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1.
Front Oncol ; 14: 1412212, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38957320

RESUMO

Introduction: Oral cavity squamous cell carcinoma (OSCC) occurs most frequently in patients >60 years old with a history of tobacco and alcohol use. Epidemiological studies describe increased incidence of OSCC in younger adults (<45 years). Despite its poor prognosis, knowledge of OSCC tumor microenvironment (TME) characteristics in younger adults is scarce and could help inform possible resistance to emerging treatment options. Methods: Patients with OSCC were evaluated using TCGA-HNSC (n=121) and a stage and subsite-matched institutional cohort (n=8) to identify differential gene expression focusing on the extracellular matrix (ECM) and epithelial-mesenchymal transition (EMT) processes in younger (≤45 years) vs. older adults (≥60 years). NanoString nCounter analysis was performed using isolated total RNA from formalin-fixed paraffin-embedded (FFPE) tumor samples. Stained tumor slides from young and old OSCC patients were evaluated for CD8+ T-cell counts using immunohistochemistry. Results: Younger OSCC patients demonstrated significantly increased expression of ECM remodeling and EMT process genes, as well as TME immunosuppression. Gene set enrichment analyses demonstrated increased ECM pathways and concurrent decreased immune pathways in young relative to old patients. Transcripts per million of genetic markers involved in ECM remodeling including LAMB3, VCAN, S100A9, COL5A1, and ITGB2 were significantly increased in tumors of younger vs. older patients (adjusted p-value < 0.10). Young patient TMEs demonstrated a 2.5-fold reduction in CD8+ T-cells as compared to older patients (p < 0.05). Conclusion: Differential gene expression impacting ECM remodeling and TME immunosuppression may contribute to disease progression in younger adult OSCC and has implications on response to evolving treatment modalities, such as immune checkpoint inhibitor therapy.

2.
3.
Front Oncol ; 8: 324, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30211114

RESUMO

Head and neck squamous cell carcinoma (HNSCC) is the 6th most common human cancer and affects approximately 50,000 new patients every year in the US. The major risk factors for HNSCC are tobacco and alcohol consumption as well as oncogenic HPV infections. Despite advances in therapy, the overall survival rate for all-comers is only 50%. Understanding the biology of HNSCC is crucial to identifying new biomarkers, implementing early diagnostic approaches and developing novel therapies. As in several other cancers, HNSCC expresses elevated levels of MCT4, a member of the SLC16 family of monocarboxylate transporters. MCT4 is a H+-linked lactate transporter which functions to facilitate lactate efflux from highly glycolytic cells. High MCT4 levels in HNSCC have been associated with poor prognosis, but the role of MCT4 in the development and progression of this cancer is still poorly understood. In this study, we used 4-nitroquinoline-1-oxide (4NQO) to induce oral cancer in MCT4-/- and wild type littermates, recapitulating the disease progression in humans. Histological analysis of mouse tongues after 23 weeks of 4NQO treatment showed that MCT4-/- mice developed significantly fewer and less extended invasive lesions than wild type. In mice, as in human samples, MCT4 was not expressed in normal oral mucosa but was detected in the transformed epithelium. In the 4NQO treated mice we detected MCT4 in foci of the basal layer undergoing transformation, and progressively in areas of carcinoma in situ and invasive carcinomas. Moreover, we found MCT4 positive macrophages within the tumor and in the stroma surrounding the lesions in both human samples of HNSCC and in the 4NQO treated animals. The results of our studies showed that MCT4 could be used as an early diagnostic biomarker of HNSCC. Our finding with the MCT4-/- mice suggest MCT4 is a driver of progression to oral squamous cell cancer and MCT4 inhibitors could have clinical benefits for preventing invasive HNSCC.

4.
Front Oncol ; 8: 341, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30211120

RESUMO

Endometrial cancer is the most common gynecologic malignancy in developed countries. Its increasing incidence is thought to be related in part to the rise of metabolic syndrome, which has been shown to be a risk factor for the development of hyperestrogenic and hyperinsulinemic states. This has consequently lead to an increase in other hormone-responsive cancers as well e.g., breast and ovarian cancer. The correlation between obesity, hyperglycemia, and endometrial cancer has highlighted the important role of metabolism in cancer establishment and persistence. Tumor-mediated reprogramming of the microenvironment and macroenvironment can range from induction of cytokines and growth factors to stimulation of surrounding stromal cells to produce energy-rich catabolites, fueling the growth, and survival of cancer cells. Such mechanisms raise the prospect of the metabolic microenvironment itself as a viable target for treatment of malignancies. Metformin is a biguanide drug that is a first-line treatment for type 2 diabetes that has beneficial effects on various markers of the metabolic syndrome. Many studies suggest that metformin shows potential as an adjuvant treatment for uterine and other cancers. Here, we review the evidence for metformin as a treatment for cancers of the endometrium. We discuss the available clinical data and the molecular mechanisms by which it may exert its effects, with a focus on how it may alter the tumor microenvironment. The pleiotropic effects of metformin on cellular energy production and usage as well as intercellular and hormone-based interactions make it a promising candidate for reprogramming of the cancer ecosystem. This, along with other treatments aimed at targeting tumor metabolic pathways, may lead to novel treatment strategies for endometrial cancer.

