AMPK regulates energy expenditure by modulating NAD+ metabolism and SIRT1 activity.

AMP-activated protein kinase (AMPK) is a metabolic fuel gauge conserved along the evolutionary scale in eukaryotes that senses changes in the intracellular AMP/ATP ratio. Recent evidence indicated an important role for AMPK in the therapeutic benefits of metformin, thiazolidinediones and exercise, which form the cornerstones of the clinical management of type 2 diabetes and associated metabolic disorders. In general, activation of AMPK acts to maintain cellular energy stores, switching on catabolic pathways that produce ATP, mostly by enhancing oxidative metabolism and mitochondrial biogenesis, while switching off anabolic pathways that consume ATP. This regulation can take place acutely, through the regulation of fast post-translational events, but also by transcriptionally reprogramming the cell to meet energetic needs. Here we demonstrate that AMPK controls the expression of genes involved in energy metabolism in mouse skeletal muscle by acting in coordination with another metabolic sensor, the NAD+-dependent type III deacetylase SIRT1. AMPK enhances SIRT1 activity by increasing cellular NAD+ levels, resulting in the deacetylation and modulation of the activity of downstream SIRT1 targets that include the peroxisome proliferator-activated receptor-gamma coactivator 1alpha and the forkhead box O1 (FOXO1) and O3 (FOXO3a) transcription factors. The AMPK-induced SIRT1-mediated deacetylation of these targets explains many of the convergent biological effects of AMPK and SIRT1 on energy metabolism.

Small molecule activators of SIRT1 as therapeutics for the treatment of type 2 diabetes.

Calorie restriction extends lifespan and produces a metabolic profile desirable for treating diseases of ageing such as type 2 diabetes. SIRT1, an NAD+-dependent deacetylase, is a principal modulator of pathways downstream of calorie restriction that produce beneficial effects on glucose homeostasis and insulin sensitivity. Resveratrol, a polyphenolic SIRT1 activator, mimics the anti-ageing effects of calorie restriction in lower organisms and in mice fed a high-fat diet ameliorates insulin resistance, increases mitochondrial content, and prolongs survival. Here we describe the identification and characterization of small molecule activators of SIRT1 that are structurally unrelated to, and 1,000-fold more potent than, resveratrol. These compounds bind to the SIRT1 enzyme-peptide substrate complex at an allosteric site amino-terminal to the catalytic domain and lower the Michaelis constant for acetylated substrates. In diet-induced obese and genetically obese mice, these compounds improve insulin sensitivity, lower plasma glucose, and increase mitochondrial capacity. In Zucker fa/fa rats, hyperinsulinaemic-euglycaemic clamp studies demonstrate that SIRT1 activators improve whole-body glucose homeostasis and insulin sensitivity in adipose tissue, skeletal muscle and liver. Thus, SIRT1 activation is a promising new therapeutic approach for treating diseases of ageing such as type 2 diabetes.

Sirtuins: novel targets for metabolic disease.

Sirtuins represent a novel family of enzymes that are collectively well situated to help regulate nutrient sensing and utilization, metabolic rate and ultimately metabolic disease. Activation of one of these enzymes, SIRT1, leads to enhanced activity of multiple proteins, including peroxisome-proliferator activated receptor coactivator-1alpha (PGC-1alpha), which helps to mediate some of the in vitro and in vivo effects of sirtuins. As such, enhanced SIRT1 activity decreases glucose levels, improves insulin sensitivity, increases mitochondrial number and function, decreases adiposity, improves exercise tolerance and potentially lowers body weight. SRT-501 is a proprietary formulation of resveratrol with improved bioavailability. As such, SRT-501 represents the first in a novel class of SIRT1 activators that has proven to be safe and well-tolerated in humans. Clinical trials in type 2 diabetic patients are currently underway.

Effects of prednisolone on the systemic release of mediators of cell-mediated cytotoxicity during human endotoxemia.

