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Pancreatic beta cell function is an important component of glucose homeostasis. Here, we investigated the function of PIMT (PRIP-interacting protein with methyl transferase domain), a transcriptional co-activator binding protein, in the pancreatic beta cells. We observed that the protein levels of PIMT, along with key beta cell markers such as PDX1 (pancreatic and duodenal homeobox 1) and MafA (MAF bZIP transcription factor A), were reduced in the beta cells exposed to hyperglycemic and hyperlipidemic conditions. Consistently, PIMT levels were reduced in the pancreatic islets isolated from high fat diet (HFD)-fed mice. The RNA sequencing analysis of PIMT knockdown beta cells identified that the expression of key genes involved in insulin secretory pathway, Ins1 (insulin 1), Ins2 (insulin 2), Kcnj11 (potassium inwardly-rectifying channel, subfamily J, member 11), Kcnn1 (potassium calcium-activated channel subfamily N member 1), Rab3a (member RAS oncogene family), Gnas (GNAS complex locus), Syt13 (synaptotagmin 13), Pax6 (paired box 6), Klf11 (Kruppel-Like Factor 11), and Nr4a1 (nuclear receptor subfamily 4, group A, member 1) was attenuated due to PIMT depletion. PIMT ablation in the pancreatic beta cells and in the rat pancreatic islets led to decreased protein levels of PDX1 and MafA, resulting in the reduction in glucose-stimulated insulin secretion (GSIS). The results from the immunoprecipitation and ChIP experiments revealed the interaction of PIMT with PDX1 and MafA, and its recruitment to the insulin promoter, respectively. Importantly, PIMT ablation in beta cells resulted in the nuclear translocation of insulin. Surprisingly, forced expression of PIMT in beta cells abrogated GSIS, while Ins1 and Ins2 transcript levels were subtly enhanced. On the other hand, the expression of genes, PRIP/Asc2/Ncoa6 (nuclear receptor coactivator 6), Pax6, Kcnj11, Syt13, Stxbp1 (syntaxin binding protein 1), and Snap25 (synaptosome associated protein 25) associated with insulin secretion, was significantly reduced, providing an explanation for the decreased GSIS upon PIMT overexpression. Our findings highlight the importance of PIMT in the regulation of insulin synthesis and secretion in beta cells.

In view of the reported chorismate mutase (CM or

Mycobacterium tuberculosis (Mtb) is a pathogen of major concern due to its ability to withstand both first- and second-line antibiotics, leading to drug resistance. Thus, there is a critical need for identification of novel anti-tuberculosis agents targeting Mtb-specific proteins. The ceaseless search for novel antimicrobial agents to combat drug-resistant bacteria can be accelerated by the development of advanced deep learning methods, to explore both existing and uncharted regions of the chemical space. The adaptation of deep learning methods to under-explored pathogens such as Mtb is a challenging aspect, as most of the existing methods rely on the availability of sufficient target-specific ligand data to design novel small molecules with optimized bioactivity. In this work, we report the design of novel anti-tuberculosis agents targeting the Mtb chorismate mutase protein using a structure-based drug design algorithm. The structure-based deep learning method relies on the knowledge of the target protein's binding site structure alone for conditional generation of novel small molecules. The method eliminates the need for curation of a high-quality target-specific small molecule dataset, which remains a challenge even for many druggable targets, including Mtb chorismate mutase. Novel molecules are proposed, that show high complementarity to the target binding site. The graph attention model could identify the probable key binding site residues, which influenced the conditional molecule generator to design new molecules with pharmacophoric features similar to the known inhibitors.

Efforts have been devoted for the discovery and development of positive allosteric modulators (PAMs) of 5-HT

That reversible protein phosphorylation by kinases and phosphatases occurs in metabolic disorders is well known. Various studies have revealed that a multi-faceted and tightly regulated phosphatase, pleckstrin homology domain leucine-rich repeat protein phosphatase (PHLPP)-1/2 displays robust effects in cardioprotection, ischaemia/reperfusion (I/R), and vascular remodelling. PHLPP1 promotes foamy macrophage development through ChREBP/AMPK-dependent pathways. Adipocyte-specific loss of PHLPP2 reduces adiposity, improves glucose tolerance,and attenuates fatty liver via the PHLPP2-HSL-PPARα axis. Discoveries of PHLPP1-mediated insulin resistance and pancreatic β cell death via the PHLPP1/2-Mst1-mTORC1 triangular loop have shed light on its significance in diabetology. PHLPP1 downregulation attenuates diabetic cardiomyopathy (DCM) by restoring PI3K-Akt-mTOR signalling. In this review, we summarise the functional role of, and cellular signalling mediated by, PHLPPs in metabolic tissues and discuss their potential as therapeutic targets.

