanism of action and identified spleen tyrosine kinase (SYK) as a novel and unexpected target in TNBC. Quantitative phosphoproteomics revealed that SYK inhibition abrogates signaling to STAT3, explaining the selectivity for basal-like breast cancer cells. This non-oncogene addiction suggests that chemical SYK inhibition may be beneficial for a specific subset of TNBC patients and demonstrates that targeting cell states could be a viable strategy to discover novel treatment strategies.

Topics: Animals; Antineoplastic Agents; Apoptosis; Cell Differentiation; Cell Line, Tumor; Cell Proliferation; Cell Survival; Drug Delivery Systems; Female; Gene Expression Profiling; Humans; Intracellular Signaling Peptides and Proteins; Mice; Molecular Docking Simulation; Molecular Targeted Therapy; Protein Interaction Domains and Motifs; Protein-Tyrosine Kinases; Proteomics; Sequence Analysis, RNA; Signal Transduction; STAT3 Transcription Factor; Staurosporine; Syk Kinase; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Xenograft Model Antitumor Assays

Transforming growth factor-β (TGF-β) functions to suppress tumorigenesis in normal mammary tissues and early-stage breast cancers and, paradoxically, acts to promote the metastasis and chemoresistance in late-stage breast cancers, particularly triple-negative breast cancers (TNBCs). Precisely how TGF-β acquires oncogenic characteristics in late-stage breast cancers remains unknown, as does the role of the endogenous mammalian target of rapamycin (mTOR) inhibitor, Dep domain-containing mTOR-interacting protein (Deptor), in coupling TGF-β to TNBC development and metastatic progression. Here we demonstrate that Deptor expression was downregulated in basal-like/TNBCs relative to their luminal counterparts. Additionally, Deptor expression was 1) inversely correlated with the metastatic ability of human (MCF10A) and mouse (4T1) TNBC progression series and 2) robustly repressed by several inducers of epithelial-mesenchymal transition programs. Functional disruption of Deptor expression in 4T07 cells significantly inhibited their proliferation and organoid growth in vitro, as well as prevented their colonization and tumor formation in the lungs of mice. In stark contrast, elevated Deptor expression was significantly associated with poorer overall survival of patients harboring estrogen receptor α-negative breast cancers. Accordingly, enforced Deptor expression in MDA-MB-231 cells dramatically enhanced their 1) organoid growth in vitro, 2) pulmonary outgrowth in mice, and 3) resistance to chemotherapies, an event dependent on the coupling of Deptor to survivin expression. Collectively, our findings highlight the dichotomous functions of Deptor in modulating the proliferation and survival of TNBCs during metastasis; they also implicate Deptor and its stimulation of survivin as essential components of TNBC resistance to chemotherapies and apoptotic stimuli.

Topics: Animals; Caspases; Cell Line, Tumor; Cell Survival; Disease Models, Animal; Drug Resistance, Neoplasm; Epithelial-Mesenchymal Transition; Estrogen Receptor alpha; Female; Gene Expression Regulation, Neoplastic; Heterografts; Humans; Inhibitor of Apoptosis Proteins; Intracellular Signaling Peptides and Proteins; Mice; Neoplasm Metastasis; Signal Transduction; Smad3 Protein; Survivin; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Triple-negative breast cancer (TNBC) is an aggressive form of breast cancer with no effective targeted therapy. Inducible nitric oxide synthase (iNOS) is associated with poor survival in patients with breast cancer by increasing tumor aggressiveness. This work aimed to investigate the potential of iNOS inhibitors as a targeted therapy for TNBC. We hypothesized that inhibition of endogenous iNOS would decrease TNBC aggressiveness by reducing tumor initiation and metastasis through modulation of epithelial-mesenchymal transition (EMT)-inducing factors.. iNOS protein levels were determined in 83 human TNBC tissues and correlated with clinical outcome. Proliferation, mammosphere-forming efficiency, migration, and EMT transcription factors were assessed in vitro after iNOS inhibition. Endogenous iNOS targeting was evaluated as a potential therapy in TNBC mouse models.. High endogenous iNOS expression was associated with worse prognosis in patients with TNBC by gene expression as well as immunohistochemical analysis. Selective iNOS (1400 W) and pan-NOS (L-NMMA and L-NAME) inhibitors diminished cell proliferation, cancer stem cell self-renewal, and cell migration in vitro, together with inhibition of EMT transcription factors (Snail, Slug, Twist1, and Zeb1). Impairment of hypoxia-inducible factor 1α, endoplasmic reticulum stress (IRE1α/XBP1), and the crosstalk between activating transcription factor 3/activating transcription factor 4 and transforming growth factor β was observed. iNOS inhibition significantly reduced tumor growth, the number of lung metastases, tumor initiation, and self-renewal.. Considering the effectiveness of L-NMMA in decreasing tumor growth and enhancing survival rate in TNBC, we propose a targeted therapeutic clinical trial by re-purposing the pan-NOS inhibitor L-NMMA, which has been extensively investigated for cardiogenic shock as an anti-cancer therapeutic.

