RESUMEN
Storage of excess calories as triglycerides is central to obesity and its associated disorders. Glycerol-3-phosphate acyltransferases (GPATs) catalyze the initial step in acylglyceride syntheses, including triglyceride synthesis. We utilized a novel small-molecule GPAT inhibitor, FSG67, to investigate metabolic consequences of systemic pharmacological GPAT inhibition in lean and diet-induced obese (DIO) mice. FSG67 administered intraperitoneally decreased body weight and energy intake, without producing conditioned taste aversion. Daily FSG67 (5 mg/kg, 15.3 µmol/kg) produced gradual 12% weight loss in DIO mice beyond that due to transient 9- to 10-day hypophagia (6% weight loss in pair-fed controls). Continued FSG67 maintained the weight loss despite return to baseline energy intake. Weight was lost specifically from fat mass. Indirect calorimetry showed partial protection by FSG67 against decreased rates of oxygen consumption seen with hypophagia. Despite low respiratory exchange ratio due to a high-fat diet, FSG67-treated mice showed further decreased respiratory exchange ratio, beyond pair-fed controls, indicating enhanced fat oxidation. Chronic FSG67 increased glucose tolerance and insulin sensitivity in DIO mice. Chronic FSG67 decreased gene expression for lipogenic enzymes in white adipose tissue and liver and decreased lipid accumulation in white adipose, brown adipose, and liver tissues without signs of damage. RT-PCR showed decreased gene expression for orexigenic hypothalamic neuropeptides AgRP or NPY after acute and chronic systemic FSG67. FSG67 given intracerebroventricularly (100 and 320 nmol icv) produced 24-h weight loss and feeding suppression, indicating contributions from direct central nervous system sites of action. Together, these data point to GPAT as a new potential therapeutic target for the management of obesity and its comorbidities.
Asunto(s)
Adiposidad/fisiología , Ingestión de Alimentos/fisiología , Inhibidores Enzimáticos/farmacología , Glicerol-3-Fosfato O-Aciltransferasa/antagonistas & inhibidores , Resistencia a la Insulina/fisiología , Obesidad/fisiopatología , Adiposidad/efectos de los fármacos , Proteína Relacionada con Agouti/metabolismo , Animales , Grasas de la Dieta/efectos adversos , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Ingestión de Alimentos/efectos de los fármacos , Hígado Graso/metabolismo , Hígado Graso/fisiopatología , Glicerol-3-Fosfato O-Aciltransferasa/fisiología , Ratones , Ratones Endogámicos , Mitocondrias Hepáticas/efectos de los fármacos , Mitocondrias Hepáticas/enzimología , Neuropéptido Y/metabolismo , Obesidad/etiología , Obesidad/metabolismo , Consumo de Oxígeno/efectos de los fármacos , Consumo de Oxígeno/fisiología , Delgadez/metabolismo , Delgadez/fisiopatología , Triglicéridos/metabolismoRESUMEN
Fatty acid synthase (FAS), the enzyme responsible for the de novo synthesis of fatty acids, is highly expressed in ovarian cancers and most common human carcinomas. Inhibition of FAS and activation of AMP-activated protein kinase (AMPK) have been shown to be cytotoxic to human cancer cells in vitro and in vivo. In this report, we explore the cytotoxic mechanism of action of FAS inhibition and show that C93, a synthetic FAS inhibitor, increases the AMP/ATP ratio, activating AMPK in SKOV3 human ovarian cancer cells, which leads to cytotoxicity. As a physiologic consequence of AMPK activation, acetyl-CoA carboxylase (ACC), the rate-limiting enzyme of fatty acid synthesis, was phosphorylated and inhibited whereas glucose oxidation was increased. Despite these attempts to conserve energy, the AMP/ATP ratio increased with worsening cellular redox status. Pretreatment of SKOV3 cells with compound C, an AMPK inhibitor, substantially rescued the cells from C93 cytotoxicity, indicating its dependence on AMPK activation. 5-(Tetradecyloxy)-2-furoic acid, an ACC inhibitor, did not activate AMPK despite inhibiting fatty acid synthesis pathway activity and was not significantly cytotoxic to SKOV3 cells. This indicates that substrate accumulation from FAS inhibition triggering AMPK activation, not end-product depletion of fatty acids, is likely responsible for AMPK activation. C93 also exhibited significant antitumor activity and apoptosis against SKOV3 xenografts in athymic mice without significant weight loss or cytotoxicity to proliferating cellular compartments such as bone marrow, gastrointestinal tract, or skin. Thus, pharmacologic FAS inhibition selectively activates AMPK in ovarian cancer cells, inducing cytotoxicity while sparing most normal human tissues from the pleiotropic effects of AMPK activation.