Transcriptional coactivators play a crucial role in regulating gene expression. PRIP interacting protein with methyl transferase domain (PIMT)/trimethyl guanosine synthase 1 (TGS1) is a co-activator interacting protein with an RNA methyl transferase domain. PIMT serves as a bridge between HAT and non-HAT coactivators and differentially modulates gene expression. Disruption of PIMT is embryonic lethal. PIMT regulates hepatic gluconeogenesis and TNF-α-induced insulin resistance in the skeletal muscle. As a methyl transferase, PIMT controls post-transcriptional regulation of HIV-1 and is essential for human telomerase RNA biogenesis. This review comprehensively describes the dual role of PIMT, which promises to be a putative target in metabolic disorders.

A series of indole based novel Schiff bases was designed as potential agonists of 5-HT

Diabetes is a chronic progressive disease associated with different co-morbidities. The availability of insulin or effective use of insulin in the body is the major cause of this disease. Enhanced blood sugar or hyperglycemia is the manifestation of this disease. If not treated timely, diabetes can damage cardiac system, blood vessels, vision, renal function and nerves leading to increased risk of cardiac disease, stroke and may cause neuronal, retinal, and kidney disease. The overall risk of morbidity and mortality among diabetes affected people is more than double compared to non-diabetic people (WHO n.d.).

Clinical studies have proven that rigorous control of hyperglycemia in type 2 diabetes patients can reduce the occurrence of chronic complications (Fig. 7.1). At present therapies of type 2 diabetes can be classified as following based on their mode of action and class

Nutritional abundance associated with chronic inflammation and dyslipidemia impairs the functioning of endoplasmic reticulum (ER) thereby hampering cellular responses to insulin. PHLPP1 was identified as a phosphatase which inactivates Akt, the master regulator of insulin mediated glucose homeostasis. Given the suggestive role of PHLPP1 phosphatase in terminating insulin signalling pathways, deeper insights into its functional role in inducing insulin resistance are warranted. Here, we show that PHLPP1 expression is enhanced in skeletal muscle of insulin resistant rodents which also displayed ER stress, an important mediator of insulin resistance. Using cultured cells and PHLPP1 knockdown mice, we demonstrate that PHLPP1 facilitates the development of ER stress. Importantly, shRNA mediated ablation of PHLPP1 significantly improved glucose clearance from systemic circulation with enhanced expression of glucose transporter 4 (GLUT-4) in skeletal muscle. Mechanistically, we show that endogenous PHLPP1 but not PP2Cα interacts with and directly dephosphorylates AMPK Thr172 in myoblasts without influencing its upstream kinase, LKB1. While the association between endogenous PHLPP1 and AMPK was enhanced in ER stressed cultured cells and soleus muscle of high fat diet fed mice, the basal interaction between PP2Ac and AMPK was minimally altered. Further, we show that PHLPP1α is phosphorylated by ERK1/2 at Ser932 under ER stress which is required for its ability to interact with and dephosphorylate AMPK and thereby induce ER stress. Taken together, our data position PHLPP1 as a key regulator of ER stress.

A series of indole based novel Schiff bases was designed as potential agonists of 5‑HT2C receptor that was supported by docking studies in silico. These compounds were synthesized via Amberlyst-15 catalysed condensation of an appropriate pyrazole based primary amine with the corresponding indole-3-aldehyde under ultrasound irradiation at ambient temperature. A number of target Schiff bases were obtained in good yields (77-87%) under mild conditions within 1 h. Notably, the methodology afforded the corresponding pyrazolo[4,3-d]pyrimidin-7(4H)-one derivatives when the primary amine was replaced by a secondary amine. Several Schiff bases showed agonist activity when tested against human 5-HT2C using luciferase assay in HEK293T cells in vitro. The SAR (Structure-Activity-Relationship) studies suggested that the imine moiety was more favorable over its cyclic form i.e. the corresponding pyrazolopyrimidinone ring. The Schiff bases 3b (EC50 1.8 nM) and 3i (EC50 5.7 nM) were identified as the most active compounds and were comparable with Lorcaserin (EC50 8.5 nM). Also like Lorcaserin, none of these compounds were found to be PAM of 5-HT2C. With ~24 and ~150 fold selectivity towards 5-HT2C over 5-HT2A and 5-HT2B respectively the compound 3i that reduced locomotor activity in zebrafish (Danio rerio) larvae model emerged as a promising hit molecule for further study.

Chronic inflammatory diseases such as insulin resistance, Type 2 diabetes, neurodegenerative diseases etc., are shown to be caused due to imbalanced activation states of macrophages. MicroRNAs which are transcriptional/post-transcriptional regulators of gene expression drive several pathophysiological processes including macrophage polarization. However the functional role of microRNAs in regulating inflammation induced insulin resistance is ill defined. In our current study we observed that the expression of miR-712 was reduced in macrophages exposed to LPS and IFN-γ. Ectopic expression of miR-712 in RAW 264.7 mouse macrophages impaired the expression of iNOS protein and secretion of pro-inflammatory cytokines such as TNF-α, IL-6 and IFN-β which in turn led to improved insulin stimulated glucose uptake in co-cultured L6 myoblasts. Mechanistically, we identified that miR-712 targets the 3'UTR of a potent inflammatory gene LRRK2 and dampens the phosphorylation of p38 and ERK1/2 kinases. Taken together, our data underscore the regulatory role of miR-712 in restoring insulin stimulated glucose uptake by myoblasts through down-regulating macrophage mediated inflammatory responses.