Topics: Activating Transcription Factor 3; Animals; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cell Transformation, Neoplastic; Disease Models, Animal; Endoplasmic Reticulum Stress; Enzyme Inhibitors; Epithelial-Mesenchymal Transition; Female; Gene Expression; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Lung Neoplasms; Mice; Molecular Targeted Therapy; Neoplasm Invasiveness; Nitric Oxide Synthase Type II; Prognosis; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Tumor Burden; Xenograft Model Antitumor Assays

Triple-negative breast cancer (TNBC) is a highly aggressive tumor subtype associated with a poor prognosis. The mechanism involved in TNBC progression remains largely unknown. To date, there are no effective therapeutic targets for this tumor subtype. In this study, by performing quantitative proteomic analyses in highly metastatic and parental breast cancer cell line, we found that RAB1B, a member of the RAS oncogene family, was significantly down-regulated in highly metastatic breast cancer cells. Moreover, down-regulation of RAB1B was also found to promote the proliferation and migration of TNBC cells in vitro and in vivo. Mechanistically, loss of RAB1B resulted in elevated expression of TGF-β receptor 1 (TβR1) through decreased degradation of ubiquitin, increased levels of phosphorylated SMAD3 and TGF-β-induced epithelial-mesenchymal transition (EMT). Furthermore, low RAB1B expression correlated with poor prognosis in breast cancer patients. Taken together, our findings reveal that RAB1B acts as a metastasis suppressor in TNBC by regulating the TGF-β/SMAD signaling pathway and RAB1B may serve as a novel biomarker of prognosis and the response to anti-tumor therapeutics for patients with TNBC.

Topics: Animals; Biomarkers, Tumor; Carcinoma, Ductal, Breast; Cell Line, Tumor; Cell Movement; Cell Proliferation; Down-Regulation; Enzyme Activation; Epithelial-Mesenchymal Transition; Female; Gene Expression Regulation, Neoplastic; HEK293 Cells; Heterografts; Humans; Lung Neoplasms; Mice; Mice, Inbred BALB C; Mice, Nude; Middle Aged; Neoplasm Transplantation; Prognosis; rab1 GTP-Binding Proteins; Receptors, Transforming Growth Factor beta; RNA Interference; RNA, Small Interfering; Signal Transduction; Smad3 Protein; Tissue Array Analysis; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Ubiquitin