Asunto(s)
Inhibidores Enzimáticos/farmacología , Ácido Graso Sintasas/antagonistas & inhibidores , Complejos Multienzimáticos/metabolismo , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/enzimología , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Quinasas Activadas por AMP , Secuencia de Aminoácidos , Animales , Línea Celular Tumoral , Activación Enzimática , Ácidos Grasos/metabolismo , Femenino , Furanos/farmacología , Glucosa/metabolismo , Humanos , Ratones , Ratones Endogámicos BALB C , Datos de Secuencia Molecular , NAD/metabolismo , Neoplasias Ováricas/metabolismo , Oxidación-Reducción , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Activation of fatty acid synthase (FAS) expression and fatty acid synthesis is a common event in human breast cancer. Sterol regulatory element binding proteins (SREBPs) are a family of transcription factors that regulate genes involved in lipid metabolism, including FAS. SREBP-1c expression is induced in liver and adipose tissue by insulin and by fasting/refeeding and is critical for nutritional regulation of lipogenic gene expression. In contrast, upregulation of fatty acid metabolism during in vitro transformation of human mammary epithelial cells and in breast cancer cells was driven by increased MAP kinase and PI 3-kinase signaling, which increased SREBP-1 levels. SREBP-1a was more abundant than SREBP-1c in many proliferative tissues and cultured cells and was thus a candidate to regulate lipogenesis for support of membrane synthesis during cell growth. We now show that SREBP-1c and FAS mRNA were both increased by H-ras transformation of MCF-10a breast epithelial cells and were both reduced by exposure of MCF-7 breast cancer cells to the MAP kinase inhibitor, PD98059, or the PI 3-kinase inhibitor, wortmannin, while SREBP-1a and SREBP-2 showed less variation. Similarly, the mRNA levels for FAS and SREBP-1c in a panel of primary human breast cancer samples showed much greater increases than did those for SREBP-1a and SREBP-2 and were significantly correlated with each other, suggesting coordinate regulation of SREBP-1c and FAS in clinical breast cancer. We conclude that regulation of FAS expression in breast cancer is achieved through modulation of SREBP-1c, similar to the regulation in liver and adipose tissue, although the upstream regulation of liopgenesis differs in these tissues.
Asunto(s)
Neoplasias de la Mama/metabolismo , Proteínas Potenciadoras de Unión a CCAAT/fisiología , Proteínas de Unión al ADN/fisiología , Ácido Graso Sintasas/biosíntesis , Regulación Neoplásica de la Expresión Génica , Factores de Transcripción , Neoplasias de la Mama/patología , Proteínas Potenciadoras de Unión a CCAAT/biosíntesis , Proteínas Potenciadoras de Unión a CCAAT/genética , Proteínas de Unión al ADN/biosíntesis , Proteínas de Unión al ADN/genética , Factor de Crecimiento Epidérmico/metabolismo , Factor de Crecimiento Epidérmico/fisiología , Ácido Graso Sintasas/genética , Femenino , Humanos , Isoformas de Proteínas/biosíntesis , Isoformas de Proteínas/genética , Isoformas de Proteínas/fisiología , ARN Mensajero/análisis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de Señal , Proteína 1 de Unión a los Elementos Reguladores de Esteroles , Células Tumorales CultivadasRESUMEN
Activation of fatty acid synthase (FAS) expression and fatty acid synthesis is a common event in tumor cells from a variety of human cancers and is closely linked to malignant transformation and to tumor virulence in population studies of human cancer. We now show that, in contrast to nutritional regulation of lipogenesis in liver or adipose tissue, changes in fatty acid metabolism during in vitro transformation of the human mammary epithelial cell line MCF-10a are driven by increases in epidermal growth factor signaling, acting in major part through the mitogen-activated protein (MAP) kinase and phosphatidylinositol (PI) 3-kinase signaling cascades. H-ras transformation of MCF-10a cells resulted in upregulation of MAP kinase and PI 3-kinase signals, upregulation of sterol regulatory element binding protein 1 (SREBP-1) transcription factor levels, and upregulation of FAS expression and FA synthesis. Deletion of the major SREBP binding site from the FAS promoter abrogated transcription in transformed MCF-10a cells. Inhibitors of MAP and PI 3-kinases downregulated SREBP-1 levels and decreased transcription from the FAS promoter, reducing FAS expression and fatty acid synthesis in transformed MCF-10a cells and in MCF-7 and HCT116 carcinoma cells. H-ras transformation sensitized MCF-10a cells to the FAS inhibitors cerulenin and C-75. These results confirm an important role for SREBP-1 in neoplastic lipogenesis, and provide a likely basis for the linkage of upregulated fatty acid metabolism with neoplastic transformation and with tumor virulence, since MAP and PI 3-kinase signaling contributes to both.