The chorismate mutase (CM) is considered as an attractive target for the identification of potential antitubercular agents due to its absence in animals but not in bacteria. A series of 3-indolylmethyl substituted pyrazolotriazinone derivatives were designed and docked into CM in silico as potential inhibitors. These compounds were efficiently synthesized using the Pd/Cu-catalyzed coupling-cyclization in a single pot involving the construction of indole ring. The methodology was later extended to the preparation of corresponding benzo analogs of pyrazolotriazinones i.e. 3-indolylmethyl substituted benzotriazinone derivatives. Several of these novel compounds showed significant inhibition of CM when tested in vitro at 30 µM. The SAR (Structure-Activity-Relationship) studies suggested that benzotriazinone moiety was more favorable over the pyrazolotriazinone ring. The two best active compounds showed IC50 ∼ 0.4–0.9 µM (better than the reference/known compounds used) and no toxicity till 30 µM in vitro.

Statins are first-line therapy drugs for cholesterol lowering. While they are highly effective at lowering cholesterol, they have propensity to induce hyperglycemia in patients. Only limited studies have been reported which studied the impact of statins on (a) whether they can worsen glucose tolerance in a high sucrose fed animal model and (b) if so, what could be the molecular mechanism. We designed studies using high sucrose fed animals to explore the above questions. The high sucrose fed animals were treated with atorvastatin and simvastatin, the two most prescribed statins. We examined the effects of statins on hyperglycemia, glucose tolerance, fatty acid accumulation and insulin signaling. We found that chronic treatment with atorvastatin made the animals hyperglycemic and glucose intolerant in comparison with diet alone. Treatment with both statins lead to fatty acid accumulation and inhibition of insulin signaling in the muscle tissue at multiple points in the pathway.

Peroxisome proliferator-activated receptor-α (PPARα) modulates the activities of all three interlinked hepatic fatty acid oxidation systems, namely mitochondrial and peroxisomal β-oxidation and microsomal ω-oxidation pathways. Hyperactivation of PPARα, by both exogenous and endogenous activators up-regulates hepatic fatty acid oxidation resulting in excess energy burning in liver contributing to the development of liver cancer in rodents. Sustained PPARα signaling disproportionately increases H2O2-generating fatty acid metabolizing enzymes as compared to H2O2-degrading enzymes in liver leading to enhanced generation of DNA damaging reactive oxygen species, progressive endoplasmic reticulum stress and inflammation. These alterations also contribute to increased liver cell proliferation with changes in apoptosis. Thus, reactive oxygen species, oxidative stress and hepatocellular proliferation are likely the main contributing factors in the pathogenesis of hepatocarcinogenesis, mediated by sustained PPARα activation-related energy burning in liver. Furthermore, the transcriptional co-activator Med1, a key subunit of the Mediator complex, is essential for PPARα signaling in that both PPARα-null and Med1-null hepatocytes are unresponsive to PPARα activators and fail to give rise to liver tumors when chronically exposed to PPARα activators.

Mediator, a large multisubunit protein complex, plays a pivotal role in gene transcription by linking gene-specific transcription factors with the preinitiation complex and RNA polymerase II. In the liver, the key subunit of the Mediator complex, Med1, interacts with several nuclear receptors and transcription factors to direct gene-specific transcription. Conditional knock-out of Med1 in the liver showed that hepatocytes lacking Med1 did not regenerate following either partial hepatectomy or treatment with certain nuclear receptor activators and failed to give rise to tumors when challenged with carcinogens. We now report that the adenovirally driven overexpression of Med1 in mouse liver stimulates hepatocyte DNA synthesis with enhanced expression of DNA replication, cell cycle control, and liver-specific genes, indicating that Med1 alone is necessary and sufficient for liver cell proliferation. Importantly, we demonstrate that AMP-activated protein kinase (AMPK), an important cellular energy sensor, interacts with, and directly phosphorylates, Med1 in vitro at serine 656, serine 756, and serine 796. AMPK also phosphorylates Med1 in vivo in mouse liver and in cultured primary hepatocytes and HEK293 and HeLa cells. In addition, we demonstrate that PPARα activators increase AMPK-mediated Med1 phosphorylation in vivo. Inhibition of AMPK by compound C decreased hepatocyte proliferation induced by Med1 and also by the PPARα activators fenofibrate and Wy-14,643. Co-treatment with compound C attenuated PPARα activator-inducible fatty acid β-oxidation in liver. Our results suggest that Med1 phosphorylation by its association with AMPK regulates liver cell proliferation and fatty acid oxidation, most likely as a downstream effector of PPARα and AMPK. Background: The Med1 subunit of the Mediator complex orchestrates metabolic pathways of energy metabolism. Results: Overexpression of Med1 in liver enhances hepatocyte proliferation and the cellular energy sensor AMPK phosphorylates Med1. Conclusion: Med1 is essential for hepatocellular proliferation and energy homeostasis downstream to AMPK. Significance: Med1 and its phosphorylation by AMPK may have important implications in the pathophysiology of liver.

