Discovery of AMP Mimetics that Exhibit High Inhibitory Potency and Specificity for AMP Deaminase

Erion, Mark D.; Kasibhatla, Srinivas Rao; Bookser, Brett C.; Poelje, Paul D. van; Reddy, M. Rami; Gruber, and Harry E.; Appleman, James R.

doi:10.1021/ja983153j

Cited by 33 publications

(48 citation statements)

References 58 publications

(56 reference statements)

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“…Other difficulties that need to be overcome include the hydrophilic nature of AMP sites and their reliance on the negatively charged phosphate group of AMP for binding affinity 13. Target-based virtual database screening has become a useful tool for the identification of inhibitors for protein-ligand and protein-protein interactions 14.…”

Section: Introductionmentioning

confidence: 99%

A library of novel allosteric inhibitors against fructose 1,6-bisphosphatase

Heng

Gryncel

Kantrowitz

2009

Bioorganic & Medicinal Chemistry

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The identification of a proper lead compound for fructose 1,6-bisphosphatase (FBPase) is a critical step in the process of developing novel therapeutics against type-2 diabetes. Herein, we have successfully generated a library of allosteric inhibitors against FBPase as potential anti-diabetic drugs, of which, the lead compound 1b was identified through utilizing a virtual high-throughput screening (vHTS) system, which we have developed. The thiazole-based core structure was synthesized via the condensation of α-bromo-keotnes with thioureas and substituents on the two aryl rings were varied. 4c was found to inhibit pig kidney FBPase approximately 5-fold better than 1b. In addition, we have also identified 10b, a tight binding fragment, which can be use for fragment-based drug design purposes.

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Section: Introductionmentioning

confidence: 99%

A library of novel allosteric inhibitors against fructose 1,6-bisphosphatase

Heng

Gryncel

Kantrowitz

2009

Bioorganic & Medicinal Chemistry

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“…We focused on the AMP binding site largely because we anticipated difficulties in discovering FBPase inhibitors that bind to the highly hydrophilic substrate binding site with sufficient affinity to compete with the buildup of intrahepatic fructose 1,6-bisphosphate. Targeting the AMP binding site is associated with its own set of well recognized design challenges arising from the hydrophilic nature of AMP sites, their reliance on the negatively charged phosphate group of AMP for binding affinity (21), and the abundance of AMP-binding enzymes controlling key biosynthetic pathways. Previous efforts targeting the AMP site identified several weak FBPase inhibitors (22), including the nonspecific inhibitor aminoimidazole-4-carboxamide riboside monophosphate (ZMP) (23,24), which lowered glucose and raised lactate levels after reaching millimolar liver levels following systemic administration of aminoimidazole-4-carboxamide riboside to normal mice at high doses (23).…”

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confidence: 99%

“…AMP deaminase (porcine heart) was purified and assayed as described in ref. 21. Glycogen phosphorylase (rabbit muscle), phosphofructokinase (rabbit liver), and adenylate kinase (rabbit muscle) were obtained from SigmaAldrich and assayed as described in refs.…”

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confidence: 99%

MB06322 (CS-917): A potent and selective inhibitor of fructose 1,6-bisphosphatase for controlling gluconeogenesis in type 2 diabetes

Erion

Poelje

Dang

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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145

