RESUMEN
Reducing premature mortality associated with age-related chronic diseases, such as cancer and cardiovascular disease, is an urgent priority. We report early results using genomics in combination with advanced imaging and other clinical testing to proactively screen for age-related chronic disease risk among adults. We enrolled active, symptom-free adults in a study of screening for age-related chronic diseases associated with premature mortality. In addition to personal and family medical history and other clinical testing, we obtained whole-genome sequencing (WGS), noncontrast whole-body MRI, dual-energy X-ray absorptiometry (DXA), global metabolomics, a new blood test for prediabetes (Quantose IR), echocardiography (ECHO), ECG, and cardiac rhythm monitoring to identify age-related chronic disease risks. Precision medicine screening using WGS and advanced imaging along with other testing among active, symptom-free adults identified a broad set of complementary age-related chronic disease risks associated with premature mortality and strengthened WGS variant interpretation. This and other similarly designed screening approaches anchored by WGS and advanced imaging may have the potential to extend healthy life among active adults through improved prevention and early detection of age-related chronic diseases (and their risk factors) associated with premature mortality.
Asunto(s)
Enfermedad/genética , Predisposición Genética a la Enfermedad , Procesamiento de Imagen Asistido por Computador/métodos , Mutación , Medicina de Precisión/métodos , Secuenciación Completa del Genoma/métodos , Adulto , Anciano , Anciano de 80 o más Años , Enfermedades Cardiovasculares/diagnóstico por imagen , Enfermedades Cardiovasculares/genética , Enfermedades Cardiovasculares/patología , Enfermedad/clasificación , Femenino , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Masculino , Persona de Mediana Edad , Neoplasias/diagnóstico por imagen , Neoplasias/genética , Neoplasias/patología , Enfermedades del Sistema Nervioso/diagnóstico por imagen , Enfermedades del Sistema Nervioso/genética , Enfermedades del Sistema Nervioso/patología , Medición de Riesgo , Análisis de Secuencia de ARN , Adulto JovenRESUMEN
Cell populations with differing proliferative, stem-like and tumorigenic states co-exist in most tumors and especially malignant gliomas. Whether metabolic variations can drive this heterogeneity by controlling dynamic changes in cell states is unknown. Metabolite profiling of human adult glioblastoma stem-like cells upon loss of their tumorigenicity revealed a switch in the catabolism of the GABA neurotransmitter toward enhanced production and secretion of its by-product GHB (4-hydroxybutyrate). This switch was driven by succinic semialdehyde dehydrogenase (SSADH) downregulation. Enhancing GHB levels via SSADH downregulation or GHB supplementation triggered cell conversion into a less aggressive phenotypic state. GHB affected adult glioblastoma cells with varying molecular profiles, along with cells from pediatric pontine gliomas. In all cell types, GHB acted by inhibiting α-ketoglutarate-dependent Ten-eleven Translocations (TET) activity, resulting in decreased levels of the 5-hydroxymethylcytosine epigenetic mark. In patients, low SSADH expression was correlated with high GHB/α-ketoglutarate ratios, and distinguished weakly proliferative/differentiated glioblastoma territories from proliferative/non-differentiated territories. Our findings support an active participation of metabolic variations in the genesis of tumor heterogeneity.
