Ovarian cancer is the most lethal of all gynecological malignancies, due in part to the diagnosis at an advanced stage caused by the lack of specific signs and symptoms and the absence of reliable tests for screening and early detection. Most patients will respond initially to treatment but about 70% of them will suffer a recurrence. Therefore, new therapeutic modalities are urgently needed to overcome chemoresistance observed in ovarian cancer patients. Evidence accumulates suggesting that the insulin/insulin growth factor (IGF) pathways could act as a good therapeutic target in several cancers, including ovarian cancer. In this paper, we will focus on the role of insulin/IGF in ovarian cancer tumorigenesis and treatment.

The worldwide epidemic of obesity and the metabolic syndrome has made nonalcoholic fatty liver disease (NAFLD), one of the most important liver diseases of our time. NAFLD is now the commonest cause of abnormal liver test results in industrialized countries and its incidence is rising. The current treatment of nonalcoholic steatohepatitis (NASH) has focused on lifestyle modification to achieve weight loss and modification of risk factors, such as insulin resistance, dyslipidemia, and hyperglycemia associated with the metabolic syndrome. With our increasing understanding of the pathogenesis of NASH, have come a plethora of new pharmacologic options with great potential to modify the natural history of NAFLD and NASH. This article focuses on a number of novel molecular targets for the treatment of NASH as well as the evidence for currently available therapy. It should be noted, however, that in part because of the long natural history of NASH and NAFLD, no therapy to date has been shown to unequivocally alter liver-related morbidity and mortality in these patients.

ABSTRACT: BACKGROUND: Diabetic cardiomyopathy (DCM) contributes to cardiac failure in diabetic patients. It is characterized by excessive lipids accumulation, with increased triacylglycerol (TAG) stores, and fibrosis in left ventricle (LV). The mechanisms responsible are incompletely known and no specific treatment is presently defined. We evaluated the possible usefulness of two molecules promoting lipid oxidation, fenofibrate and metformin, in an experimental model of DCM, the Zucker diabetic rat (ZDF). METHODS: ZDF and controls (C) rats were studied at 7, 14 and 21 weeks. After an initial study at 7 weeks, ZDF rats received no treatment, metformin or fenofibrate until final studies (at 14 or 21 weeks). C rats received no treatment. Each study comprised measurements of metabolic parameters (plasma glucose, TAG, insulin levels) and sampling of heart for histology and measurements of TAG content and relevant mRNA concentration. RESULTS: ZDF rats were insulin-resistant at 7 weeks, type 2 diabetic at 14 weeks and diabetic with insulin deficiency at 21 weeks. Their plasma TAG levels were increased. ZDF rats had at 7 weeks an increased LV TAG content with some fibrosis. LV TAG content increased in untreated ZDF rats at 14 and 21 weeks and was always higher than in C. Fibrosis increased also moderately in untreated ZDF rats. Metformin and fenofibrate decreased plasma TAG concentrations. LV TAG content was decreased by metformin (14 and 21 weeks) and by fenofibrate (14 weeks). Fibrosis was reduced by fenofibrate only and was increased by metformin. Among the mRNA measured, fenofibrate increased Acyl-CoA Oxidase mRNA level, metformin decreased Acyl-CoA Synthase and increased AdipoR1 and pro-inflammatory mRNA levels. CONCLUSION: Fenofibrate had favourable actions on DCM. Metformin had beneficial effect on TAG content but not on fibrosis. PPARa agonists could be useful for the prevention and treatment of DCM.

AMP-activated protein kinase (AMPK), a phylogenetically conserved serine/threonine protein kinase, has been proposed to function as a fuel gauge to monitor cellular energy status in response to nutritional environmental variations. AMPK system is a regulator of energy balance that, once activated by low energy status, switches on ATP-producing catabolic pathways (such as fatty acid oxidation and glycolysis), and switches off ATP-consuming anabolic pathways (such as lipogenesis), both by short-term effect on phosphorylation of regulatory proteins and by long-term effect on gene expression. Numerous observations obtained with pharmacological activators and agents that deplete intracellular ATP have been supportive of AMPK playing a role in the control of energy metabolism but none of these studies have provided conclusive evidence. Relatively recent developments in our understanding of precisely how AMPK complexes might operate to control energy metabolism is due in part to the development of transgenic and knockout mouse models. Although there are inevitable caveats with genetic models, some important findings have emerged. In the present review, we discuss recent findings obtained from animal models with inhibition or activation of AMPK signaling pathway.