Triple-negative breast cancers (TNBCs) are a heterogeneous group of breast tumours that are often associated with adverse pathological characteristics, poorer clinical outcomes and lack of targeted therapeutic options. Epithelial-mesenchymal transition, which plays a crucial role in tumour development and progression, is characterised by a transition from epithelial to mesenchymal phenotype and loss of proteins involved in maintaining cell junctions. We aimed to correlate protein expression of E-cadherin, Snail2 and transforming growth factor beta (TGF-β) with clinicopathological parameters and survivals of a series of patients with TNBC. The study cohort comprised 767 TNBCs diagnosed at the Department of Pathology, Singapore General Hospital from 1994 to 2012. Immunohistochemistry was performed on sections cut from tissue microarrays using the polymeric method. Staining intensity and percentage of positive tumour cells were evaluated and correlated with clinicopathological findings and clinical outcomes. Loss of E-cadherin expression, Snail2 positivity, cytoplasmic and nuclear expression of TGF-β were observed in 265 (35.2 %), 241 (32.0 %), 272 (36.2 %) and 262 (34.8 %) tumours, respectively. Histological grade significantly correlated with Snail2 positivity (P < 0.001) and loss of membranous E-cadherin expression (P = 0.003). Nuclear expression of TGF-β was inversely correlated with histological grade (P = 0.010). Median follow-up was 73 months, with a maximum of 236 months. Despite a graphical curve for earlier recurrence in patients with tumours harbouring a combinational phenotype of loss of membranous E-cadherin and positive Snail2 expression, there was no statistical significance. Similarly for women with tumours expressing cytoplasmic TGF-β, graphical representation showed poorer metastasis-free survival but without statistical significance. Loss of membranous E-cadherin and positive Snail2 expression are significantly associated with high-grade TNBCs. More work is needed to improve understanding of the role of TGF-β in TNBC.

Topics: Adult; Aged; Aged, 80 and over; Biomarkers, Tumor; Cadherins; Epithelial-Mesenchymal Transition; Female; Humans; Middle Aged; Snail Family Transcription Factors; Survival Analysis; Transcription Factors; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Triple-negative breast cancer (TNBC) is an aggressive cancer with a poor prognosis due to its epithelial‑to-mesenchymal transition (EMT) phenotype. Cancer patients often experience several detrimental effects of cancer treatment, such as chemoresistance, radioresistance and the maintenance of cancer stem cells due to EMT. Thus, EMT signaling is considered to be a valuable therapeutic target for cancer treatment, and its inhibition is being attempted as a new treatment option for TNBC patients. Previously, we showed that 3-(2-chlorobenzyl)-1,7-dimethyl-1H-imidazo[2,1-f]purine‑2,4(3H,8H)-dione (IM-412) inhibits transforming growth factor-β (TGF-β)-induced differentiation of human lung fibroblasts through both Smad-dependent and -independent pathways. In the present study, we examined the inhibitory effect of IM-412 on EMT pathways and invasiveness in TNBC cells since the TGF-β signaling pathway is a typical signaling pathway that functions in EMT. IM-412 not only potently suppressed the migration and invasion of MDA-MB-231 cells, but also lowered the expression of mesenchymal markers and EMT-activating transcription factors in these cells. IM-412 inhibited the activation of several signaling proteins, including Smad2/Smad3, p38MAPK, Akt and JNK, and it also attenuated the phosphorylation of FGFR1 and FGFR3. Collectively, our findings suggest that the synthetic compound IM-412 suppressed the EMT process in MDA-MB-231 cells and thereby effectively inhibited the migration and invasion of these cancer cells. Thus, IM-412 could serve as a novel therapeutic agent for malignant cancers.

Topics: Biomarkers, Tumor; Cell Differentiation; Cell Line, Tumor; Cell Movement; Epithelial-Mesenchymal Transition; Female; Humans; Imidazoles; Phosphorylation; Purines; Receptor, Fibroblast Growth Factor, Type 1; Receptor, Fibroblast Growth Factor, Type 3; Signal Transduction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Tumours frequently activate genes whose expression is otherwise biased to the testis, collectively known as cancer-testis antigens (CTAs). The extent to which CTA expression represents epiphenomena or confers tumorigenic traits is unknown. In this study, to address this, we implemented a multidimensional functional genomics approach that incorporates 7 different phenotypic assays in 11 distinct disease settings. We identify 26 CTAs that are essential for tumor cell viability and/or are pathological drivers of HIF, WNT or TGFβ signalling. In particular, we discover that Foetal and Adult Testis Expressed 1 (FATE1) is a key survival factor in multiple oncogenic backgrounds. FATE1 prevents the accumulation of the stress-sensing BH3-only protein, BCL-2-Interacting Killer (BIK), thereby permitting viability in the presence of toxic stimuli. Furthermore, ZNF165 promotes TGFβ signalling by directly suppressing the expression of negative feedback regulatory pathways. This action is essential for the survival of triple negative breast cancer cells in vitro and in vivo. Thus, CTAs make significant direct contributions to tumour biology.