Asunto(s)
Neoplasias Encefálicas/metabolismo , Carcinogénesis/metabolismo , Glioma/metabolismo , Hidroxibutiratos/metabolismo , Células Madre Neoplásicas/metabolismo , Ácido gamma-Aminobutírico/metabolismo , Anciano , Animales , Encéfalo/metabolismo , Encéfalo/patología , Encéfalo/cirugía , Neoplasias Encefálicas/patología , Neoplasias Encefálicas/cirugía , Carcinogénesis/patología , Muerte Celular/fisiología , Proliferación Celular/fisiología , Niño , Preescolar , Femenino , Glioma/patología , Glioma/cirugía , Humanos , Masculino , Ratones Desnudos , Persona de Mediana Edad , Trasplante de Neoplasias , Células Madre Neoplásicas/patología , Succionato-Semialdehído Deshidrogenasa/metabolismoRESUMEN
HOXB13 is a member of the homeodomain family of sequence-specific transcription factors and, together with the androgen receptor (AR), plays a critical role in the normal development of the prostate gland. We demonstrate here that, in prostate cancer cells, HOXB13 is a key determinant of the response to androgens. Specifically, it was determined that HOXB13 interacts with the DNA-binding domain of AR and inhibits the transcription of genes that contain an androgen-response element (ARE). In contrast, the AR:HOXB13 complex confers androgen responsiveness to promoters that contain a specific HOXB13-response element. Further, HOXB13 and AR synergize to enhance the transcription of genes that contain a HOX element juxtaposed to an ARE. The profound effects of HOXB13 knockdown on androgen-regulated proliferation, migration, and lipogenesis in prostate cancer cells highlight the importance of the observed changes in gene expression.
Asunto(s)
Proteínas de Homeodominio/metabolismo , Metribolona/farmacología , Receptores Androgénicos/metabolismo , Secuencia de Aminoácidos , Western Blotting , Línea Celular , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Inmunoprecipitación de Cromatina , Análisis por Conglomerados , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Perfilación de la Expresión Génica , Regulación de la Expresión Génica/efectos de los fármacos , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Proteínas de Homeodominio/genética , Humanos , Metabolismo de los Lípidos/efectos de los fármacos , Masculino , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/patología , Unión Proteica/efectos de los fármacos , ARN Interferente Pequeño/genética , Receptores Androgénicos/genética , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Elementos de Respuesta/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , TransfecciónRESUMEN
We show that HEXIM1 (hexamethylene bis-acetamide inducible 1) functions as an AR (androgen receptor) co-repressor as it physically interacts with the AR and is required for the ability of anti-androgens to inhibit androgen-induced target gene expression and cell proliferation. Oncomine™ database and IHC (immunohistochemistry) analyses of human prostate tissues revealed that expression of HEXIM1 mRNA and protein are down-regulated during the development and progression of prostate cancer. Enforced down-regulation of HEXIM1 in parental hormone-dependent LNCaP cells results in resistance to the inhibitory action of anti-androgens. Conversely, ectopic expression of HEXIM1 in the CRPC (castration-resistant prostate cancer) cell line, C4-2, enhances their sensitivity to the repressive effects of the anti-androgen bicalutamide. Novel insight into the mechanistic basis for HEXIM1 inhibition of AR activity is provided by the present studies showing that HEXIM1 induces expression of the histone demethylase KDM5B (lysine-specific demethylase 5B) and inhibits histone methylation, resulting in the inhibition of FOXA1 (forkhead box A1) licensing activity. This is a new mechanism of action attributed to HEXIM1, and distinct from what has been reported so far to be involved in HEXIM1 regulation of other nuclear hormone receptors, including the oestrogen receptor.
Asunto(s)
Antagonistas de Andrógenos/farmacología , Neoplasias de la Próstata/metabolismo , Proteínas de Unión al ARN/metabolismo , Receptores Androgénicos/metabolismo , Anilidas/farmacología , Línea Celular Tumoral , Elementos de Facilitación Genéticos , Células Epiteliales/metabolismo , Expresión Génica/efectos de los fármacos , Histonas/metabolismo , Humanos , Histona Demetilasas con Dominio de Jumonji/metabolismo , Masculino , Metribolona/farmacología , Nitrilos/farmacología , Proteínas Nucleares/metabolismo , Próstata/metabolismo , Neoplasias de la Próstata Resistentes a la Castración/metabolismo , Transporte de Proteínas , Proteínas Represoras/metabolismo , Compuestos de Tosilo/farmacología , Factores de Transcripción , Factores de Elongación Transcripcional/metabolismoRESUMEN
The impact of ligand binding on nuclear receptor (NR) structure and the ability of target cells to distinguish between different receptor-ligand complexes are key determinants of the pharmacological activity of NR ligands. However, until relatively recently, these mechanistic insights have not been used in a prospective manner to develop screens for NR modulators with specific therapeutic activities. Driven by the need for unique androgen receptor (AR) antagonists that retain activity in hormone-refractory prostate cancer, we developed and applied a conformation-based screen to identify AR antagonists that were mechanistically distinct from existing drugs of this class. Two molecules were identified by using this approach, D36 and D80, which interact with AR in a unique manner and allosterically inhibit AR agonist activity. Unlike the clinically important antiandrogens, casodex and hydroxyflutamide, both D36 and D80 block androgen action in cellular models of hormone-refractory prostate cancer. Mechanistically, these compounds further distinguish themselves from classical AR antagonists in that they do not promote AR nuclear translocation and quantitatively inhibit the association of AR with DNA even under conditions of overexpression. Although the therapeutic potential of these antiandrogens is apparent, it is the demonstration that it is possible, to modulate the interaction of cofactors with agonist-activated AR, using second-site modulators, that has the greatest potential with respect to the therapeutic exploitation of AR and other NRs.