Nutritional excess and/or obesity represent well-known predisposition factors for the development of non-insulin-dependent diabetes mellitus (NIDDM). However, molecular links between obesity and NIDDM are only beginning to emerge. Here, we demonstrate that nutrients suppress phosphatidylinositol 3 (PI3)-kinase/Akt signaling via Raptor-dependent mTOR (mammalian target of rapamycin)-mediated phosphorylation of insulin receptor substrate 1 (IRS-1). Raptor directly binds to and serves as a scaffold for mTOR-mediated phosphorylation of IRS-1 on Ser636/639. These serines lie close to the Y(632)MPM motif that is implicated in the binding of p85alpha/p110alpha PI3-kinase to IRS-1 upon insulin stimulation. Phosphomimicking mutations of these serines block insulin-stimulated activation of IRS-1-associated PI3-kinase. Knockdown of Raptor as well as activators of the LKB1/AMPK pathway, such as the widely used antidiabetic compound metformin, suppress IRS-1 Ser636/639 phosphorylation and reverse mTOR-mediated inhibition on PI3-kinase/Akt signaling. Thus, diabetes-related hyperglycemia hyperactivates the mTOR pathway and may lead to insulin resistance due to suppression of IRS-1-dependent PI3-kinase/Akt signaling.

In 2000, more than 151 million people in the world are diabetic. It is predicted that by 2010, 221 million people and by 2025, 324 million will be diabetic. In the U.S., for the population born in 2000, the estimated lifetime risk for diabetes is more than 1 in 3. The economic and human cost of this disease is devastating. The current cost of diabetes in the U.S. is estimated to be at $132 billion, which includes $92 billion of direct medical costs and $40 billion of indirect costs such as disability, work loss and premature mortality. The outbreak of the current diabetic epidemic has been accompanied by a similarly drastic increase in obesity. The relation between the two is a matter of debate but presumably both are caused by changes in dietary habits and an increasingly sedentary modern lifestyle. Compelling scientific evidence indicates that lifestyle modification effectively prevents or delays the occurrence of type 2 diabetes. Recent clinical trials also demonstrate that success in the treatment of obesity, either surgically or pharmacologically, leads to the prevention of type 2 diabetes among the obese. Clinical data have also revealed that the insulin sensitizing agent troglitazone is efficacious in both beta-cell preservation and delaying the onset of type 2 diabetes. Future safe and more effective anti-obesity medicines and insulin sensitizing agents that help to preserve beta-cell function, in addition to efforts of lifestyle modification, thus hold promise for the overweight population with potential for reduction in the development of diabetics.

Oxygen deprivation leads to the accumulation of misfolded proteins in the endoplasmic reticulum (ER), causing ER stress. Under conditions of ER stress, inhibition of protein synthesis and up-regulation of ER chaperone expression reduce the misfolded proteins in the ER. AMP-activated protein kinase (AMPK) is a key regulatory enzyme involved in energy homeostasis during hypoxia. It has been shown that AMPK activation is associated with inhibition of protein synthesis via phosphorylation of elongation factor 2 (eEF2) in cardiomyocytes. We therefore examined whether AMPK attenuates hypoxia-induced ER stress in neonatal rat cardiomyocytes. We found that hypoxia induced ER stress, as assessed by the expression of CHOP and BiP and cleavage of caspase 12. Knockdown of CHOP or caspase 12 through small interfering RNA (siRNA) resulted in decreased expression of cleaved poly(ADP-ribose) polymerase following exposure to hypoxia. We also found that hypoxia-induced CHOP expression and cleavage of caspase 12 were significantly inhibited by pretreatment with 5-aminoimidazole-4-carboxyamide-1-beta-D-ribofuranoside (AICAR), a pharmacological activator of AMPK. In parallel, adenovirus expressing dominant-negative AMPK significantly attenuated the cardioprotective effects of AICAR. Knockdown of eEF2 phosphorylation using eEF2 kinase siRNA abolished these cardioprotective effects of AICAR. Taken together, these findings demonstrate that activation of AMPK contributes to protection of the heart against hypoxic injury through attenuation of ER stress and that attenuation of protein synthesis via eEF2 inactivation may be the mechanism of cardioprotection by AMPK.