Amberlyst-15 has been identified as a green and reusable catalyst in the synthesis of spiro 2,3-dihydroquinazolin-4(1H)-ones under ultrasound irradiation at room temperature. The reaction can be performed in an open flask as the conversion was found to be not sensitive to the presence of air or atmospheric moisture. The methodology was found to be general and a wide variety of spiro 2,3-dihydroquinazolin-4(1H)-ones were prepared from 2-aminobenzamides and cyclic ketones within a few minutes in quantitative yields. The products isolated do not require any chromatographic purification. This method provides advantages such as shorter reaction time, high yields of products, and simple operational procedures. Some of the compounds showed chorismate mutase inhibitory properties when tested in vitro.

Mycobacterium tuberculosis chorismate mutase (MtbCM) catalyzes the rearrangement of chorismate to prephenate in the shikimate biosynthetic pathway to form the essential amino acids, phenylalanine and tyrosine. Two genes encoding chorismate mutase have been identified in Mtb. The secretory form,∗MtbCM (encoded by Rv1885c) is assumed to play a key role in pathogenesis of tuberculosis. Also, the inhibition of MtbCM may hinder the supply of nutrients to the organism. Indeed, the existence of chorismate mutase (CM) in bacteria, fungi and higher plants but not in human and low sequence homology among known CM makes it an interesting target for the discovery of anti-tubercular agents. The present article mainly focuses on the recent developments in the structure, function and inhibition of MtbCM. The understanding of various aspects of MtbCM as presented in the current article may facilitate the design and subsequent chemical synthesis of new inhibitors against ∗MtbCM, that could lead to the discovery and development of novel and potent anti-tubercular agents in future.

A direct and single-step method has been developed for the synthesis of mono and 2,3-disubstituted quinoxalines by using a AlCl3 induced (hetero)arylation of 2,3-dichloroquinoxaline. Both symmetrical and unsymmetrical 2,3-disubstituted quinoxalines can be prepared conveniently by using this method under appropriate reaction conditions. The reaction proceeds via C–C bond formation and can be utilized for the preparation of a variety of quinoxaline derivatives from readily available starting materials and reagents. The molecular structure of a representative compound was confirmed by single crystal X-ray diffraction study. Some of the compounds synthesized were tested for chorismate mutase inhibitory properties in vitro and one compound showed promising activity representing one of the few examples of chorismate mutase inhibition by a heteroarene based small molecule.

Metabolic syndrome is characterized by a cluster of metabolic disorders, such as reduced glucose tolerance, hyperinsulinemia, hypertension, visceral obesity and lipid disorders. The benefit of exercise in maintaining total metabolic control is well known and recent research indicates that AMP-activated protein kinase (AMPK) may play an important role in exercise-related effects. AMPK is considered as a master switch in regulating glucose and lipid metabolism. AMPK is an enzyme that works as a fuel gauge, being activated in conditions of high phosphate depletion. In the liver, activation of AMPK results in decreased production of plasma glucose, cholesterol, triglyceride and enhanced fatty acid oxidation. AMPK is also robustly activated by skeletal muscle contraction and myocardial ischemia, and is involved in the stimulation of glucose transport and fatty acid oxidation by these stimuli. In adipose tissue, activated AMPK inhibits deposition of fat, but enhances breakdown and burning of stored fat, resulting in reduction of body weight. The two leading diabetic drugs, namely metformin and rosiglitazone, and adipokines, such as adiponectin and leptin, show their metabolic effects partially through AMPK. These data suggest that AMPK may be a key player in the development of new treatments for obesity, Type 2 diabetes and the metabolic syndrome. In this review, the author provide insight into the role of AMPK as a probable target for treatment of metabolic syndrome.

The mechanisms underlying inflammation induced insulin resistance are poorly understood. Here, we report that the expression of PIMT, a transcriptional co-activator binding protein, was up-regulated in the soleus muscle of high sucrose diet (HSD) induced insulin resistant rats and TNF-α exposed cultured myoblasts. Moreover, TNF-α induced phosphorylation of PIMT at the ERK1/2 target site Ser298. Wild type (WT) PIMT or phospho-mimic Ser298Asp mutant but not phospho-deficient Ser298Ala PIMT mutant abrogated insulin stimulated glucose uptake by L6 myotubes and neonatal rat skeletal myoblasts. Whereas, PIMT knock down relieved TNF-α inhibited insulin signaling. Mechanistic analysis revealed that PIMT differentially regulated the expression of GLUT4, MEF2A, PGC-1α and HDAC5 in cultured cells and skeletal muscle of Wistar rats. Further characterization showed that PIMT was recruited to GLUT4, MEF2A and HDAC5 promoters and overexpression of PIMT abolished the activity of WT but not MEF2A binding defective mutant GLUT4 promoter. Collectively, we conclude that PIMT mediates TNF-α induced insulin resistance at the skeletal muscle via the transcriptional modulation of GLUT4, MEF2A, PGC-1α and HDAC5 genes.