Topics: Adenocarcinoma; Adenocarcinoma of Lung; Animals; Antigens, Neoplasm; Apoptosis Regulatory Proteins; Carcinogenesis; Cell Line; Cell Line, Tumor; Cell Survival; Colorectal Neoplasms; DNA-Binding Proteins; Fluorescent Antibody Technique; Gene Expression Regulation, Neoplastic; HCT116 Cells; HEK293 Cells; Humans; Immunoblotting; In Vitro Techniques; Lung Neoplasms; Membrane Proteins; Mice, Inbred NOD; Mitochondrial Proteins; Neoplasm Transplantation; Neoplasms; Prognosis; Proportional Hazards Models; Real-Time Polymerase Chain Reaction; Signal Transduction; Smad7 Protein; Transcription Factors; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Ubiquitin-Protein Ligases; Wnt Signaling Pathway

Interactions with the extracellular matrix (ECM) through integrin adhesion receptors provide cancer cells with physical and chemical cues that act together with growth factors to support survival and proliferation. Antagonists that target integrins containing the β1 subunit inhibit tumor growth and sensitize cells to irradiation or cytotoxic chemotherapy in preclinical breast cancer models and are under clinical investigation. We found that the loss of β1 integrins attenuated breast tumor growth but markedly enhanced tumor cell dissemination to the lungs. When cultured in three-dimensional ECM scaffolds, antibodies that blocked β1 integrin function or knockdown of β1 switched the migratory behavior of human and mouse E-cadherin-positive triple-negative breast cancer (TNBC) cells from collective to single cell movement. This switch involved activation of the transforming growth factor-β (TGFβ) signaling network that led to a shift in the balance between miR-200 microRNAs and the transcription factor zinc finger E-box-binding homeobox 2 (ZEB2), resulting in suppressed transcription of the gene encoding E-cadherin. Reducing the abundance of a TGFβ receptor, restoring the ZEB/miR-200 balance, or increasing the abundance of E-cadherin reestablished cohesion in β1 integrin-deficient cells and reduced dissemination to the lungs without affecting growth of the primary tumor. These findings reveal that β1 integrins control a signaling network that promotes an epithelial phenotype and suppresses dissemination and indicate that targeting β1 integrins may have undesirable effects in TNBC.

Topics: Animals; Blotting, Western; Cadherins; Cell Line, Tumor; Cell Movement; DNA-Binding Proteins; Extracellular Matrix; Flow Cytometry; Gene Silencing; Homeodomain Proteins; Humans; Immunohistochemistry; Integrin beta1; Luciferases; Lung Neoplasms; Mice; Mice, Knockout; MicroRNAs; Neoplasm Metastasis; Repressor Proteins; Signal Transduction; Time-Lapse Imaging; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Zebrafish; Zinc Finger E-box Binding Homeobox 2

Transforming growth factor beta (TGFβ) proteins are multitasking cytokines, in which high levels at tumor sites generally correlate with poor prognosis in human patients with cancer. Previously, it was reported that TGFβ downregulates the expression of ataxia telangiectasia-mutated (ATM) and mutS homolog 2 (MSH2) in breast cancer cells through an miRNA-mediated mechanism. In this study, expression of a panel of DNA-repair genes was examined, identifying breast cancer 1, early onset (BRCA1) as a target downregulated by TGFβ through the miR181 family. Correlations between the expression levels of TGFβ1 and the miR181/BRCA1 axis were observed in primary breast tumor specimens. By downregulating BRCA1, ATM, and MSH2, TGFβ orchestrates DNA damage response in certain breast cancer cells to induce a "BRCAness" phenotype, including impaired DNA-repair efficiency and synthetic lethality to the inhibition of poly (ADP-ribose) polymerase (PARP). Xenograft tumors with active TGFβ signaling exhibited resistance to the DNA-damaging agent doxorubicin but increased sensitivity to the PARP inhibitor ABT-888. Combination of doxorubicin with ABT-888 significantly improved the treatment efficacy in TGFβ-active tumors. Thus, TGFβ can induce "BRCAness" in certain breast cancers carrying wild-type BRCA genes and enhance the responsiveness to PARP inhibition, and the molecular mechanism behind this is characterized.. These findings enable better selection of patients with sporadic breast cancer for PARP interventions, which have exhibited beneficial effects in patients carrying BRCA mutations.