Asunto(s)
Antagonistas de Receptores Androgénicos , Antineoplásicos/farmacología , Neoplasias de la Próstata/patología , Regulación Alostérica/efectos de los fármacos , Animales , Antineoplásicos/química , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Resistencia a Antineoplásicos/efectos de los fármacos , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Ligandos , Masculino , Ratones , Conformación Molecular , Neoplasias de la Próstata/genética , Receptores Androgénicos , Transcripción Genética/efectos de los fármacosRESUMEN
Background: Polyamine levels are intricately controlled by biosynthetic, catabolic enzymes and antizymes. The complexity suggests that minute alterations in levels lead to profound abnormalities. We described the therapeutic course for a rare syndrome diagnosed by whole exome sequencing caused by gain-of-function variants in the C-terminus of ornithine decarboxylase (ODC), characterized by neurological deficits and alopecia. Methods: N-acetylputrescine levels with other metabolites were measured using ultra-performance liquid chromatography paired with mass spectrometry and Z-scores established against a reference cohort of 866 children. Results: From previous studies and metabolic profiles, eflornithine was identified as potentially beneficial with therapy initiated on FDA approval. Eflornithine normalized polyamine levels without disrupting other pathways. She demonstrated remarkable improvement in both neurological symptoms and cortical architecture. She gained fine motor skills with the capacity to feed herself and sit with support. Conclusions: This work highlights the strategy of repurposing drugs to treat a rare disease. Funding: No external funding was received for this work.
Asunto(s)
Transportadores de Ácidos Dicarboxílicos/genética , Reposicionamiento de Medicamentos , Eflornitina/farmacología , Eflornitina/uso terapéutico , Mutación con Ganancia de Función/genética , Proteínas de Transporte de Membrana Mitocondrial/genética , Alopecia , Preescolar , Transportadores de Ácidos Dicarboxílicos/química , Variación Genética , Humanos , Proteínas de Transporte de Membrana Mitocondrial/química , Ornitina Descarboxilasa/genética , Poliaminas , Putrescina/análogos & derivados , Enfermedades Raras/tratamiento farmacológico , Enfermedades Raras/genética , Secuenciación del ExomaRESUMEN
One subclass of antiestrogens, the selective estrogen receptor down-regulators (SERDs), have received considerable attention of late as they competitively inhibit estrogen binding and induce a rapid, proteasome-dependent degradation of the receptor. Contained within this class of molecules is the steroidal antiestrogen ICI182,780 (faslodex), recently approved for the treatment of metastatic cancer, and GW5638/DPC974, a SERD that is currently being evaluated in the clinic. Given that mechanistic differences between different selective estrogen receptor modulators have been translated into important clinical profiles, it was of interest to determine if the SERD subclass of ligands were likewise functionally or mechanistically distinguishable. In this study, we show that although the steroidal and nonsteroidal SERDs target ERalpha for degradation, the underlying mechanism(s) are different. Of note was the identification of a specific protein-protein interaction surface presented on ERalpha in the presence of the ICI182,780-activated receptor which is required for degradation. Interestingly, this surface is also presented on ERalpha in the presence of RU58,668, a SERD that is chemically distinct from ICI182,780. This surface is not required for GW5638-mediated degradation, and thus, this SERD seems to affect ERalpha down-regulation by a different mechanism. These data suggest that sequencing of therapies using drugs of this class is likely to be possible. Finally, because of the unmet need for orally active SERDS that function similarly to ICI182,780, we have used the insights from these mechanistic studies to develop and validate a high-throughput screen for compounds of this class with improved pharmaceutical properties.