Uncontrolled inflammation leads to several diseases such as insulin resistance, T2D and several types of cancers. The functional role of microRNAs in inflammation induced insulin resistance is poorly studied. MicroRNAs are post-transcriptional regulatory molecules which mediate diverse biological processes. We here show that miR-16 expression levels are down-regulated in different inflammatory conditions such as LPS/IFNγ or palmitate treated macrophages, palmitate exposed myoblasts and insulin responsive tissues of high sucrose diet induced insulin resistant rats. Importantly, forced expression of miR-16 in macrophages impaired the production of TNF-α, IL-6 and IFN-β leading to enhanced insulin stimulated glucose uptake in co-cultured skeletal myoblasts. Further, ectopic expression of miR-16 enhanced insulin stimulated glucose uptake in skeletal myoblasts via the up-regulation of GLUT4 and MEF2A, two key players involved in insulin stimulated glucose uptake. Collectively, our data highlight the important role of miR-16 in ameliorating inflammation induced insulin resistance.

Diverse analogs of 1,5-diarylpyrazoles having 3-hydroxymethyl-4-sulfamoyl (SO2NH2)/methyl sulfonyl (SO2Me)-pheny group at N1 were synthesized and evaluated for their in vitro cyclooxygenase (COX-1/COX-2) inhibitory activity. The SAR study mainly involved the variations at positions C-3, C-5 and N1 of the pyrazole ring. Several small hydrophobic groups at/around position-4 of C-5 phenyl, viz. 3,4-dimethylphenyl analog 9, 3-methyl-4-methylsulfanylphenyl analog 14 and 2,3-dihydrobenzo[b]thiophenyl analog 17, exhibited impressive COX-2 inhibitory potency. In general, the sulfonamide analogues with a CHF2 at C-3 were found to be more potent than those having a CF3 group. The three dimensional quantitative structure activity relationship comprising comparative molecular field analysis (3D-QSAR-CoMFA) afforded the models with high predictivity which further validated the acceptance of hydroxymethyl (CH2OH) group in the hydrophilic pocket of the COX-2 enzyme.

A number of novel 1,5-diarylpyrazoles possessing N-substitution on the sulfonamide (-SO2NH2) moiety were synthesized and tested for COX-1/COX-2 inhibition in vitro. Many of these 1,1-dioxo-2,3- dihydrobenzo(d)isothiazolyl substituted 1,5-diarylpyrazoles, where the SO2NH2 group was a part of the fused ring, showed COX inhibitory activity. Few of them were identified as selective COX-2 inhibitors. Structure Activity Relationship study within the series are discussed.

Ragaglitazar [(−) DRF 2725; NNC 61-0029] is a coligand of PPARα and PPARγ. In ob/ob mice, ragaglitazar showed significant reduction in plasma glucose, triglyceride and insulin (ED50 values

A number of naphthofuranones were synthesized and tested for COX-1 and COX-2 inhibition. Few of them were identified as selective COX-2 inhibitors. Structure-activity relationship studies within the series are discussed.

Objective: Preclinical evaluation of DRF 2655, a peroxisome proliferator-activated receptor alpha (PPARα) and PPARγ agonist, as a body-weight lowering, hypolipidemic and euglycemic agent. Research Methods and Procedures: DRF 2655 was studied in different genetic, normal, and hyperlipidemic animal models. HEK 293 cells were used to conduct the reporter-based transactivation of PPARα and PPARγ. To understand the biochemical mechanism of lipid-, body-weight-, and glucose-lowering effects, activities of key β-oxidation and lipid catabolism enzymes and gluconeogenic enzymes were studied in db/db mice treated with DRF 2655. 3T3L1 cells were used for adipogenesis study, and HepG2 cells were used to study the effect of DRF 2655 on total cholesterol and triglyceride synthesis using [14C]acetate and [3H]glycerol. Results: DRF 2655 showed concentration-dependent transactivation of PPARα and PPARγ. In the 3T3L1 cell-differentiation study, DRF 2655 and rosiglitazone showed 369% and 471% increases, respectively, in triglyceride accumulation. DRF 2655 showed body-weight lowering and euglycemic and hypolipidemic effects in various animal models. db/db mice treated with DRF 2655 showed 5- and 3.6-fold inhibition in phosphoenolpyruvate carboxykinase and glucose 6-phosphatase activity and 651% and 77% increases in the β-oxidation enzymes carnitine palmitoyltransferase and carnitine acetyltransferase, respectively. HepG2 cells treated with DRF 2655 showed significant reduction in lipid synthesis. Discussion: DRF 2655 showed excellent euglycemic and hypolipidemic activities in different animal models. An exciting finding is its body-weight lowering effect in these models, which might be mediated by the induction of target enzymes involved in hepatic lipid catabolism through PPARα activation.