Corticosteroids are widely used for the suppression of cell-mediated cytoxicity. This process is mediated by natural killer cells and cytotoxic T lymphocytes, and their activation can be monitored by levels of the chemokines CXCL9 and CXCL10, the degranulation product granzymes A and B, and by levels of secretory phospholipase A2. The current study aimed to determine the effects of increasing doses of prednisolone on the release of these mediators in healthy humans exposed to LPS. Therefore, 32 healthy men received prednisolone orally at doses of 0, 3, 10, or 30 mg (n = 8 per group) at 2 h before intravenous injection of Escherichia coil LPS (4 ng/kg). Prednisolone dose-dependently attenuated the LPS-induced rises in the plasma concentrations of the chemokines CXCL9 and CXCL10, as well as of granzymes A and B levels. CXCL10 and granzyme B release were most sensitive to prednisolone, with a significant inhibition already achieved at the lowest prednisolone dose (3 mg). The levels of secretory phospholipase A2 were increased after LPS administration but were not significantly affected by prednisolone. This study demonstrates that prednisolone differentially inhibits the systemic release of mediators involved in cell-mediated cytotoxicity in humans in vivo.

The influence of corticosteroids on the release of novel biomarkers in human endotoxemia.

OBJECTIVE: Sepsis intervention studies need better patient stratification methods, and one way to realize this is the introduction of stable biomarkers. A set of recently developed novel biomarkers, based upon precursor-fragments of short-lived hormones, was previously shown to be increased during sepsis. However, it is not known whether these biomarkers are influenced by sepsis intervention strategies. Therefore we investigated the markers in a model of human endotoxemia intervened by increasing doses of prednisolone. DESIGN AND SETTING: Prospective, open-label study in a specialized clinical research unit of a university hospital. SUBJECTS: Thirty-two healthy male volunteers. INTERVENTIONS: Subjects received prednisolone orally at doses of 0, 3, 10 or 30 mg (n=8 per group) at 2 h before intravenous injection of Escherichia coli lipopolysaccharide (LPS) (4 ng/kg). Blood samples were drawn during 24 h after LPS injection. MEASUREMENTS AND RESULTS: LPS injection caused an increase in levels of midregional pro-adrenomedullin (MR-proADM), midregional pro-atrial natriuretic peptide (MR-proANP), C-terminal pro-arginine-vasopressin (CT-proAVP) and procalcitonin (PCT). Prednisolone caused a dose dependent inhibition of MR-proADM, MR-proANP and CT-proAVP levels. CONCLUSIONS: These results show that a set of novel, highly stable sepsis biomarkers was increased during human endotoxemia and was dose-dependently inhibited by corticosteroid pre-treatment.

Nicotinamide Improves Aspects of Healthspan, but Not Lifespan, in Mice.

The role in longevity and healthspan of nicotinamide (NAM), the physiological precursor of NAD+, is elusive. Here, we report that chronic NAM supplementation improves healthspan measures in mice without extending lifespan. Untargeted metabolite profiling of the liver and metabolic flux analysis of liver-derived cells revealed NAM-mediated improvement in glucose homeostasis in mice on a high-fat diet (HFD) that was associated with reduced hepatic steatosis and inflammation concomitant with increased glycogen deposition and flux through the pentose phosphate and glycolytic pathways. Targeted NAD metabolome analysis in liver revealed depressed expression of NAM salvage in NAM-treated mice, an effect counteracted by higher expression of de novo NAD biosynthetic enzymes. Although neither hepatic NAD+ nor NADP+ was boosted by NAM, acetylation of some SIRT1 targets was enhanced by NAM supplementation in a diet- and NAM dose-dependent manner. Collectively, our results show health improvement in NAM-supplemented HFD-fed mice in the absence of survival effects.

Proteasome inhibition reduces superantigen-mediated T cell activation and the severity of psoriasis in a SCID-hu model.

There is increasing evidence that bacterial superantigens contribute to inflammation and T cell responses in psoriasis. Psoriatic inflammation entails a complex series of inductive and effector processes that require the regulated expression of various proinflammatory genes, many of which require NF-kappa B for maximal trans-activation. PS-519 is a potent and selective proteasome inhibitor based upon the naturally occurring compound lactacystin, which inhibits NF-kappa B activation by blocking the degradation of its inhibitory protein I kappa B. We report that proteasome inhibition by PS-519 reduces superantigen-mediated T cell-activation in vitro and in vivo. Proliferation was inhibited along with the expression of very early (CD69), early (CD25), and late T cell (HLA-DR) activation molecules. Moreover, expression of E-selectin ligands relevant to dermal T cell homing was reduced, as was E-selectin binding in vitro. Finally, PS-519 proved to be therapeutically effective in a SCID-hu xenogeneic psoriasis transplantation model. We conclude that inhibition of the proteasome, e.g., by PS-519, is a promising means to treat T cell-mediated disorders such as psoriasis.