Muscle wasting (cachexia) is a consequence of chronic diseases, such as cancer, and is associated with degradation of muscle proteins such as MyoD. The cytokines tumor necrosis factor alpha and gamma interferon induce muscle degeneration by activating the transcription factor NF-kappaB and its target genes. Here, we show that a downstream target of NF-kappaB is the nitric oxide (NO) synthase gene (iNos) and suggest that NO production stimulates MyoD mRNA loss. In fact, although cytokine treatment of iNos(-/-) mice activated NF-kappaB, it did not trigger MyoD mRNA degeneration, demonstrating that NF-kappaB-mediated muscle wasting requires an active iNOS-NO pathway. The induced expression of iNOS by cytokines relies on both transcriptional activation via NF-kappaB and increased mRNA stability via the RNA-binding protein HuR. Moreover, we show that HuR regulates iNOS expression in an AMP-activated protein kinase (AMPK)-dependent manner. Furthermore, AMPK activation results in HuR nuclear sequestration, inhibition of iNOS synthesis, and reduction in cytokine-induced MyoD loss. These results define iNOS and HuR as critical players in cytokine-induced cachexia, establishing them as potential therapeutic targets.

Cell growth and proliferation requires an intricate coordination between the stimulatory signals arising from nutrients and growth factors and the inhibitory signals arising from intracellular and extracellular stresses. Alteration of the coordination often causes cancer. In mammals, the mTOR (mammalian target of rapamycin) protein kinase is the central node in nutrient and growth factor signaling, and p53 plays a critical role in sensing genotoxic and other stresses. The results presented here demonstrate that activation of p53 inhibits mTOR activity and regulates its downstream targets, including autophagy, a tumor suppression process. Moreover, the mechanisms by which p53 regulates mTOR involves AMP kinase activation and requires the tuberous sclerosis (TSC) 1/TSC2 complex, both of which respond to energy deprivation in cells. In addition, glucose starvation not only signals to shut down mTOR, but also results in the transient phosphorylation of the p53 protein. Thus, p53 and mTOR signaling machineries can cross-talk and coordinately regulate cell growth, proliferation, and death.

Recent studies indicate that the LKB1 tumour suppressor protein kinase is the major "upstream" activator of the energy sensor AMP-activated protein kinase (AMPK). We have used mice in which LKB1 is expressed at only approximately 10% of the normal levels in muscle and most other tissues, or that lack LKB1 entirely in skeletal muscle. Muscle expressing only 10% of the normal level of LKB1 had significantly reduced phosphorylation and activation of AMPKalpha2. In LKB1-lacking muscle, the basal activity of the AMPKalpha2 isoform was greatly reduced and was not increased by the AMP-mimetic agent, 5-aminoimidazole-4-carboxamide riboside (AICAR), by the antidiabetic drug phenformin, or by muscle contraction. Moreover, phosphorylation of acetyl CoA carboxylase-2, a downstream target of AMPK, was profoundly reduced. Glucose uptake stimulated by AICAR or muscle contraction, but not by insulin, was inhibited in the absence of LKB1. Contraction increased the AMP:ATP ratio to a greater extent in LKB1-deficient muscles than in LKB1-expressing muscles. These studies establish the importance of LKB1 in regulating AMPK activity and cellular energy levels in response to contraction and phenformin.

Sensitivity of glucose transport to stimulation by insulin has been shown to occur concomitant with activation of the AMP-activated protein kinase (AMPK) in skeletal muscle, suggesting a role of AMPK in regulation of insulin action. The purpose of the present study was to evaluate a possible role of AMPK in potentiation of insulin action in muscle cells. The experimental model involved insulin-responsive C2C12 myotubes that exhibit a twofold increase in glucose transport in the presence of insulin. Treatment of myotubes with the AMPK activator 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR), followed by a 2-h recovery, augmented the ability of insulin to stimulate glucose transport. Similarly, incubation in hyperosmotic medium, another AMPK-activating treatment, acted synergistically with insulin to stimulate glucose transport. Furthermore, the increase in insulin action caused by hyperosmotic stress was prevented by inclusion of compound C, an AMPK inhibitor, in hyperosmotic medium. In addition, iodotubercidin, a general kinase inhibitor that is effective against AMPK, also prevented the combined effects of insulin and hyperosmotic stress on glucose transport. The new information provided by these data is that previously reported AICAR effects on insulin action are generalizable to myotubes, hyperosmotic stress and insulin synergistically increase glucose transport, and AMPK appears to mediate potentiation of insulin action.