5.
Front Oncol ; 8: 331, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30234009

RESUMO

For a long time, pioneers in the field of cancer cell metabolism, such as Otto Warburg, have focused on the idea that tumor cells maintain high glycolytic rates even with adequate oxygen supply, in what is known as aerobic glycolysis or the Warburg effect. Recent studies have reported a more complex situation, where the tumor ecosystem plays a more critical role in cancer progression. Cancer cells display extraordinary plasticity in adapting to changes in their tumor microenvironment, developing strategies to survive and proliferate. The proliferation of cancer cells needs a high rate of energy and metabolic substrates for biosynthesis of biomolecules. These requirements are met by the metabolic reprogramming of cancer cells and others present in the tumor microenvironment, which is essential for tumor survival and spread. Metabolic reprogramming involves a complex interplay between oncogenes, tumor suppressors, growth factors and local factors in the tumor microenvironment. These factors can induce overexpression and increased activity of glycolytic isoenzymes and proteins in stromal and cancer cells which are different from those expressed in normal cells. The fructose-6-phosphate/fructose-1,6-bisphosphate cycle, catalyzed by 6-phosphofructo-1-kinase/fructose 1,6-bisphosphatase (PFK1/FBPase1) isoenzymes, plays a key role in controlling glycolytic rates. PFK1/FBpase1 activities are allosterically regulated by fructose-2,6-bisphosphate, the product of the enzymatic activity of the dual kinase/phosphatase family of enzymes: 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase (PFKFB1-4) and TP53-induced glycolysis and apoptosis regulator (TIGAR), which show increased expression in a significant number of tumor types. In this review, the function of these isoenzymes in the regulation of metabolism, as well as the regulatory factors modulating their expression and activity in the tumor ecosystem are discussed. Targeting these isoenzymes, either directly or by inhibiting their activating factors, could be a promising approach for treating cancers.

6.
Front Cell Dev Biol ; 5: 27, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28421181

RESUMO

Introduction: Monocarboxylate transporter 1 (MCT1) is an importer of monocarboxylates such as lactate and pyruvate and a marker of mitochondrial metabolism. MCT1 is highly expressed in a subgroup of cancer cells to allow for catabolite uptake from the tumor microenvironment to support mitochondrial metabolism. We studied the protein expression of MCT1 in a broad group of breast invasive ductal carcinoma specimens to determine its association with breast cancer subtypes and outcomes. Methods: MCT1 expression was evaluated by immunohistochemistry on tissue micro-arrays (TMA) obtained through our tumor bank. Two hundred and fifty-seven cases were analyzed: 180 cases were estrogen receptor and/or progesterone receptor positive (ER+ and/or PR+), 62 cases were human epidermal growth factor receptor 2 positive (HER2+), and 56 cases were triple negative breast cancers (TNBC). MCT1 expression was quantified by digital pathology with Aperio software. The intensity of the staining was measured on a continuous scale (0-black to 255-bright white) using a co-localization algorithm. Statistical analysis was performed using a linear mixed model. Results: High MCT1 expression was more commonly found in TNBC compared to ER+ and/or PR+ and compared to HER-2+ (p < 0.001). Tumors with an in-situ component were less likely to stain strongly for MCT1 (p < 0.05). High nuclear grade was associated with higher MCT1 staining (p < 0.01). Higher T stage tumors were noted to have a higher expression of MCT1 (p < 0.05). High MCT1 staining in cancer cells was associated with shorter progression free survival, increased risk of recurrence, and larger size independent of TNBC status (p < 0.05). Conclusion: MCT1 expression, which is a marker of high catabolite uptake and mitochondrial metabolism, is associated with recurrence in breast invasive ductal carcinoma. MCT1 expression as quantified with digital image analysis may be useful as a prognostic biomarker and to design clinical trials using MCT1 inhibitors.