Phase I trial of the proteasome inhibitor PS-341 in patients with refractory hematologic malignancies.

PURPOSE: To determine the maximum-tolerated dose (MTD), dose-limiting toxicity (DLT), and pharmacodynamics (PD) of the proteasome inhibitor bortezomib (previously known as PS-341) in patients with refractory hematologic malignancies. PATIENTS AND METHODS: Patients received PS-341 twice weekly for 4 weeks at either 0.40, 1.04, 1.20, or 1.38 mg/m(2), followed by a 2-week rest. The PD of PS-341 was evaluated by measurement of whole blood 20S proteasome activity. RESULTS: Twenty-seven patients received 293 doses of PS-341, including 24 complete cycles. DLTs at doses above the 1.04-mg/m(2) MTD attributed to PS-341 included thrombocytopenia, hyponatremia, hypokalemia, fatigue, and malaise. In three of 10 patients receiving additional therapy, serious reversible adverse events appeared during cycle 2, including one episode of postural hypotension, one systemic hypersensitivity reaction, and grade 4 transaminitis in a patient with hepatitis C and a substantial acetaminophen ingestion. PD studies revealed PS-341 induced 20S proteasome inhibition in a time-dependent manner, and this inhibition was also related to both the dose in milligrams per meter squared, and the absolute dose of PS-341. Among nine fully assessable patients with heavily pretreated plasma cell dyscrasias completing one cycle of therapy, there was one complete response and a reduction in paraprotein levels and/or marrow plasmacytosis in eight others. In addition, one patient with mantle cell lymphoma and another with follicular lymphoma had shrinkage of nodal disease. CONCLUSION: PS-341 was well tolerated at 1.04 mg/m(2) on this dose-intensive schedule, although patients need to be monitored for electrolyte abnormalities and late toxicities. Additional studies are indicated to determine whether incorporation of dose/body surface area yields a superior PD model to dosing without normalization. PS-341 showed activity against refractory multiple myeloma and possibly non-Hodgkin's lymphoma in this study, and merits further investigation in these populations.

The proteasome inhibitor bortezomib (Velcade) sensitizes some human tumor cells to Apo2L/TRAIL-mediated apoptosis.

On testing a panel of different human cancer cell lines, we observed that the proteasome inhibitor bortezomib could dramatically sensitize some lines to the apoptotic effects of Apo2L/TRAIL. Certain renal, colon, or breast tumor cell lines were dramatically sensitized, whereas other tumor lines from the same tissue of origin remained resistant. This sensitization did not correlate with either the p53 status of the individual tumor cell lines or their intrinsic sensitivity to Apo2L/TRAIL. Colon cancer cell lines lacking p53 or Bax were sensitized by bortezomib, suggesting that neither p53 nor Bax levels were crucial for sensitization. Although the molecular basis of bortezomib sensitization of tumor cells to Apo2L/TRAIL remains to be determined, this combination can have an enhanced apoptotic effect over either agent alone for certain human cancer cells.

Mechanisms of proteasome inhibitor PS-341-induced G(2)-M-phase arrest and apoptosis in human non-small cell lung cancer cell lines.

PURPOSE: PS-341 is a novel dipeptide boronic acid proteasome inhibitor with in vitro and in vivo antitumor activity that induces mechanisms of apoptosis by unknown mechanisms. EXPERIMENTAL DESIGN: Human non-small cell lung cancer cell lines were used to investigate effects PS-341 on cell proliferation, cell cycle progression, and the induction of apoptosis. RESULTS: PS-341 was 38-360-fold more cytotoxic against H460 cells when compared with the proteasome inhibitors MG-132 and PSI. Differential PS-341 cytotoxic effects were found with respect to P53 function: H322 cells (p53 mutant) were 6-fold less sensitive as compared with H460 cells (p53 wild type); and H358 cells (p53 null) were 1.6-fold more sensitive as compared with H460 cells (p53 wild type). A concentration- and time-dependent cell cycle blockade at G(2)-M phase was seen for H460 cells without any direct effects on microtubule polymerization or depolymerization. PS-341 exposure in H460 cells led to stabilization of p53, induction of p21(cip/waf-1) and MDM2 expression, an increase in cyclin B and cyclin A, and the activation of cyclin B and cyclin A kinases. MDM2 induction was found only in H460 cells, whereas in H322 and H358 cells, G(2)-M-phase arrest, p21(cip/waf-1) induction, and an increase in cyclin B1 were found. The commitment of G(2)-M-phase cells to apoptosis was verified by the activation of caspase-3 and cleavage of poly(ADP-ribose) polymerase in drug-free medium. CONCLUSIONS: Our data suggest that the PS-341-induced G(2)-M-phase arrest may be associated with the inhibition of degradation of cell cycle regulators and that the up-regulation of p21(cip/waf-1) expression may be via p53-dependent and/or -independent pathways. The resulting disturbance of cell cycle progression leads either to growth inhibition or to the initiation of apoptotic pathways.