Peroxisome proliferator-activated receptor (PPAR)-binding protein (PBP) is an important coactivator for PPARgamma and other transcription factors. PBP is an integral component of a multiprotein thyroid hormone receptor-associated protein (TRAP)/vitamin D(3) receptor-interacting protein (DRIP)/activator-recruited cofactor (ARC) complex required for transcriptional activity. To study the regulation of PBP by cellular signaling pathways, we identified the phosphorylation sites of PBP. Using a combination of in vitro and in vivo approaches and mutagenesis of PBP phosphorylation sites, we identified six phosphorylation sites on PBP: one exclusive protein kinase A (PKA) phosphorylation site at serine 656, two protein kinase C (PKC) sites at serine 796 and serine 1345, a common PKA/PKC site at serine 756, and two extracellular signal-regulated kinase 2 sites of the mitogen-activated protein kinase (MAPK) family at threonine 1017 and threonine 1444. Binding of PBP to PPARgamma1 or retinoid-X-receptor for 9-cis-retinoic acid (RXR) is independent of their phosphorylation states, implying no changes in protein-protein interaction after modification by phosphorylation. Overexpression of RafBXB, an activated upstream kinase of the MAPK signal transduction pathway, exerts a significant additive inductive effect on PBP coactivator function. This effect is significantly diminished by overexpression of RafBXB301, a dominant negative mutant of RafBXB. These results identify phosphorylation as a regulatory modification event of PBP and demonstrate that PBP phosphorylation by Raf/MEK/MAPK cascade exerts a positive effect on PBP coactivator function. The functional role of PKA and PKC phosphorylation sites in PBP remains to be elucidated.

Statins are a class of oral drugs that are widely used for treatment of hypercholesterolemia. Recent clinical data suggest that chronic use of these drugs increases the frequency of new onset diabetes. Studies to define the risks of statin-induced diabetes and its underlying mechanisms are clearly necessary. We explored the possible mechanism of statin induced insulin resistance using a well-established cell based model and simvastatin as a prototype statin. Our data show that simvastatin induces insulin resistance in a cholesterol biosynthesis inhibition independent fashion but does so by a fatty acid mediated effect on insulin signaling pathway. These data may help design strategies for prevention of statin induced insulin resistance and diabetes in patients with hypercholesterolemia.

We report here the synthesis of a series of 5-[4-[2-[substituted phthalazinones-2(or 4)yl]ethoxy]phenylmethyl]thiazolidine-2,4-diones and 5-[4-[2-[2,3-benzoxazine-4-one-2-yl]ethoxy]phenylmethyl]thiazolidine-2,4-diones and their plasma glucose and plasma triglyceride lowering activity in db/db mice. In vitro PPARgamma transactivation assay was performed in HEK 293T cells. In vitro and in vivo pharmacological studies showed that the phthalazinone analogue has better activity. PHT46 (compound 5a), the best compound in this series, showed better in vitro PPARgamma transactivation potential than troglitazone and pioglitazone. In insulin resistant db/db mice, PHT46 showed better plasma glucose and triglyceride lowering activity than the standard drugs. Pharmacokinetic study in Wistar rats showed good systemic exposure of PHT46. Subchronic toxicity study in Wistar rats did not show any treatment-related adverse effect.

The phosphate-specific transporter (Pst) in bacteria is a multi-subunit system which belongs to the ABC family of transporters. The gene forms part of an operon and it is involved in phosphate uptake in prokaryotes. Its import function is known to be operative only under conditions of phosphate starvation. However, we found overexpression of this transporter in a Mycobacterium smegmatis strain selected for ciprofloxacin resistance (CIPr) which was grown under conditions in which the phosphate-scavenging function of this operon was inoperative. In CIPr cells, active efflux of the drug plays a predominant role in conferring high levels of fluoroquinolone resistance. We therefore investigated the role of this transporter in the process of efflux-mediated drug resistance by inactivating the pst operon in the CIPr strain. Phenotypic characterization of the resulting strain, CIPrd, showed a striking reduction in the minimal inhibitory concentration (MIC) of ciprofloxacin and in the drug extrusion profile as well. Genotype analysis, on the other hand, revealed partial disruption of the pst operon in CIPrd as a consequence of transporter gene amplification. Furthermore, disruption of this operon in wild-type cells resulted in hypersensitivity to ciprofloxacin and other xenobiotics to which CIPr cells exhibited cross-resistance. Thus our results provide strong evidence that Pst is a natural membrane transport system that has the ability to promote drug efflux in addition to its phosphate-scavenging function in the CIPr strain.

Peroxisomes are subcellular organelles that are found in the cytoplasm of most animal cells. They perform diverse metabolic functions, including H2O2-derived respiration, β-oxidation of fatty acids, and cholesterol metabolism. Peroxisome proliferators are a large class of structurally dissimilar industrial and pharmaceutical chemicals that were originally identified as inducers of both the size and the number of peroxisomes in rat and mouse livers or hepatocytes in vitro. Exposure to peroxisome proliferators leads to a stereotypical orchestration of adaptations consisting of hepatocellular hypertrophy and hyperplasia, and transcriptional induction of fatty acid metabolizing enzymes regulated in parallel with peroxisome proliferation. Chronic exposure to peroxisome proliferators causes liver tumors in both male and female mice and rats. Evidence indicates a pivotal role for a subset of nuclear receptor superfamily members, called peroxisome proliferator-activated receptors (PPARs), in mediating energy metabolism. Upon activation, PPARs regulate the expression of genes involved in lipid metabolism and peroxisome proliferation, as well as genes involved in cell growth. In this review, we describe the molecular mode of action of PPAR transcription factors, including ligand binding, interaction with specific DNA response elements, transcriptional activation, and cross talk with other signaling pathways. We discuss the evidence that suggests that PPARα and transcriptional coactivator Med1/PBP, a key subunit of the Mediator complex play a central role in mediating hepatic steatosis to hepatocarcinogenesis. Disproportionate increases in H2O2-generating enzymes generates excess reactive oxygen species resulting in sustained oxidative stress and progressive endoplasmic reticulum (ER) stress with activation of unfolded protein response signaling. Thus, these major contributors coupled with hepatocellular proliferation are the key players of peroxisome proliferators-induced hepatocarcinogenesis.