8.
Oncotarget ; 8(6): 9868-9884, 2017 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-28039467

RESUMO

In this report, we systematically examined the role of telomerase activity in lung and ovarian cancer stem cell (CSC) propagation. For this purpose, we indirectly gauged telomerase activity, by linking the hTERT-promoter to eGFP. Using lung (A549) and ovarian (SKOV3) cancer cells, transduced with the hTERT-GFP reporter, we then employed GFP-expression levels to fractionate these cell lines into GFP-high and GFP-low populations. We functionally compared the phenotype of these GFP-high and GFP-low populations. More specifically, we now show that the cancer cells with higher telomerase activity (GFP-high) are more energetically activated, with increased mitochondrial mass and function, as well as increased glycolytic activity. This was further validated and confirmed by unbiased proteomics analysis. Cells with high telomerase activity also showed an increased capacity for stem cell activity (as measured using the 3D-spheroid assay) and cell migration (as measured using a Boyden chamber approach). These enhanced biological phenotypes were effectively inhibited by classical modulators of energy metabolism, which target either i) mitochondrial metabolism (i.e., oligomycin) or ii) glycolysis (i.e., 2-deoxy-glucose), or iii) by using the FDA-approved antibiotic doxycycline, which inhibits mitochondrial biogenesis. Finally, the level of telomerase activity also determined the ability of hTERT-high cells to proliferate, as assessed by measuring DNA synthesis via EdU incorporation. Consistent with these observations, treatment with an FDA-approved CDK4/6 inhibitor (PD-0332991/palbociclib) specifically blocked the propagation of both lung and ovarian CSCs. Virtually identical results were obtained with breast CSCs, which were also highly sensitive to palbociclib at concentrations in the nanomolar range. In summary, CSCs with high telomerase activity are among the most energetically activated, migratory and proliferative cell sub-populations. These observations may provide a mechanistic explanation for tumor metabolic heterogeneity, based on telomerase activity. FDA-approved drugs, such as doxycycline and palbociclib, were both effective at curtailing CSC propagation. Thus, these FDA-approved drugs could be used to target telomerase-high proliferative CSCs, in multiple cancer types. Finally, our experiments also allowed us to distinguish two different cellular populations of hTERT-high cells, one that was proliferative (i.e., replicative immortality) and the other that was non-proliferative (i.e., quiescent). We speculate that the non-proliferative population of hTERT-high cells that we identified could be mechanistically involved in tumor dormancy.


Assuntos
Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Neoplasias Pulmonares/tratamento farmacológico , Células-Tronco Neoplásicas/efeitos dos fármacos , Neoplasias Ovarianas/tratamento farmacológico , Piperazinas/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Piridinas/farmacologia , Telomerase/metabolismo , Células A549 , Movimento Celular/efeitos dos fármacos , Quinase 4 Dependente de Ciclina/metabolismo , Quinase 6 Dependente de Ciclina/metabolismo , Relação Dose-Resposta a Droga , Metabolismo Energético/efeitos dos fármacos , Feminino , Humanos , Neoplasias Pulmonares/enzimologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/enzimologia , Mitocôndrias/patologia , Células-Tronco Neoplásicas/enzimologia , Células-Tronco Neoplásicas/patologia , Neoplasias Ovarianas/enzimologia , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/patologia , Fenótipo , Proteômica/métodos , Transdução de Sinais/efeitos dos fármacos , Telomerase/genética , Fatores de Tempo , Transdução Genética
9.
Nat Rev Clin Oncol ; 14(1): 11-31, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27141887

RESUMO

Awareness that the metabolic phenotype of cells within tumours is heterogeneous - and distinct from that of their normal counterparts - is growing. In general, tumour cells metabolize glucose, lactate, pyruvate, hydroxybutyrate, acetate, glutamine, and fatty acids at much higher rates than their nontumour equivalents; however, the metabolic ecology of tumours is complex because they contain multiple metabolic compartments, which are linked by the transfer of these catabolites. This metabolic variability and flexibility enables tumour cells to generate ATP as an energy source, while maintaining the reduction-oxidation (redox) balance and committing resources to biosynthesis - processes that are essential for cell survival, growth, and proliferation. Importantly, experimental evidence indicates that metabolic coupling between cell populations with different, complementary metabolic profiles can induce cancer progression. Thus, targeting the metabolic differences between tumour and normal cells holds promise as a novel anticancer strategy. In this Review, we discuss how cancer cells reprogramme their metabolism and that of other cells within the tumour microenvironment in order to survive and propagate, thus driving disease progression; in particular, we highlight potential metabolic vulnerabilities that might be targeted therapeutically.