Asunto(s)
Cinamatos/farmacología , Estradiol/análogos & derivados , Moduladores de los Receptores de Estrógeno/farmacología , Receptor alfa de Estrógeno/metabolismo , Estilbenos/farmacología , Secuencia de Aminoácidos , Sitios de Unión , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Cinamatos/química , Estradiol/química , Estradiol/farmacología , Antagonistas de Estrógenos/química , Antagonistas de Estrógenos/farmacología , Moduladores de los Receptores de Estrógeno/química , Receptor alfa de Estrógeno/química , Fulvestrant , Células HeLa , Humanos , Datos de Secuencia Molecular , Conformación Proteica/efectos de los fármacos , Estilbenos/química , Fracciones Subcelulares/efectos de los fármacos , Fracciones Subcelulares/metabolismoRESUMEN
Metabolomics is the untargeted measurement of the metabolome, which is composed of the complement of small molecules detected in a biological sample. As such, metabolomic analysis produces a global biochemical phenotype. It is a technology that has been utilized in the research setting for over a decade. The metabolome is directly linked to and is influenced by genetics, epigenetics, environmental factors, and the microbiome-all of which affect health. Metabolomics can be applied to human clinical diagnostics and to other fields such as veterinary medicine, nutrition, exercise, physiology, agriculture/plant biochemistry, and toxicology. Applications of metabolomics in clinical testing are emerging, but several aspects of its use as a clinical test differ from applications focused on research or biomarker discovery and need to be considered for metabolomics clinical test data to have optimum impact, be meaningful, and be used responsibly. In this review, we deconstruct aspects and challenges of metabolomics for clinical testing by illustrating the significance of test design, accurate and precise data acquisition, quality control, data processing, n-of-1 comparison to a reference population, and biochemical pathway analysis. We describe how metabolomics technology is integral to defining individual biochemical phenotypes, elaborates on human health and disease, and fits within the precision medicine landscape. Finally, we conclude by outlining some future steps needed to bring metabolomics into the clinical space and to be recognized by the broader medical and regulatory fields.
Asunto(s)
Metabolómica/métodos , Técnicas de Química Analítica/métodos , Pruebas de Química Clínica/métodos , Humanos , Metaboloma , Metabolómica/normasRESUMEN
Estrogen receptor alpha (ERalpha) regulates transcription of specific genes and is believed to play a major role in breast tumorigenesis. We previously identified estrogen down regulated gene 1 (EDG1 (also known as HEXIM1)) using the C-terminus of ERalpha (E/F domain) as bait in yeast two-hybrid screenings. Here we report on the role of EDG1 as a coregulator of ERalpha transcriptional activity. We observe an interaction between EDG1 and ERalpha. EDG1 inhibits the transcriptional activity of ERalpha and this is dependent upon the C-terminus of EDG1. The C-terminus of EDG1/HEXIM1 was recently shown to inhibit the positive transcription elongation factor b (P-TEFb) by interacting with the cyclin T1 subunit. Here we show that ERalpha interacts with cyclin T1, cyclin T1 and ER co-occupancy on the promoter region of an ER target gene, and that this interaction plays an important role in ERalpha-induced gene expression. The interaction of ERalpha with cyclin T1 also allows ERalpha to compete with EDG1 for cyclin T1, and may release cyclin T1 from EDG1 repression. Conversely, increased EDG1 expression results in inhibition of cyclin T1 recruitment and ERalpha DNA binding. Our results support a novel functional interaction between ERalpha and cyclin T1 that is modulated by EDG1.