A series of novel alkynyl substituted 3,4-dihydropyrimidin-2(1H)-one (DHPM) derivatives were designed, synthesized and evaluated in vitro as potential inhibitors of chorismate mutase (CM). All these compounds were prepared via a multi-component reaction (MCR) involving sequential I2-mediated Biginelli reaction followed by Cu-free Sonogashira coupling. Some of them showed promising inhibitory activities when tested at 30 μM. One compound showed dose dependent inhibition of CM with IC50 value of 14.76 ± 0.54 μM indicating o-alkynylphenyl substituted DHPM as a new scaffold for the discovery of promising inhibitors of CM.

The acetylation of indole (IIIe) in the presence of AlCl3 leads to desired 3-acetyl derivative (VII), whilst the analogous reaction of 2-tert-butylindoles (IIIa)—(IIId) is accompanied by an unexpected migration of the sulfonyl group.

Peroxisomes are subcellular organelles that are found in the cytoplasm of most animal cells. They perform diverse metabolic functions, including H2O2-derived respiration, β-oxidation of fatty acids, and cholesterol metabolism. Peroxisome proliferators are a large class of structurally dissimilar industrial and pharmaceutical chemicals that were originally identified as inducers of both the size and the number of peroxisomes in rat and mouse livers or hepatocytes in vitro. Exposure to peroxisome proliferators leads to a stereotypical orchestration of adaptations consisting of hepatocellular hypertrophy and hyperplasia, and transcriptional induction of fatty acid metabolizing enzymes regulated in parallel with peroxisome proliferation. Chronic exposure to peroxisome proliferators causes liver tumors in both male and female mice and rats. Evidence indicates a pivotal role for a subset of nuclear receptor superfamily members, called peroxisome proliferator-activated receptors (PPARs), in mediating energy metabolism. Upon activation, PPARs regulate the expression of genes involved in lipid metabolism and peroxisome proliferation, as well as genes involved in cell growth. In this review, we describe the molecular mode of action of PPAR transcription factors, including ligand binding, interaction with specific DNA response elements, transcriptional activation, and cross talk with other signaling pathways. We discuss the evidence that suggests that PPARα and transcriptional coactivator Med1/PBP, a key subunit of the Mediator complex play a central role in mediating hepatic steatosis to hepatocarcinogenesis. Disproportionate increases in H2O2-generating enzymes generates excess reactive oxygen species resulting in sustained oxidative stress and progressive endoplasmic reticulum (ER) stress with activation of unfolded protein response signaling. Thus, these major contributors coupled with hepatocellular proliferation are the key players of peroxisome proliferators-induced hepatocarcinogenesis.

A rapid and direct access to N-aryl substituted fused triazinone derivatives is accomplished via Cu-mediated N-arylation.

A series of novel N-aryl substituted thieno[2,3-d]pyrimidin-4(3H)-ones were designed and synthesized as potential inhibitors of chorismate mutase. Synthesis of this class of compounds was carried out by using Cu-mediated C-N bond forming reaction between thieno[2,3-d]pyrimidin-4(3H)-ones and aryl boronic acids. The reaction can be performed in an open flask as the conversion was found to be not sensitive to the presence of air or atmospheric moisture. A range of compounds were prepared by using this method and single crystal X-ray diffraction study was performed using a representative compound. In vitro pharmacological data of some of the compounds synthesized along with dose response studies using active molecules are presented. In silico interactions of these molecules with chorismate mutase are also presented.

A new compound library based on the 2-aminochromene framework is synthesized via a novel strategy involving a multi-component reaction and Pd-catalyzed C—C bond forming reaction.

A rapid and direct access to N-aryl substituted fused triazinone derivatives has been accomplished via N-arylation of 1,2,3-triazin-4-one ring involving a Cu-mediated coupling between triazinone derivatives and aryl boronic acids. A combination of Cu(OAc)(2)-Et(3)N in 1,2-dichloroethane was found to be effective and various fused triazinone derivatives have been prepared by using this methodology. Molecular structure of a representative compound was confirmed by single crystal X-ray diffraction study. The scope and limitations of this reaction is discussed. Some of the compounds synthesized were tested for chorismate mutase inhibitory properties in vitro. The in vitro dose response study of an active compound is presented.