Assuntos
Neoplasias/metabolismo , Acetilcoenzima A/metabolismo , Adaptação Fisiológica , Aminoácidos/metabolismo , Antineoplásicos/uso terapêutico , Antioxidantes/metabolismo , Autofagia/fisiologia , Glicemia/metabolismo , Metabolismo Energético/efeitos dos fármacos , Epigenômica , Ácidos Graxos/metabolismo , Heterogeneidade Genética , Ácido Glutâmico/metabolismo , Glutamina/metabolismo , Humanos , Corpos Cetônicos/metabolismo , Ácido Láctico/metabolismo , Lipídeos/biossíntese , Mitocôndrias/efeitos dos fármacos , Ribossomos Mitocondriais/efeitos dos fármacos , Neoplasias/tratamento farmacológico , Ácidos Nucleicos/biossíntese , Estresse Oxidativo/efeitos dos fármacos , Ácido Pirúvico/metabolismo , Serina-Treonina Quinases TOR/antagonistas & inibidores , Fatores de Transcrição/metabolismo , Microambiente Tumoral
10.
Appl Immunohistochem Mol Morphol ; 25(1): e1-e8, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-27093448

RESUMO

We report a case of Epstein-Barr virus (EBV)-associated T-cell lymphoma of gastrointestinal (GI) tract from a 70-year-old white woman who initially presented with a widespread GI inflammation and gastric obstruction. Initial biopsies of the GI tract showed severe chronic inflammation in the esophagus, stomach, and the small intestine. Celiac disease and inflammatory bowel disease were ruled out. The patient was treated with partial gastrectomy. Histology showed gastric wall thickening with EBV-positive, mixed lymphocytic and plasma cell infiltration in the mucosa, and thickening and fibrosis of the submucosa. She developed EBV-associated T-cell lymphoma of the GI tract one and a half years later and expired due to multiorgan failure. The T-cell lymphoma diffusely infiltrated the GI wall without forming a mass lesion. The lymphoma expressed EBV and cytotoxic molecules but lacked common features of extranodal natural killer/T-cell lymphoma nasal type, such as angioinvasion/angiodestruction, necrosis, or CD56 expression. Immunoglobulin heavy chain (IGH) gene and T-cell receptor-γ gene rearrangements and EBV-positive cells were detected at the early stage of the disease. While IGH clones were transient, T-cell clones and EBV-positive cells progressively increased over the disease course. We conclude that this case is best classified as EBV-associated peripheral T-cell lymphoma of GI tract. Age-related immune senescence may have contributed to the uncontrolled GI inflammation and subsequent transformation to T-cell lymphoma.


Assuntos
Infecções por Vírus Epstein-Barr/patologia , Inflamação/patologia , Linfoma de Células T Periférico/patologia , Idoso , Doença Crônica , Feminino , Humanos , Hibridização In Situ , Linfoma de Células T Periférico/virologia
11.
Oncotarget ; 7(45): 72395-72414, 2016 Nov 08.
Artigo em Inglês | MEDLINE | ID: mdl-27590350

RESUMO

Despite ubiquitous activation in human cancer, essential downstream effector pathways of the MYC transcription factor have been difficult to define and target. Using a structure/function-based approach, we identified the mitochondrial RNA polymerase (POLRMT) locus as a critical downstream target of MYC. The multifunctional POLRMT enzyme controls mitochondrial gene expression, a process required both for mitochondrial function and mitochondrial biogenesis. We further demonstrate that inhibition of this newly defined MYC effector pathway causes robust and selective tumor cell apoptosis, via an acute, checkpoint-like mechanism linked to aberrant electron transport chain complex assembly and mitochondrial reactive oxygen species (ROS) production. Fortuitously, MYC-dependent tumor cell death can be induced by inhibiting the mitochondrial gene expression pathway using a variety of strategies, including treatment with FDA-approved antibiotics. In vivo studies using a mouse model of Burkitt's Lymphoma provide pre-clinical evidence that these antibiotics can successfully block progression of MYC-dependent tumors.


Assuntos
Regulação Neoplásica da Expressão Gênica , Genes Mitocondriais , Genes myc , Neoplasias/genética , Animais , Linhagem Celular Tumoral , RNA Polimerases Dirigidas por DNA/genética , RNA Polimerases Dirigidas por DNA/metabolismo , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Neoplasias/patologia , Proteínas Proto-Oncogênicas c-myc , Espécies Reativas de Oxigênio/metabolismo , Transfecção
12.
Aging (Albany NY) ; 8(8): 1593-607, 2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-27344270