Asunto(s)
Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Ciclinas/metabolismo , Receptor alfa de Estrógeno/metabolismo , Proteínas de Unión al ARN/metabolismo , Animales , Unión Competitiva , Neoplasias de la Mama/genética , Línea Celular , Cricetinae , Ciclina T , Receptor alfa de Estrógeno/antagonistas & inhibidores , Eliminación de Gen , Humanos , Mutación/genética , Regiones Promotoras Genéticas/genética , Unión Proteica , Proteínas de Unión al ARN/química , Proteínas de Unión al ARN/genética , Proteínas Represoras/química , Proteínas Represoras/genética , Proteínas Represoras/metabolismo , Factores de Transcripción , Transcripción Genética/genéticaRESUMEN
Lifetime exposure to estrogens is a major risk factor in breast cancer, but the mechanism for this action is not fully defined. To better determine this mechanism, the activation domain of estrogen receptor (ER) alpha was used in yeast two-hybrid screenings. These screenings resulted in the identification of a novel antiproliferative protein, estrogen down-regulated gene 1 (EDG1), of which the mRNA and protein were shown to be down-regulated directly by estrogens. Our studies additionally suggested an important role for EDG1 in ER alpha-mediated breast cancer development. Analysis of 43 invasive breast cancer samples and 40 adjacent normal breast samples demonstrated EDG1 protein levels to be significantly higher in normal breast epithelial tissue as compared with breast epithelial tumor tissue. EDG1 expression levels were also correlated with the proliferation activity and ER alpha status of the tumors to examine the prognostic value of EDG1 in invasive breast tumors. EDG1 expression was more disassociated from proliferative activity as compared with ER alpha expression in tumor cells. A growth regulatory function for EDG1 is additionally indicated by studies wherein overexpression of EDG1 protein in breast cells resulted in decreased cell proliferation and decreased anchorage-independent growth. Conversely, inhibiting EDG1 expression in breast cells resulted in increased breast cell growth. Thus, we have identified a novel growth inhibitor that is down-regulated by estrogens and colocalizes with ER alpha in breast tissue. These studies support a role for EDG1 in breast cancer.
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Neoplasias de la Mama/etiología , Mama/efectos de los fármacos , Estrógenos/farmacología , Inhibidores de Crecimiento/aislamiento & purificación , Receptores de Estrógenos/aislamiento & purificación , Acetamidas/farmacología , Mama/química , Mama/citología , División Celular/efectos de los fármacos , Regulación hacia Abajo , Células Epiteliales/química , Receptor alfa de Estrógeno , Femenino , Inhibidores de Crecimiento/análisis , Inhibidores de Crecimiento/farmacología , Humanos , Antígeno Ki-67/análisis , Pérdida de Heterocigocidad , Receptores de Estrógenos/análisis , Receptores de Lisoesfingolípidos , Receptores de Esfingosina-1-FosfatoRESUMEN
Antiestrogens have found widespread use in the treatment of breast cancer (reviewed in ref. 1). There is also interest in the use of tamoxifen as a preventive agent for breast cancer. However, the mechanism for the antitumor effects of antiestrogens is relatively unknown. For the most part it is thought that the basis for the anticancer action of antiestrogens is the inhibition of estradiol (E2)-stimulated tumor growth. We have observed however that antiestrogens can activate detoxifying enzymes, like quinone reductase (NQO1), which protect cells against the toxic and tumor-promoting effects of carcinogens (2). Studies characterizing the molecular mechanisms behind the regulation of NQO1 by the Estrogen Receptor (ER) support an important role of the ER in pathways regulating antioxidant defenses. Moreover these findings represent a novel mechanism through which antiestrogens function. The activation of NQO1 may contribute to the beneficial anticancer and antioxidant activity of antiestrogens in breast cancer and possibly other estrogen target tissues. It is possible that the basis for some of the anticancer action of antiestrogens is that the induction of NQO1 inhibits the genotoxic effects induced by the oxidative metabolism of estrogens.