Diabetes mellitus is a chronic disease that is growing in prevalence worldwide. Pharmacologic therapy is often necessary to achieve optimal glycemic control in the management of diabetes. Orally administered antihyperglycemic agents (OHAs) can be used either alone or in combination with other OHAs or insulin. The number of available OHAs has increased significantly in the last decade, which translates into more therapeutic options and complex decision-making for physicians. This review article is designed to help with these decisions. We review the mechanism of action, efficacy and side effects of the different classes of OHAs (alpha-glucosidase inhibitors, biguanides, insulin secretagogues, insulin sensitizers and intestinal lipase inhibitor) and discuss the current recommendations for their use.

AMPK (AMP-activated protein kinase) is a key sensor of energy status within the cell. Activated by an increase in the AMP/ATP ratio, AMPK acts to limit cellular energy depletion by down-regulating selective ATP-dependent processes. The purpose of the present study was to determine the role of AMPK in regulating intestinal glucose transport.[3H]3-O-methyl glucose fluxes were measured in murine jejunum in the presence and absence of the AMPK activators AICAR (5-aminoimidazole-4-carboxamide riboside) and metformin and the p38 inhibitor, SB203580. To differentiate between a sodium-coupled (SGLT1) and diffusive (GLUT2) route of entry, fluxes were measured in the presence of the SGLT1 and GLUT2 inhibitors phloridzin and phloretin. Glucose transporter mRNA levels were measured by reverse transcriptase-PCR, and localization by Western blotting. Surface-expressed GLUT2 was assessed by luminal biotinylation. Activation of p38 mitogen-activated protein kinase was analysed by Western blotting. We found that treatment of jejunal tissue with AICAR resulted in enhanced net glucose uptake and was associated with phosphorylation of p38 mitogen-activated protein kinase. Inhibition of p38 abrogated the stimulation of AICAR-stimulated glucose uptake. Phloretin abolished the AICAR-mediated increase in glucose flux, whereas phloridzin had no effect, suggesting the involvement of GLUT2. In addition, AICAR decreased total protein levels of SGLT1, concurrently increasing levels of GLUT2 in the brush-border membrane. The anti-diabetic drug metformin, a known activator of AMPK, also induced the localization of GLUT2 to the luminal surface. We conclude that the activation of AMPK results in an up-regulation of non-energy requiring glucose uptake by GLUT2 and a concurrent down-regulation of sodium-dependent glucose transport.

Three kinases, Pak1, Tos3, and Elm1, activate Snf1 protein kinase in Saccharomyces cerevisiae. This cascade is conserved in mammals, where LKB1 activates AMP-activated protein kinase. We address the specificity of the activating kinases for the three forms of Snf1 protein kinase containing the beta-subunit isoforms Gal83, Sip1, and Sip2. Pak1 is the most important kinase for activating Snf1-Gal83 in response to glucose limitation, but Elm1 also has a significant role; moreover, both Pak1 and Elm1 affect Snf1-Sip2. These findings exclude the possibility of a one-to-one correspondence between the activating kinases and the Snf1 complexes. We further identify a second, unexpected role for Pak1 in regulating Snf1-Gal83: the catalytic activity of Pak1 is required for the nuclear enrichment of Snf1-Gal83 in response to carbon stress. The nuclear enrichment of Snf1 fused to green fluorescent protein (GFP) depends on both Gal83 and Pak1 and is abolished by a mutation of the activation loop threonine; in contrast, the nuclear enrichment of Gal83-GFP occurs in a snf1Delta mutant and depends on Pak1 only when Snf1 is present. Snf1-Gal83 is the only form of the kinase that localizes to the nucleus. These findings, that Pak1 both activates Snf1-Gal83 and controls its nuclear localization, implicate Pak1 in regulating nuclear Snf1 protein kinase activity.

Obesity and its associated comorbidities are among the most prevalent and challenging conditions confronting the medical profession in the 21st century. A major metabolic consequence of obesity is insulin resistance, which is strongly associated with the deposition of triglycerides in the liver. Hepatic steatosis can either be a benign, noninflammatory condition that appears to have no adverse sequelae or can be associated with steatohepatitis: a condition that can result in end-stage liver disease, accounting for up to 14% of liver transplants in the US. Here we highlight recent advances in our understanding of the molecular events contributing to hepatic steatosis and nonalcoholic steatohepatitis.