RESUMO

Bedaquiline (a.k.a., Sirturo) is an anti-microbial agent, which is approved by the FDA for the treatment of multi-drug resistant pulmonary tuberculosis (TB). Bedaquiline is a first-in-class diaryl-quinoline compound, that mechanistically inhibits the bacterial ATP-synthase, and shows potent activity against both drug-sensitive and drug-resistant TB. Interestingly, eukaryotic mitochondria originally evolved from engulfed aerobic bacteria. Thus, we hypothesized that, in mammalian cells, bedaquiline might also target the mitochondrial ATP-synthase, leading to mitochondrial dysfunction and ATP depletion. Here, we show that bedaquiline has anti-cancer activity, directed against Cancer Stem-like Cells (CSCs). More specifically, we demonstrate that bedaquiline treatment of MCF7 breast cancer cells inhibits mitochondrial oxygen-consumption, as well as glycolysis, but induces oxidative stress. Importantly, bedaquiline significantly blocks the propagation and expansion of MCF7-derived CSCs, with an IC-50 of approx. 1-µM, as determined using the mammosphere assay. Similarly, bedaquiline also reduces both the CD44+/CD24low/- CSC and ALDH+ CSC populations, under anchorage-independent growth conditions. In striking contrast, bedaquiline significantly increases oxygen consumption in normal human fibroblasts, consistent with the fact that it is well-tolerated in patients treated for TB infections. As such, future pre-clinical studies and human clinical trials in cancer patients may be warranted. Interestingly, we also highlight that bedaquiline shares certain structural similarities with trans-piceatannol and trans-resveratrol, which are known natural flavonoid inhibitors of the mitochondrial ATP-synthase (complex V) and show anti-aging properties.


Assuntos
Antituberculosos/farmacologia , Proliferação de Células/efeitos dos fármacos , Diarilquinolinas/farmacologia , Mitocôndrias/efeitos dos fármacos , Células-Tronco Neoplásicas/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Humanos , Células MCF-7 , Mitocôndrias/metabolismo , Células-Tronco Neoplásicas/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Consumo de Oxigênio/efeitos dos fármacos
13.
Oncotarget ; 7(23): 34084-99, 2016 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-27136895

RESUMO

Atovaquone is an FDA-approved anti-malarial drug, which first became clinically available in the year 2000. Currently, its main usage is for the treatment of pneumocystis pneumonia (PCP) and/or toxoplasmosis in immune-compromised patients. Atovaquone is a hydroxy-1,4-naphthoquinone analogue of ubiquinone, also known as Co-enzyme Q10 (CoQ10). It is a well-tolerated drug that does not cause myelo-suppression. Mechanistically, it is thought to act as a potent and selective OXPHOS inhibitor, by targeting the CoQ10-dependence of mitochondrial complex III. Here, we show for the first time that atovaquone also has anti-cancer activity, directed against Cancer Stem-like Cells (CSCs). More specifically, we demonstrate that atovaquone treatment of MCF7 breast cancer cells inhibits oxygen-consumption and metabolically induces aerobic glycolysis (the Warburg effect), as well as oxidative stress. Remarkably, atovaquone potently inhibits the propagation of MCF7-derived CSCs, with an IC-50 of 1 µM, as measured using the mammosphere assay. Atovaquone also maintains this selectivity and potency in mixed populations of CSCs and non-CSCs. Importantly, these results indicate that glycolysis itself is not sufficient to maintain the proliferation of CSCs, which is instead strictly dependent on mitochondrial function. In addition to targeting the proliferation of CSCs, atovaquone also induces apoptosis in both CD44+/CD24low/- CSC and ALDH+ CSC populations, during exposure to anchorage-independent conditions for 12 hours. However, it has no effect on oxygen consumption in normal human fibroblasts and, in this cellular context, behaves as an anti-inflammatory, consistent with the fact that it is well-tolerated in patients treated for infections. Future studies in xenograft models and human clinical trials may be warranted, as the IC-50 of atovaquone's action on CSCs (1 µM) is >50 times less than its average serum concentration in humans.


Assuntos
Antineoplásicos/farmacologia , Atovaquona/farmacologia , Células-Tronco Neoplásicas/efeitos dos fármacos , Fosforilação Oxidativa/efeitos dos fármacos , Antimaláricos/farmacologia , Reposicionamento de Medicamentos , Complexo III da Cadeia de Transporte de Elétrons/antagonistas & inibidores , Humanos , Células MCF-7
14.
Breast Cancer Res ; 18(1): 55, 2016 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-27220421

RESUMO

Cancer is now viewed as a stem cell disease. There is still no consensus on the metabolic characteristics of cancer stem cells, with several studies indicating that they are mainly glycolytic and others pointing instead to mitochondrial metabolism as their principal source of energy. Cancer stem cells also seem to adapt their metabolism to microenvironmental changes by conveniently shifting energy production from one pathway to another, or by acquiring intermediate metabolic phenotypes. Determining the role of cancer stem cell metabolism in carcinogenesis has become a major focus in cancer research, and substantial efforts are conducted towards discovering clinical targets.