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Antineoplásicos/uso terapéutico , Neoplasias de la Mama/tratamiento farmacológico , Antagonistas de Estrógenos/uso terapéutico , NAD(P)H Deshidrogenasa (Quinona)/metabolismo , Receptores de Estrógenos/fisiología , Tamoxifeno/uso terapéutico , Neoplasias de la Mama/inducido químicamente , Estrógenos , Humanos , Células Tumorales CultivadasRESUMEN
Bladder cancer (BCa) is a common malignancy worldwide and has a high probability of recurrence after initial diagnosis and treatment. As a result, recurrent surveillance, primarily involving repeated cystoscopies, is a critical component of post diagnosis patient management. Since cystoscopy is invasive, expensive and a possible deterrent to patient compliance with regular follow-up screening, new non-invasive technologies to aid in the detection of recurrent and/or primary bladder cancer are strongly needed. In this study, mass spectrometry based metabolomics was employed to identify biochemical signatures in human urine that differentiate bladder cancer from non-cancer controls. Over 1000 distinct compounds were measured including 587 named compounds of known chemical identity. Initial biomarker identification was conducted using a 332 subject sample set of retrospective urine samples (cohort 1), which included 66 BCa positive samples. A set of 25 candidate biomarkers was selected based on statistical significance, fold difference and metabolic pathway coverage. The 25 candidate biomarkers were tested against an independent urine sample set (cohort 2) using random forest analysis, with palmitoyl sphingomyelin, lactate, adenosine and succinate providing the strongest predictive power for differentiating cohort 2 cancer from non-cancer urines. Cohort 2 metabolite profiling revealed additional metabolites, including arachidonate, that were higher in cohort 2 cancer vs. non-cancer controls, but were below quantitation limits in the cohort 1 profiling. Metabolites related to lipid metabolism may be especially interesting biomarkers. The results suggest that urine metabolites may provide a much needed non-invasive adjunct diagnostic to cystoscopy for detection of bladder cancer and recurrent disease management.
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Biomarcadores de Tumor/metabolismo , Biomarcadores de Tumor/orina , Metabolómica , Neoplasias de la Vejiga Urinaria/metabolismo , Neoplasias de la Vejiga Urinaria/orina , Anciano , Estudios de Casos y Controles , Estudios de Cohortes , Femenino , Humanos , Masculino , Persona de Mediana Edad , Pronóstico , Neoplasias de la Vejiga Urinaria/diagnósticoRESUMEN
Pulmonary arterial hypertension (PAH) is a progressive and fatal disease of the lung vasculature for which the molecular etiologies are unclear. Specific metabolic alterations have been identified in animal models and in PAH patients, though existing data focus mainly on abnormalities of glucose homeostasis. We hypothesized that analysis of the entire metabolome in PAH would reveal multiple other metabolic changes relevant to disease pathogenesis and possible treatment. Layered transcriptomic and metabolomic analyses of human pulmonary microvascular endothelial cells (hPMVEC) expressing two different disease-causing mutations in the bone morphogenetic protein receptor type 2 (BMPR2) confirmed previously described increases in aerobic glycolysis but also uncovered significant upregulation of the pentose phosphate pathway, increases in nucleotide salvage and polyamine biosynthesis pathways, decreases in carnitine and fatty acid oxidation pathways, and major impairment of the tricarboxylic acid (TCA) cycle and failure of anaplerosis. As a proof of principle, we focused on the TCA cycle, predicting that isocitrate dehydrogenase (IDH) activity would be altered in PAH, and then demonstrating increased IDH activity not only in cultured hPMVEC expressing mutant BMPR2 but also in the serum of PAH patients. These results suggest that widespread metabolic changes are an important part of PAH pathogenesis, and that simultaneous identification and targeting of the multiple involved pathways may be a more fruitful therapeutic approach than targeting of any one individual pathway.