Topics: Animals; Ataxia Telangiectasia Mutated Proteins; BRCA1 Protein; Breast Neoplasms; Cell Line, Tumor; Disease Progression; DNA Repair; Down-Regulation; Doxorubicin; Female; Gene Expression Regulation, Neoplastic; Genomic Instability; Humans; Mice; MicroRNAs; MutS Homolog 2 Protein; Poly(ADP-ribose) Polymerase Inhibitors; Poly(ADP-ribose) Polymerases; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Triple-negative breast cancer is associated with poor prognosis because of a high rate of tumor recurrence and metastasis. Previous studies demonstrated that the synthetic triterpenoid, CDDO-Imidazolide (CDDO-Im) induced cell cycle arrest and apoptosis in triple-negative breast cancer. Since a small subpopulation of cancer stem cells has been suggested to be responsible for drug resistance and metastasis of tumors, our present study determined whether the effects of CDDO-Im in triple-negative breast cancer are due to the inhibition of a cancer stem cell subpopulation. CDDO-Im treatment markedly induced cell cycle arrest at G2/M-phase and apoptosis in the triple-negative breast cancer cell lines, SUM159 and MDA-MB-231. Because SUM159 cells were more sensitive to CDDO-Im than MDA-MB-231 cells, the effects of CDDO-Im on the cancer stem cell subpopulation were further investigated in SUM159 cells. SUM159 cells formed tumorspheres in culture, and the cancer stem cell subpopulation, CD24-/EpCAM+ cells, was markedly enriched in SUM159 tumorspheres. The CD24-/EpCAM+ cells in SUM159 tumorspheres were significantly inhibited by CDDO-Im treatment. CDDO-Im also significantly decreased sphere forming efficiency and tumorsphere size in both primary and secondary sphere cultures. PCR array of stem cell signaling genes showed that expression levels of many key molecules in the stem cell signaling pathways, such as Notch, TGF-β/Smad, Hedgehog and Wnt, were significantly down-regulated by CDDO-Im in SUM159 tumorspheres. Protein levels of Notch receptors (c-Notch1, Notch1 and Notch3), TGF-β/Smad (pSmad2/3) and Hedgehog downstream effectors (GLI1) also were markedly reduced by CDDO-Im. In conclusion, the present study demonstrates that the synthetic triterpenoid, CDDO-Im, is a potent anti-cancer agent against triple-negative breast cancer cells by targeting the cancer stem cell subpopulation.

Topics: Antineoplastic Agents; Apoptosis; Cell Cycle Checkpoints; Cell Line, Tumor; Dose-Response Relationship, Drug; G2 Phase Cell Cycle Checkpoints; Hedgehog Proteins; Humans; Imidazoles; M Phase Cell Cycle Checkpoints; Neoplastic Stem Cells; Oleanolic Acid; Receptors, Notch; Signal Transduction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms

Deregulation of the cell cycle machinery is often found in human cancers. Modulations in the cell cycle regulator function and expression result not only in proliferative advantages, but also lead to tumor progression and invasiveness of the cancer. In particular, cyclin D1 and p21 are often over-expressed in human cancers, correlating with high tumor grade, poor prognosis and increased metastasis. This prompted us to investigate the role of the cyclin D1/p21 signaling axis downstream of transforming growth factor beta (TGFβ) in breast cancer progression.. Cyclins mRNA and protein expressions were assessed by quantitative real-time PCR and Western blot in triple negative breast cancer cell lines. Co-localization and interaction between cyclin D1 and p21 were performed by immunocytochemistry and co-immunoprecipitation, respectively. Cell migration was assessed by wound healing and quantitative time-lapse imaging assays. In addition, the effects of cyclin D1 on cellular structure and actin organization were examined by staining with F-actin marker phalloidin and mesenchymal intermediate filament vimentin. Finally, a mammary fat pad xenograft mouse model was used to assess mammary tumor growth and local invasion.. We found TGFβ to specifically up-regulate the expression of cyclin D1 in triple negative breast cancer cells. Induction of cyclin D1 is also required for TGFβ-mediated cell migration. Suppression of cyclin D1 expression not only resulted in a rounded and epithelial-like phenotype, but also prevented TGFβ-induced vimentin and F-actin co-localization at the cell edge as well as invadopodia formation. Furthermore, TGFβ promoted the nuclear co-localization and physical interaction between cyclin D1 and p21. The co-expression of cyclin D1 and p21 proteins are required for the initial steps of tumor development, as double knockdown of these two molecules prevented primary tumor formation in a Xenograft mouse model. Moreover, the in vivo studies indicated that locally advanced features of the invasive tumors, including skeletal muscle, mammary fat pad and lymphovascular invasion, as well as ulcerated skin, were attenuated in the absence of cyclin D1 and p21.. Thus, our findings highlight the cyclin D1/p21 signaling axis as a critical regulator of TGFβ-mediated tumor growth initiation and local tumor cell invasion, both in vitro and in vivo.

Topics: Animals; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Movement; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; Female; Gene Expression Regulation, Neoplastic; Humans; Mice; Neoplasm Invasiveness; Signal Transduction; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Xenograft Model Antitumor Assays

Breast cancer is the second leading cause of cancer death in women in the United States. Metastasis accounts for the death of ~90 % of these patients, yet the mechanisms underlying this event remain poorly defined. WAVE3 belongs to the WASP/WAVE family of actin-binding proteins that play essential roles in regulating cell morphology, actin polymerization, cytoskeleton remodeling, cell motility, and invasion. Accordingly, we demonstrated previously that WAVE3 promotes the acquisition of invasive and metastatic phenotypes by human breast cancers. Herein, we show that transforming growth factor-β (TGF-β) selectively and robustly induced the expression of WAVE3 in metastatic breast cancer cells, but not in their nonmetastatic counterparts. Moreover, the induction of WAVE3 expression in human and mouse triple-negative breast cancer cells (TNBCs) by TGF-β likely reflects its coupling to microRNA expression via a Smad2- and β3 integrin-dependent mechanism. We further demonstrate the requirement for WAVE3 expression in mediating the initiation of epithelial-mesenchymal transition (EMT) programs stimulated by TGF-β. Indeed, stable depletion of WAVE3 expression in human TNBC cells prevented TGF-β from inducing EMT programs and from stimulating the proliferation, migration, and the formation of lamellipodia in metastatic TNBC cells. Lastly, we observed WAVE3 deficiency to abrogate the outgrowth of TNBC cell organoids in 3-dimensional organotypic cultures as well as to decrease the growth and metastasis of 4T1 tumors produced in syngeneic Balb/C mice. Indeed, WAVE3 deficiency significantly reduced the presence of sarcomatoid morphologies indicative of EMT phenotypes in pulmonary TNBC tumors as compared to those detected in their parental counterparts. Collectively, these findings indicate the necessity for WAVE3 expression and activity during EMT programs stimulated by TGF-β; they also suggest that measures capable of inactivating WAVE3 may play a role in alleviating metastasis stimulated by TGF-β.

Topics: Animals; Cell Movement; Cell Proliferation; Epithelial-Mesenchymal Transition; Female; Humans; Integrin beta3; Mice; Mice, Inbred BALB C; Smad2 Protein; Transforming Growth Factor beta; Triple Negative Breast Neoplasms; Up-Regulation; Wiskott-Aldrich Syndrome Protein Family; Xenograft Model Antitumor Assays