Assuntos
Metabolismo Energético , Neoplasias/metabolismo , Células-Tronco Neoplásicas/metabolismo , Animais , Feminino , Humanos , Redes e Vias Metabólicas , Mitocôndrias/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/imunologia , Neoplasias/patologia , Células-Tronco Neoplásicas/efeitos dos fármacos , Microambiente Tumoral/imunologia
15.
Cell Metab ; 22(6): 956-8, 2015 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-26636492

RESUMO

The role of reactive oxygen species (ROS) and antioxidants in cancer is controversial because of their context-dependent ability to promote or suppress tumorigenesis. Piskounova et al. (2015) now show that ROS limit distant metastasis: only cells with increased antioxidant capacity are able to succeed in their purpose to metastasize.


Assuntos
Melanoma/metabolismo , Melanoma/patologia , Metástase Neoplásica/prevenção & controle , Estresse Oxidativo , Animais , Feminino , Humanos , Masculino
16.
Oncotarget ; 6(31): 30453-71, 2015 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-26421711

RESUMO

Here, we developed an isogenic cell model of "stemness" to facilitate protein biomarker discovery in breast cancer. For this purpose, we used knowledge gained previously from the study of the mouse mammary tumor virus (MMTV). MMTV initiates mammary tumorigenesis in mice by promoter insertion adjacent to two main integration sites, namely Int-1 (Wnt1) and Int-2 (Fgf3), which ultimately activates Wnt/ß-catenin signaling, driving the propagation of mammary cancer stem cells (CSCs). Thus, to develop a humanized model of MMTV signaling, we over-expressed WNT1 and FGF3 in MCF7 cells, an ER(+) human breast cancer cell line. We then validated that MCF7 cells over-expressing both WNT1 and FGF3 show a 3.5-fold increase in mammosphere formation, and that conditioned media from these cells is also sufficient to promote stem cell activity in untransfected parental MCF7 and T47D cells, as WNT1 and FGF3 are secreted factors. Proteomic analysis of this model system revealed the induction of i) EMT markers, ii) mitochondrial proteins, iii) glycolytic enzymes and iv) protein synthesis machinery, consistent with an anabolic CSC phenotype. MitoTracker staining validated the expected WNT1/FGF3-induced increase in mitochondrial mass and activity, which presumably reflects increased mitochondrial biogenesis. Importantly, many of the proteins that were up-regulated by WNT/FGF-signaling in MCF7 cells, were also transcriptionally over-expressed in human breast cancer cells in vivo, based on the bioinformatic analysis of public gene expression datasets of laser-captured patient samples. As such, this isogenic cell model should accelerate the discovery of new biomarkers to predict clinical outcome in breast cancer, facilitating the development of personalized medicine.Finally, we used mitochondrial mass as a surrogate marker for increased mitochondrial biogenesis in untransfected MCF7 cells. As predicted, metabolic fractionation of parental MCF7 cells, via MitoTracker staining, indicated that high mitochondrial mass is a new metabolic biomarker for the enrichment of anabolic CSCs, as functionally assessed by mammosphere-forming activity. This observation has broad implications for understanding the role of mitochondrial biogenesis in the propagation of stem-like cancer cells. Technically, this general metabolic approach could be applied to any cancer type, to identify and target the mitochondrial-rich CSC population.The implications of our work for understanding the role of mitochondrial metabolism in viral oncogenesis driven by random promoter insertions are also discussed, in the context of MMTV and ALV infections.


Assuntos
Biomarcadores Tumorais/fisiologia , Neoplasias da Mama/patologia , Fator 3 de Crescimento de Fibroblastos/biossíntese , Mitocôndrias/fisiologia , Proteína Wnt1/biossíntese , Meios de Cultivo Condicionados/farmacologia , Feminino , Fator 3 de Crescimento de Fibroblastos/metabolismo , Humanos , Células MCF-7 , Vírus do Tumor Mamário do Camundongo/genética , Vírus do Tumor Mamário do Camundongo/patogenicidade , Potencial da Membrana Mitocondrial/fisiologia , Mitocôndrias/metabolismo , Modelos Biológicos , Células-Tronco Neoplásicas/citologia , Células-Tronco Neoplásicas/patologia , Esferoides Celulares/citologia , Células Tumorais Cultivadas , Via de Sinalização Wnt/fisiologia , Proteína Wnt1/metabolismo
17.
Oncotarget ; 6(26): 21892-905, 2015 Sep 08.
Artigo em Inglês | MEDLINE | ID: mdl-26323205