RESUMEN
While patients with advanced prostate cancer initially respond favorably to androgen ablation therapy, most experience a relapse of the disease within 1-2 years. Although hormone-refractory disease is unresponsive to androgen-deprivation, androgen receptor (AR)-regulated signaling pathways remain active and are necessary for cancer progression. Thus, both AR itself and the processes downstream of the receptor remain viable targets for therapeutic intervention. Microarray analysis of multiple clinical cohorts showed that the serine/threonine kinase Ca2+/calmodulin-dependent protein kinase kinase ß (CaMKKß) is both highly expressed in the prostate and further elevated in prostate cancers. Using cellular models of prostate cancer, we have determined that androgens (a) directly increase the expression of a CaMKKß splice variant and (b) increase functional CaMKKß protein levels as determined by the phosphorylation of both CaMKI and AMP-activated protein kinase (AMPK), two of CaMKKß's primary substrates. Importantly, inhibition of the CaMKKß-AMPK, but not CaMKI, signaling axis in prostate cancer cells by pharmacological inhibitors or siRNA-mediated knockdown blocks androgen-mediated migration and invasion. Conversely, overexpression of CaMKKß alone leads to both increased AMPK phosphorylation and cell migration. Given the key roles of CaMKKß and AMPK in the biology of prostate cancer cells, we propose that these enzymes are potential therapeutic targets in prostate cancer.
Asunto(s)
Andrógenos/farmacología , Quinasa de la Proteína Quinasa Dependiente de Calcio-Calmodulina/metabolismo , Movimiento Celular/fisiología , Neoplasias de la Próstata/enzimología , Neoplasias de la Próstata/patología , Proteínas Quinasas/metabolismo , Quinasas de la Proteína-Quinasa Activada por el AMP , Andrógenos/metabolismo , Quinasa de la Proteína Quinasa Dependiente de Calcio-Calmodulina/biosíntesis , Quinasa de la Proteína Quinasa Dependiente de Calcio-Calmodulina/genética , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Humanos , Isoenzimas , Masculino , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/metabolismo , Isoformas de Proteínas , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Receptores Androgénicos/metabolismo , Transducción de Señal/efectos de los fármacos , Regulación hacia ArribaRESUMEN
Both pro- and antimitogenic activities have been ascribed to progesterone receptor (PR) agonists and antagonists in breast cancer cells; however, the transcriptional responses that underlie these paradoxical functions are not apparent. Using nontransformed, normal human mammary epithelial cells engineered to express PR and standard microarray technology, we defined 2370 genes that were significantly regulated by the PR agonist R5020. Gene ontology (GO) analysis revealed that GO terms involved in inflammation and nuclear factor-κB (NF-κB) signaling were among the most significantly regulated. Interestingly, on those NF-κB responsive genes that were inhibited by agonist-activated PR, antagonists either 1) mimicked the actions of agonists or 2) reversed the inhibitory actions of agonists. This difference in pharmacological response could be attributed to the fact that although agonist- and antagonist-activated PR is recruited to NF-κB-responsive promoters, the physical presence of PR tethered to the promoter of some genes is sufficient for transcriptional inhibition, whereas on others, an agonist-activated PR conformation is required for inhibition of NF-κB signaling. Importantly, the actions of PR on the latter class of genes were reversed by an activation function-2-inhibiting, LXXLL-containing peptide. Consideration of the relative activities of these distinct antiinflammatory pathways in breast cancer may be instructive with respect to the likely therapeutic activity of PR agonists or antagonists in the treatment of breast cancer.