A phase I trial of the novel proteasome inhibitor PS341 in advanced solid tumor malignancies.

PURPOSE: The purpose of this study was to evaluate the toxicity and pharmacodynamic behavior of the novel proteasome inhibitor PS341 administered as a twice weekly i.v. bolus for 2 weeks, followed by a 1-week recovery period in patients with advanced solid tumor malignancies. EXPERIMENTAL DESIGN: In this Phase I trial, 43 patients were treated with PS341 in doses ranging from 0.13 to 1.56 mg/m2/dose. A standard Phase I design was used. Pharmacodynamic studies were performed to access 20S proteasome activity. RESULTS: Forty-three patients were treated with 89 cycles of PS341. Patients were heavily pretreated. Dose-limiting toxicities on this schedule were diarrhea and sensory neurotoxicity. Other side effects seen were fatigue, fever, anorexia, nausea, vomiting, rash, pruritus, and headache. There was no dose-limiting hematological toxicity. A dose-related inhibition of 20S proteasome activity with increasing dose of PS341 was seen. There was one major response in a patient with refractory non-small cell lung carcinoma. CONCLUSIONS: Given the results of this trial, it is safe and reasonable to recommend treatment with PS341 on the schedule used in this trial at 1.56 mg/m2/dose in Phase II trials. Particular care should be taken with patients with preexisting neuropathy. Further testing in Phase II trials is warranted.

Proteasome inhibition: a new anti-inflammatory strategy.

The ubiquitin-proteasome pathway has a central role in the selective degradation of intracellular proteins. Among the key proteins modulated by the proteasome are those involved in the control of inflammatory processes, cell cycle regulation, and gene expression. Consequently proteasome inhibition is a potential treatment option for cancer and inflammatory conditions. Thus far, proof of principle has been obtained from studies in numerous animal models for a variety of human diseases including cancer, reperfusion injury, and inflammatory conditions such as rheumatoid arthritis, asthma, multiple sclerosis, and psoriasis. Two proteasome inhibitors, each representing a unique chemical class, are currently under clinical evaluation. Velcade (PS-341) is currently being evaluated in multiple phase II clinical trials for several solid tumor indications and has just entered a phase III trial for multiple myeloma. PS-519, representing another class of inhibitors, focuses on the inflammatory events following ischemia and reperfusion injury. Since proteasome inhibitors exhibit anti-inflammatory and antiproliferative effects, diseases characterized by both of these processes simultaneously, as is the case in rheumatoid arthritis or psoriasis, might also represent clinical opportunities for such drugs.

PS-341, a novel proteasome inhibitor, induces Bcl-2 phosphorylation and cleavage in association with G2-M phase arrest and apoptosis.

Treatment with the proteasome inhibitor, PS-341 resulted in concentration- and time-dependent effects on Bcl-2 phosphorylation and cleavage in H460 cells that coincided with the PS-341-induced G2-M phase arrest. The observed Bcl-2 cleavage paralleled the degree of PS-341-induced apoptosis but was detected to a similar extent with comparable concentrations of two other proteasome inhibitors (MG-132 and PSI). Calpain inhibitors, ALLM and ALLN, and the caspase inhibitors, Z-VAD and AC-YVAD did not induce BcI-2 phosphorylation and cleavage. Exposure to PS-341 resulted in an additional Mr 25,000 cleavage fragment of Bcl-2, whereas only a Mr 23,000 fragment was observed with other anticancer agents. The formation of the Mr 25,000 fragment was not prevented by caspase inhibitors unlike the Mr 23,000 fragment, which suggests mediation by a caspase-independent pathway. Cell fractionation studies revealed that the Bcl-2 cleaved fragments localize within membrane structures and was an early event (at approximately 12 h, posttreatment), and before the observed cleavage of poly(ADP-ribose) polymerase (PARP), beta-catenin, and DNA fragmentation (at approximately 36 h posttreatment). The Mr 23,000 Bcl-2 cleavage product was inhibited by the pan-caspase inhibitor and the inhibitors of capase-3, -8, -9; but the PARP cleavage was prevented only by the pan-caspase and caspase-3 inhibitors, which suggests that the Mr 23,000 Bcl-2 cleavage occurred at both the initiation and execution stages of apoptosis. The inhibition of the ubiquitin/proteasome pathway by PS-341 leads, at an early stage of apoptosis, to Bcl-2 phosphorylation and a unique proteolytic cleavage product, which are associated with G2-M phase arrest and the induction of apoptosis.