RESUMO

Tumor cell metabolic heterogeneity is thought to contribute to tumor recurrence, distant metastasis and chemo-resistance in cancer patients, driving poor clinical outcome. To better understand tumor metabolic heterogeneity, here we used the MCF7 breast cancer line as a model system to metabolically fractionate a cancer cell population. First, MCF7 cells were stably transfected with an hTERT-promoter construct driving GFP expression, as a surrogate marker of telomerase transcriptional activity. To enrich for immortal stem-like cancer cells, MCF7 cells expressing the highest levels of GFP (top 5%) were then isolated by FACS analysis. Notably, hTERT-GFP(+) MCF7 cells were significantly more efficient at forming mammospheres (i.e., stem cell activity) and showed increased mitochondrial mass and mitochondrial functional activity, all relative to hTERT-GFP(-) cells. Unbiased proteomics analysis of hTERT-GFP(+) MCF7 cells directly demonstrated the over-expression of 33 key mitochondrial proteins, 17 glycolytic enzymes, 34 ribosome-related proteins and 17 EMT markers, consistent with an anabolic cancer stem-like phenotype. Interestingly, MT-CO2 (cytochrome c oxidase subunit 2; Complex IV) expression was increased by >20-fold. As MT-CO2 is encoded by mt-DNA, this finding is indicative of increased mitochondrial biogenesis in hTERT-GFP(+) MCF7 cells. Importantly, most of these candidate biomarkers were transcriptionally over-expressed in human breast cancer epithelial cells in vivo. Similar results were obtained using cell size (forward/side scatter) to fractionate MCF7 cells. Larger stem-like cells also showed increased hTERT-GFP levels, as well as increased mitochondrial mass and function. Thus, this simple and rapid approach for the enrichment of immortal anabolic stem-like cancer cells will allow us and others to develop new prognostic biomarkers and novel anti-cancer therapies, by specifically and selectively targeting this metabolic sub-population of aggressive cancer cells. Based on our proteomics and functional analysis, FDA-approved inhibitors of protein synthesis and/or mitochondrial biogenesis, may represent novel treatment options for targeting these anabolic stem-like cancer cells.


Assuntos
Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Mitocôndrias/metabolismo , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Telomerase/metabolismo , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/genética , Linhagem Celular Tumoral , Feminino , Humanos , Células MCF-7 , Mitocôndrias/genética , Proteômica/métodos , Telomerase/genética , Regulação para Cima
18.
Oncotarget ; 6(16): 14005-25, 2015 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-26087309

RESUMO

DNA-PK is an enzyme that is required for proper DNA-repair and is thought to confer radio-resistance in cancer cells. As a consequence, it is a high-profile validated target for new pharmaceutical development. However, no FDA-approved DNA-PK inhibitors have emerged, despite many years of drug discovery and lead optimization. This is largely because existing DNA-PK inhibitors suffer from poor pharmacokinetics. They are not well absorbed and/or are unstable, with a short plasma half-life. Here, we identified the first FDA-approved DNA-PK inhibitor by "chemical proteomics". In an effort to understand how doxycycline targets cancer stem-like cells (CSCs), we serendipitously discovered that doxycycline reduces DNA-PK protein expression by nearly 15-fold (> 90%). In accordance with these observations, we show that doxycycline functionally radio-sensitizes breast CSCs, by up to 4.5-fold. Moreover, we demonstrate that DNA-PK is highly over-expressed in both MCF7- and T47D-derived mammospheres. Interestingly, genetic or pharmacological inhibition of DNA-PK in MCF7 cells is sufficient to functionally block mammosphere formation. Thus, it appears that active DNA-repair is required for the clonal expansion of CSCs. Mechanistically, doxycycline treatment dramatically reduced the oxidative mitochondrial capacity and the glycolytic activity of cancer cells, consistent with previous studies linking DNA-PK expression to the proper maintenance of mitochondrial DNA integrity and copy number. Using a luciferase-based assay, we observed that doxycycline treatment quantitatively reduces the anti-oxidant response (NRF1/2) and effectively blocks signaling along multiple independent pathways normally associated with stem cells, including STAT1/3, Sonic Hedgehog (Shh), Notch, WNT and TGF-beta signaling. In conclusion, we propose that the efficacy of doxycycline as a DNA-PK inhibitor should be tested in Phase-II clinical trials, in combination with radio-therapy. Doxycycline has excellent pharmacokinetics, with nearly 100% oral absorption and a long serum half-life (18-22 hours), at a standard dose of 200-mg per day. In further support of this idea, we show that doxycycline effectively inhibits the mammosphere-forming activity of primary breast cancer samples, derived from metastatic disease sites (pleural effusions or ascites fluid). Our results also have possible implications for the radio-therapy of brain tumors and/or brain metastases, as doxycycline is known to effectively cross the blood-brain barrier. Further studies will be needed to determine if other tetracycline family members also confer radio-sensitivity.