Tuberous sclerosis complex (TSC) and Peutz-Jeghers syndrome (PJS) are dominantly inherited benign tumor syndromes that share striking histopathological similarities. Here we show that LKB1, the gene mutated in PJS, acts as a tumor suppressor by activating TSC2, the gene mutated in TSC. Like TSC2, LKB1 inhibits the phosphorylation of the key translational regulators S6K and 4EBP1. Furthermore, we show that LKB1 activates TSC2 through the AMP-dependent protein kinase (AMPK), indicating that LKB1 plays a role in cell growth regulation in response to cellular energy levels. Our results suggest that PJS and other benign tumor syndromes could be caused by dysregulation of the TSC2/mTOR pathway.

Metformin, a drug widely used in the treatment of Type II diabetes, has recently received attention owing to new findings regarding its mitochondrial and cellular effects. In the present study, the effects of metformin on respiration, complex 1 activity, mitochondrial permeability transition, cytochrome c release and cell death were investigated in cultured cells from a human carcinoma-derived cell line (KB cells). Metformin significantly decreased respiration both in intact cells and after permeabilization. This was due to a mild and specific inhibition of the respiratory chain complex 1. In addition, metformin prevented to a significant extent mitochondrial permeability transition both in permeabilized cells, as induced by calcium, and in intact cells, as induced by the glutathione-oxidizing agent t-butyl hydroperoxide. This effect was equivalent to that of cyclosporin A, the reference inhibitor. Finally, metformin impaired the t-butyl hydroperoxide-induced cell death, as judged by Trypan Blue exclusion, propidium iodide staining and cytochrome c release. We propose that metformin prevents the permeability transition-related commitment to cell death in relation to its mild inhibitory effect on complex 1, which is responsible for a decreased probability of mitochondrial permeability transition.

Stearoyl-CoA desaturase (SCD) catalyzes the rate-limiting step in the biosynthesis of monounsaturated fatty acids. Mice with a targeted disruption of the SCD1 isoform have reduced body adiposity, increased energy expenditure, and up-regulated expression of several genes encoding enzymes of fatty acid beta-oxidation in liver. The mechanisms by which SCD deficiency leads to these metabolic changes are presently unknown. Here we show that the phosphorylation and activity of AMP-activated protein kinase (AMPK), a metabolic sensor that regulates lipid metabolism during increased energy expenditure is significantly increased (approximately 40%, P < 0.01) in liver of SCD1 knockout mice (SCD1-/-). In parallel with the activation of AMPK, the phosphorylation of acetyl-CoA carboxylase at Ser-79 was increased and enzymatic activity was decreased (approximately 35%, P < 0.001), resulting in decreased intracellular levels of malonyl-CoA (approximately 47%, P < 0.001). An SCD1 mutation also increased AMPK phosphorylation and activity and increased acetyl-CoA carboxylase phosphorylation in leptin-deficient ob/ob mice. Lower malonyl-CoA concentrations are known to derepress carnitine palmitoyltransferase 1 (CPT1). In SCD1-/- mice, CPT1 and CPT2 activities were significantly increased (in both cases approximately 60%, P < 0.001) thereby stimulating the oxidation of mitochondrial palmitoyl-CoA. Our results identify AMPK as a mediator of increased fatty acid oxidation in liver of SCD1-deficient mice.

AMP-activated protein kinase (AMPK) is a highly conserved sensor of cellular energy status found in all eukaryotic cells. AMPK is activated by stimuli that increase the cellular AMP/ATP ratio. Essential to activation of AMPK is its phosphorylation at Thr-172 by an upstream kinase, AMPKK, whose identity in mammalian cells has remained elusive. Here we present biochemical and genetic evidence indicating that the LKB1 serine/threonine kinase, the gene inactivated in the Peutz-Jeghers familial cancer syndrome, is the dominant regulator of AMPK activation in several mammalian cell types. We show that LKB1 directly phosphorylates Thr-172 of AMPKalpha in vitro and activates its kinase activity. LKB1-deficient murine embryonic fibroblasts show nearly complete loss of Thr-172 phosphorylation and downstream AMPK signaling in response to a variety of stimuli that activate AMPK. Reintroduction of WT, but not kinase-dead, LKB1 into these cells restores AMPK activity. Furthermore, we show that LKB1 plays a biologically significant role in this pathway, because LKB1-deficient cells are hypersensitive to apoptosis induced by energy stress. On the basis of these results, we propose a model to explain the apparent paradox that LKB1 is a tumor suppressor, yet cells lacking LKB1 are resistant to cell transformation by conventional oncogenes and are sensitive to killing in response to agents that elevate AMP. The role of LKB1/AMPK in the survival of a subset of genetically defined tumor cells may provide opportunities for cancer therapeutics.