Quantitative real-time RT-PCR analysis of inflammatory gene expression associated with ischemia-reperfusion brain injury.

Ischemia-reperfusion brain injury initiates an inflammatory response involving the expression of adhesion molecules and cytokines, some of which are regulated by the nuclear transcription factor NF-kappaB. In this study the authors examined mRNA expression levels for several important genes associated with inflammation at five time points (3, 6, 12, 24, and 72 hours) after transient middle cerebral artery occlusion (MCAO) in Sprague-Dawley rats. A sensitive and quantitative technique (TaqMan real-time QRT-PCR) was used to simultaneously measure mRNA levels for key cell adhesion molecules and inflammatory cytokines. Gene expression increased significantly in the injured hemisphere for interleukin (IL)-1beta (12-fold increase at 24 hours), IL-6 (25-fold increase at 6 hours) and ICAM-1 (4-fold increase at 24 hours), and the interhemispheric differences for these genes were significant for every time point examined (P < 0.05 for all values). Tumor necrosis factor-alpha mRNA was upregulated in the injured versus uninjured hemisphere from 3 to 24 hours (5-fold increase at 6 hours), while E-selectin showed a significant increase in mRNA levels from 6 to 24 hours after MCAO (10-fold increase at 6 hours) (P < 0.05 for all values). VCAM-1 mRNA levels did not respond differentially to injury at any time point between the two brain hemispheres. At all time points examined, activated NF-kappaB immunoreactivity was observed in cells throughout the infarct-damaged tissue. These results are consistent with the proinflammatory properties of the induced molecules, which are involved in the initiation of the inflammatory cascade, and may thus contribute to secondary cellular responses that lead to further brain damage.

Delayed treatment with MLN519 reduces infarction and associated neurologic deficit caused by focal ischemic brain injury in rats via antiinflammatory mechanisms involving nuclear factor-kappaB activation, gliosis, and leukocyte infiltration.

Secondary brain injury due to ischemia includes the infiltration of leukocytes into the brain parenchyma mediated by activation of nuclear factor-kappaB (NF-kappaB), which is activated by proteasome degradation. Neuroprotection with the proteasome inhibitor MLN519 has previously been reported to decrease ischemic brain injury in rats. The authors used higher doses of MLN519 to evaluate the neuroprotection therapeutic window after 24 hours of brain injury in rats as correlated to proteasome levels, activated NF-kappaB immunoreactivity, and leukocyte infiltration. Male Sprague-Dawley rats were subjected to 2-hour middle cerebral artery occlusion (MCAO) and recovery. MLN519 or vehicle was administered after injury with a single injection given in delayed increments of 2 hours (i.e., 4, 6, or 8 hours after MCAO). Treatment with MLN519 up to 6 hours after MCAO (4 hours after reperfusion) effectively reduced neuronal and astrocytic degeneration, decreased cortical infarct volume, and increased neurologic recovery. These effects were related to >80% reductions in blood proteasome levels, reduced neutrophil infiltration, and a decrease in activated NF-kappaB immunoreactivity. This improved neuroprotection profile and antiinflammatory effect of MLN519 provides an exciting avenue for potential treatment of focal ischemic brain injury in humans.

Sirt1 inhibition promotes in vivo arterial thrombosis and tissue factor expression in stimulated cells.