A multi component based synthesis involving palladium catalyzed C-C bond forming reaction has been developed as a new strategy to access systematically modified functionalized 2-aminochromenes. This MCR involves the use of bromobenzaldehyde as a key component and is highlighted by generating a new compound library. Many of these compounds showed Mycobacterium tuberculosis H37Rv chorismate mutase inhibiting properties in vitro representing the lead example of chorismate mutase inhibition by heteroarene based compounds.

Type 2 diabetes is characterized by abnormal metabolism of glucose and fat, due in part to resistance to the actions of insulin in peripheral tissues. If untreated it leads to several complications such as blindness, kidney failure, neuropathy and amputations. The benefit of exercise in diabetic patients is well known and recent research indicates that AMP activated protein kinase (AMPK) plays a major role in this exercise related effect. AMPK is considered as a master switch regulating glucose and lipid metabolism. The AMPK is an enzyme that works as a fuel gauge, being activated in conditions of high energy phosphate depletion. AMPK is also activated robustly by skeletal muscle contraction and myocardial ischaemia, and is involved in the stimulation of glucose transport and fatty acid oxidation produced by these stimuli. In liver, activation of AMPK results in enhanced fatty acid oxidation and decreased production of glucose, cholesterol, and triglycerides. The two leading diabetic drugs namely, metformin and rosiglitazone, show their metabolic effects partially through AMPK. These data, along with evidence from studies showing that chemical activation of AMPK in vivo with 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR) improves blood glucose concentrations and lipid profiles, make this enzyme an attractive pharmacological target for the treatment of type 2 diabetes and other metabolic disorders.

Several chemical modifications in the N 1-benzenesulfonamide ring of celecoxib are presented. The series with a hydroxymethyl group adjacent to the sulfonamide was found to be the most potent modification that yielded many compounds selectively active against COX-2 enzyme in vitro. ? 2003 Elsevier Ltd. All rights reserved.

We investigated the biological activity of Dr. Reddy's Research Foundation (DRF) 2519, a benzoxazinone analogue of the thiazolidinedione class of compounds. In the in vitro transactivation assay, DRF 2519 showed interesting dual activation of Peroxisome Proliferator Activated Receptor (PPAR) alpha and gamma. In insulin-resistant ob/ob mouse model, DRF 2519 showed significant alleviation of insulin resistance and dyslipidemia, which is better than rosiglitazone. Fatty Zucker rats treated with DRF 2519 showed better reduction of plasma insulin, triglyceride and free fatty acid levels than those treated with rosiglitazone. In addition, these rats were able to clear plasma lipids better when challenged with exogenous lipid (i.v.). DRF 2519 treatment resulted in improved plasma lipid profiles in high-fat-fed Sprague-Dawley rats. Treated rats showed better plasma lipid clearance and hepatic triglyceride secretion. When compared to DRF 2519, fenofibrate was comparatively less efficacious while rosigltiazone showed no activity in these models. In ex vivo studies, DRF 2519 showed induction of liver acyl CoA oxidase mRNA and increase in lipoprotein lipase (LPL) protein expression and activity in adipose tissue. In the in vitro studies, DRF 2519 inhibited the lipid biosynthesis and secretion of apolipoprotein B from human hepatoma (Hep)G2 cells. It also enhanced insulin-induced relaxation of rat aortic smooth muscle. These results indicate that DRF 2519, a dual activator of PPAR-alpha and gamma, could be an interesting development candidate in the management of metabolic disorders and associated complications.

Ragaglitazar [(-) DRF 2725; NNC 61-0029] is a coligand of PPARalpha and PPARgamma. In ob/ob mice, ragaglitazar showed significant reduction in plasma glucose, triglyceride and insulin (ED50 values0.03, 6.1 and0.1 mg kg-1). These effects are three-fold better than rosiglitazone and KRP-297. In Zucker fa/fa rats, ragaglitazar showed dose-dependent reduction in triglyceride and insulin, hepatic triglyceride secretion and triglyceride clearance kinetics (maximum of 74, 53, 32 and 50% at 3 mg kg-1), which are better than rosiglitazone and KRP-297. In a high-fat-fed hyperlipidaemic rat model, the compound showed an ED50 of 3.95, 3.78 mg kg-1 for triglyceride and cholesterol lowering, and 0.29 mg kg-1 for HDL-C increase. It also showed improvement in clearance of plasma triglyceride and hepatic triglyceride secretion rate. All these effects are 3-10-fold better than fenofibrate and KRP-297. Ragaglitazar treatment showed significant reduction in plasma Apo B and Apo CIII levels, and increase in liver CPT1 and CAT activity and ACO mRNA. Significant increase of both liver and fat LPL activity and fat aP2 mRNA was also observed. In a high-fat-fed hamster model, ragaglitazar at 1 mg kg-1 showed 83 and 61% reduction in triglyceride and total cholesterol, and also 17% reduction in fat feed-induced body weight increase. In these hyperlipidaemic animal models, PPARgamma ligands failed to show any significant efficacy. Taken together, ragaglitazar shows better insulin-sensitizing and lipid-lowering potential, as compared to the standard compounds.