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Neoplasias Inflamatorias de la Mama/patología , Receptores de Progesterona/fisiología , Secuencia de Aminoácidos , Línea Celular , Línea Celular Tumoral , Elementos de Facilitación Genéticos , Células Epiteliales/metabolismo , Células Epiteliales/patología , Femenino , Gonanos/farmacología , Humanos , Neoplasias Inflamatorias de la Mama/genética , Neoplasias Inflamatorias de la Mama/metabolismo , Interleucina-1beta/antagonistas & inhibidores , Interleucina-1beta/metabolismo , Interleucina-1beta/farmacología , Mifepristona/farmacología , Datos de Secuencia Molecular , FN-kappa B/antagonistas & inhibidores , FN-kappa B/genética , FN-kappa B/metabolismo , Progesterona/genética , Progesterona/metabolismo , Progestinas/metabolismo , Regiones Promotoras Genéticas , Isoformas de Proteínas , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Receptores de Progesterona/agonistas , Receptores de Progesterona/antagonistas & inhibidores , Transducción de Señal , Activación Transcripcional , Translocación GenéticaRESUMEN
Selective estrogen receptor modulators (SERMs), such as tamoxifen (TAM), have been used extensively for the treatment and prevention of breast cancer and other pathologies associated with aberrant estrogen receptor (ER) signaling. These compounds exhibit cell-selective agonist/antagonist activities as a consequence of their ability to induce different conformational changes in ER, thereby enabling it to recruit functionally distinct transcriptional coregulators. However, the observation that SERMs can also regulate aspects of calcium signaling and apoptosis in an ER-independent manner in some systems suggests that some of the activity of drugs within this class may also arise as a consequence of their ability to interact with targets other than ER. In this study, we demonstrate that 4-hydroxy-TAM (4OHT), an active metabolite of TAM, directly binds to and modulates the transcriptional activity of the aryl hydrocarbon receptor (AHR). Of specific interest was the observation, that in the absence of ER, 4OHT can induce the expression of AHR target genes involved in estradiol metabolism, cellular proliferation, and metastasis in cellular models of breast cancer. The potential role for AHR in SERM pharmacology was further underscored by the ability of 4OHT to suppress osteoclast differentiation in vitro in part through AHR. Cumulatively, these findings provide evidence that it is necessary to reevaluate the relative roles of ER and AHR in manifesting the pharmacological actions and therapeutic efficacy of TAM and other SERMs.
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Receptores de Hidrocarburo de Aril/fisiología , Receptores de Estrógenos/fisiología , Moduladores Selectivos de los Receptores de Estrógeno/farmacología , Tamoxifeno/análogos & derivados , Animales , Translocador Nuclear del Receptor de Aril Hidrocarburo/genética , Translocador Nuclear del Receptor de Aril Hidrocarburo/metabolismo , Neoplasias de la Mama/metabolismo , Diferenciación Celular/efectos de los fármacos , Línea Celular , Línea Celular Tumoral , Inmunoprecipitación de Cromatina , Relación Dosis-Respuesta a Droga , Femenino , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Humanos , Masculino , Ratones , Osteoclastos/metabolismo , Osteoclastos/fisiología , Receptores de Hidrocarburo de Aril/genética , Receptores de Hidrocarburo de Aril/metabolismo , Receptores de Estrógenos/deficiencia , Receptores de Estrógenos/genética , Receptores de Estrógenos/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Moduladores Selectivos de los Receptores de Estrógeno/metabolismo , Tamoxifeno/metabolismo , Tamoxifeno/farmacologíaRESUMEN
Advanced prostate cancers preferentially metastasize to bone, suggesting that this tissue produces factors that provide a suitable microenvironment for prostate cancer cells. Recently, it has become clear that even in antiandrogen-resistant cancers, the androgen receptor (AR)-signaling axis is required for prostate cancer progression. Therefore, we hypothesized that AR may be involved in the regulation of pathways that are responsible for the homing of prostate cancer cells to select microenvironments. In support of this hypothesis, we have determined that chemokine (C-X-C motif) receptor 4 (CXCR4), the receptor for the chemokine CXCL12, is up-regulated in prostate cancer cells in response to androgens. Given that the levels of CXCL12 are elevated at sites of known prostate cancer metastases such as bone, these results suggest that androgens may influence prostate cancer metastasis. Specifically, we demonstrate that androgens increase the levels of both CXCR4 mRNA and functional protein in LNCaP prostate cancer cells. Importantly, androgens enhanced the migration of LNCaP cells toward a CXCL12 gradient, an effect that could be blocked by the specific CXCR4 antagonist AMD3100. Interestingly, CXCR4 is not directly regulated by androgens but rather is positively up-regulated by Krüppel-like factor 5 (KLF5), a transcription factor that we have shown to be an early, direct target of AR. Further, KLF5 is both required and sufficient for androgen-mediated CXCR4 expression and migration toward CXCL12. Taken together, these findings demonstrate that AR can utilize the CXCL12/CXCR4 axis through induction of KLF5 expression to promote prostate cancer progression and highlight the potential utility of CXCR4 antagonists as prostate cancer therapeutics.