Assuntos
Neoplasias da Mama/radioterapia , Proteína Quinase Ativada por DNA/antagonistas & inibidores , Doxiciclina/farmacologia , Células-Tronco Neoplásicas/efeitos da radiação , Radiossensibilizantes/farmacologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Proteína Quinase Ativada por DNA/genética , Proteína Quinase Ativada por DNA/metabolismo , Regulação para Baixo/efeitos dos fármacos , Feminino , Humanos , Células MCF-7 , Células-Tronco Neoplásicas/enzimologia , Células-Tronco Neoplásicas/patologia , Proteômica/métodos
19.
Oncotarget ; 6(17): 14777-95, 2015 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-26087310

RESUMO

Here, we show that new mitochondrial biogenesis is required for the anchorage independent survival and propagation of cancer stem-like cells (CSCs). More specifically, we used the drug XCT790 as an investigational tool, as it functions as a specific inhibitor of the ERRα-PGC1 signaling pathway, which governs mitochondrial biogenesis. Interestingly, our results directly demonstrate that XCT790 efficiently blocks both the survival and propagation of tumor initiating stem-like cells (TICs), using the MCF7 cell line as a model system. Mechanistically, we show that XCT790 suppresses the activity of several independent signaling pathways that are normally required for the survival of CSCs, such as Sonic hedgehog, TGFß-SMAD, STAT3, and Wnt signaling. We also show that XCT790 markedly reduces oxidative mitochondrial metabolism (OXPHOS) and that XCT790-mediated inhibition of CSC propagation can be prevented or reversed by Acetyl-L-Carnitine (ALCAR), a mitochondrial fuel. Consistent with our findings, over-expression of ERRα significantly enhances the efficiency of mammosphere formation, which can be blocked by treatment with mitochondrial inhibitors. Similarly, mammosphere formation augmented by FOXM1, a downstream target of Wnt/ß-catenin signaling, can also be blocked by treatment with three different classes of mitochondrial inhibitors (XCT790, oligomycin A, or doxycycline). In this context, our unbiased proteomics analysis reveals that FOXM1 drives the expression of >90 protein targets associated with mitochondrial biogenesis, glycolysis, the EMT and protein synthesis in MCF7 cells, processes which are characteristic of an anabolic CSC phenotype. Finally, doxycycline is an FDA-approved antibiotic, which is very well-tolerated in patients. As such, doxycycline could be re-purposed clinically as a 'safe' mitochondrial inhibitor, to target FOXM1 and mitochondrial biogenesis in CSCs, to prevent tumor recurrence and distant metastasis, thereby avoiding patient relapse.


Assuntos
Proliferação de Células/fisiologia , Mitocôndrias/metabolismo , Células-Tronco Neoplásicas/metabolismo , Biogênese de Organelas , Acetilcarnitina/farmacologia , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Adesão Celular/efeitos dos fármacos , Adesão Celular/fisiologia , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Cromatografia Líquida , Doxiciclina/farmacologia , Proteína Forkhead Box M1 , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Glicólise/efeitos dos fármacos , Humanos , Células MCF-7 , Mitocôndrias/efeitos dos fármacos , Células-Tronco Neoplásicas/efeitos dos fármacos , Nitrilas/farmacologia , Oligomicinas/farmacologia , Fosforilação Oxidativa/efeitos dos fármacos , Proteômica/métodos , Receptores de Estrogênio/genética , Receptores de Estrogênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Esferoides Celulares/efeitos dos fármacos , Esferoides Celulares/metabolismo , Espectrometria de Massas em Tandem , Tiazóis/farmacologia , Via de Sinalização Wnt/efeitos dos fármacos , Receptor ERRalfa Relacionado ao Estrogênio
20.
Nat Rev Cancer ; 15(4): 225-37, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25801618

RESUMO

It has been over 20 years since the discovery that caveolar lipid rafts function as signalling organelles. Lipid rafts create plasma membrane heterogeneity, and caveolae are the most extensively studied subset of lipid rafts. A newly emerging paradigm is that changes in caveolae also generate tumour metabolic heterogeneity. Altered caveolae create a catabolic tumour microenvironment, which supports oxidative mitochondrial metabolism in cancer cells and which contributes to dismal survival rates for cancer patients. In this Review, we discuss the role of caveolae in tumour progression, with a special emphasis on their metabolic and cell signalling effects, and their capacity to transform the tumour microenvironment.


Assuntos
Cavéolas/metabolismo , Neoplasias/metabolismo , Neoplasias/patologia , Transdução de Sinais , Animais , Humanos , Microdomínios da Membrana
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