AIMS: The mammalian silent information regulator-two 1 (Sirt1) blunts the noxious effects of cardiovascular risk factors such as type 2 diabetes mellitus and obesity. Nevertheless, the role of Sirt1 in regulating the expression of tissue factor (TF), the key trigger of coagulation, and arterial thrombus formation remains unknown. METHODS AND RESULTS: Human as well as mouse cell lines were used for in vitro experiments, and C57Bl/6 mice for in vivo procedures. Sirt1 inhibition by splitomicin or sirtinol enhanced cytokine-induced endothelial TF protein expression as well as surface activity, while TF pathway inhibitor protein expression did not change. Sirt1 inhibition further enhanced TF mRNA expression, TF promoter activity, and nuclear translocation as well as DNA binding of the p65 subunit of nuclear factor-kappa B (NFκB/p65). Sirt1 siRNA enhanced TF protein and mRNA expression, and this effect was reduced in NFκB/p65(-/-) mouse embryonic fibroblasts reconstituted with non-acetylatable Lys(310)-mutant NFκB/p65. Activation of the mitogen-activated protein kinases p38, c-Jun NH(2)-terminal kinase, and p44/42 (ERK) remained unaffected. In vivo, mice treated with the Sirt1 inhibitor splitomicin exhibited enhanced TF activity in the arterial vessel wall and accelerated carotid artery thrombus formation in a photochemical injury model. CONCLUSION: We provide pharmacological and genetic evidence that Sirt1 inhibition enhances TF expression and activity by increasing NFκB/p65 activation in human endothelial cells. Furthermore, Sirt1 inhibition induces arterial thrombus formation in vivo. Hence, modulation of Sirt1 may offer novel therapeutic options for targeting thrombosis.

SRT1720 improves survival and healthspan of obese mice.

Sirt1 is an NAD(+)-dependent deacetylase that extends lifespan in lower organisms and improves metabolism and delays the onset of age-related diseases in mammals. Here we show that SRT1720, a synthetic compound that was identified for its ability to activate Sirt1 in vitro, extends both mean and maximum lifespan of adult mice fed a high-fat diet. This lifespan extension is accompanied by health benefits including reduced liver steatosis, increased insulin sensitivity, enhanced locomotor activity and normalization of gene expression profiles and markers of inflammation and apoptosis, all in the absence of any observable toxicity. Using a conditional SIRT1 knockout mouse and specific gene knockdowns we show SRT1720 affects mitochondrial respiration in a Sirt1- and PGC-1α-dependent manner. These findings indicate that SRT1720 has long-term benefits and demonstrate for the first time the feasibility of designing novel molecules that are safe and effective in promoting longevity and preventing multiple age-related diseases in mammals.

Small molecule activators of SIRT1 replicate signaling pathways triggered by calorie restriction in vivo.

BACKGROUND: Calorie restriction (CR) produces a number of health benefits and ameliorates diseases of aging such as type 2 diabetes. The components of the pathways downstream of CR may provide intervention points for developing therapeutics for treating diseases of aging. The NAD+-dependent protein deacetylase SIRT1 has been implicated as one of the key downstream regulators of CR in yeast, rodents, and humans. Small molecule activators of SIRT1 have been identified that exhibit efficacy in animal models of diseases typically associated with aging including type 2 diabetes. To identify molecular processes induced in the liver of mice treated with two structurally distinct SIRT1 activators, SIRT501 (formulated resveratrol) and SRT1720, for three days, we utilized a systems biology approach and applied Causal Network Modeling (CNM) on gene expression data to elucidate downstream effects of SIRT1 activation. RESULTS: Here we demonstrate that SIRT1 activators recapitulate many of the molecular events downstream of CR in vivo, such as enhancing mitochondrial biogenesis, improving metabolic signaling pathways, and blunting pro-inflammatory pathways in mice fed a high fat, high calorie diet. CONCLUSION: CNM of gene expression data from mice treated with SRT501 or SRT1720 in combination with supporting in vitro and in vivo data demonstrates that SRT501 and SRT1720 produce a signaling profile that mirrors CR, improves glucose and insulin homeostasis, and acts via SIRT1 activation in vivo. Taken together these results are encouraging regarding the use of small molecule activators of SIRT1 for therapeutic intervention into type 2 diabetes, a strategy which is currently being investigated in multiple clinical trials.

Sirtuins--novel therapeutic targets to treat age-associated diseases.

Sirtuins post-translationally modulate the function of many cellular proteins that undergo reversible acetylation-deacetylation cycles, affecting physiological responses that have implications for treating diseases of ageing. Potent small-molecule modulators of sirtuins have shown efficacy in preclinical models of metabolic, neurodegenerative and inflammatory diseases, and so hold promise for drug discovery efforts in multiple therapeutic areas. Here, we discuss current knowledge and data that strengthens sirtuins as a druggable set of enzymes for the treatment of age-associated diseases, including activation of SIRT1 in type 2 diabetes.