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In type 2 diabetes, the liver produces excessive amounts of glucose through the gluconeogenesis (GNG) pathway and consequently is partly responsible for the elevated glucose levels characteristic of the disease. In an effort to find safe and efficacious GNG inhibitors, we targeted the AMP binding site of fructose 1,6-bisphosphatase (FBPase). The hydrophilic nature of AMP binding sites and their widespread use for allosteric regulation of enzymes in metabolic pathways has historically made discovery of AMP mimetics suitable for drug development difficult. By using a structure-based drug design strategy, we discovered a series of compounds that mimic AMP but bear little structural resemblance. The lead compound, MB05032, exhibited high potency and specificity for human FBPase. Oral delivery of MB05032 was achieved by using the bisamidate prodrug MB06322 (CS-917), which is converted to MB05032 in two steps through the action of an esterase and a phosphoramidase. MB06322 inhibited glucose production from a variety of GNG substrates in rat hepatocytes and from bicarbonate in male Zucker diabetic fatty rats. Analysis of liver GNG pathway intermediates confirmed FBPase as the site of action. Oral administration of MB06322 to Zucker diabetic fatty rats led to a dose-dependent decrease in plasma glucose levels independent of insulin levels and nutritional status. Glucose lowering occurred without signs of hypoglycemia or significant elevations in plasma lactate or triglyceride levels. The findings suggest that potent and specific FBPase inhibitors represent a drug class with potential to treat type 2 diabetes through inhibition of GNG.endogenous glucose production ͉ AMP mimetic ͉ structure-based drug design ͉ phosphonate prodrug ͉ antihyperglycemic T he liver (1, 2) and, to a lesser extent, the kidneys (3) are the primary organs responsible for endogenous glucose production (EGP). In type 2 diabetes mellitus (T2DM), excessive EGP in the fasted state and fed state contributes to the chronic elevation of blood glucose levels found in patients with advanced (4, 5) and mild diabetes (6, 7). Moreover, fasting plasma glucose levels correlate with EGP rates in patients with fasting plasma glucose Ͼ180 mg͞dl (10 mM) (8) and possibly in patients with lower levels (6, 7). Studies using 13 C NMR spectroscopy (9) as well as more recent studies using deuterated water (7, 10) attribute the excessive EGP in T2DM patients to increased flux through the gluconeogenesis (GNG) pathway.Efforts over the past 40 years to discover inhibitors of GNG produced few safe and effective drug candidates (11). Metformin, the only marketed drug that acts, at least partially, through inhibition of GNG (12), inhibits GNG indirectly and only 33-36% at the rarely prescribed maximal human dose (13). Direct GNG inhibitors show more pronounced glucose lowering in animals but not without eliciting safety-related concerns. Hypoglycemia, lactic acidosis, and hypertriglyceridemia are the principle safety risks associated with GNG inhibition as highlighted in studie...

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“…Further studies are needed to clarify the molecular mechanisms how AMPD1 levels and concomitantly modified cellular adenine nucleotide levels regulate the expression levels of mTOR and related molecules and the formation of mTORC1. Studies using in vitro culture system in combination with overexpression and knockdown of Ampd1 gene and pharmacological intervention using AMP mimetics [ 15 ] could be useful for this purpose. We are currently studying in this context.…”

Section: Discussionmentioning

confidence: 99%

AMPD1 regulates mTORC1-p70 S6 kinase axis in the control of insulin sensitivity in skeletal muscle

et al. 2015

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BackgroundInsulin resistance triggered by excess fat is a key pathogenic factor that promotes type 2 diabetes. Understanding molecular mechanisms of insulin resistance may lead to the identification of a novel therapeutic target for type 2 diabetes. AMPD1, an isoform of AMP deaminase (AMPD), is suggested to play roles in the regulation of glucose metabolism through controlling AMP-activated protein kinase (AMPK) activation. We reported that the diet-induced insulin resistance was improved in AMPD1-deficient mice compared to wild type mice. To further delineate this observation, we studied changes of insulin signaling in skeletal muscle of wild type (WT) and AMPD1-deficient mice.MethodsPhosphorylation levels of kinases and expression levels of mTOR components were quantified by immunoblotting using protein extracts from tissues. The interaction between mTOR and Raptor was determined by immunoblotting of mTOR immunoprecipitates with anti-Raptor antibody. Gene expression was studied by quantitative PCR using RNA extracted from tissues.ResultsPhosphorylation levels of AMPK, Akt and p70 S6 kinase in skeletal muscle were higher in AMPD1-deficient mice compared to WT mice after high fat diet challenge, while they did not show such difference in normal chow diet. Also, no significant changes in phosphorylation levels of AMPK, Akt or p70 S6 kinase were observed in liver and white adipose tissue between WT and AMPD1-deficient mice. The expression levels of mTOR, Raptor and Rictor tended to be increased by AMPD1 deficiency compared to WT after high fat diet challenge. AMPD1 deficiency increased Raptor-bound mTOR in skeletal muscle compared to WT after high fat diet challenge. Gene expression of peroxisome proliferator-activated receptor-γ coactivator 1α and β, downstream targets of p70 S6 kinase, in skeletal muscles was not changed significantly by AMPD1 deficiency compared to the wild type after high fat diet challenge.ConclusionThese data suggest that AMPD1 deficiency activates AMPK/Akt/mTORC1/p70 S6 kinase axis in skeletal muscle after high fat diet challenge, but not in normal chow diet. These changes may contribute to improve insulin resistance.

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Discovery of AMP Mimetics that Exhibit High Inhibitory Potency and Specificity for AMP Deaminase

Cited by 33 publications

References 58 publications

A library of novel allosteric inhibitors against fructose 1,6-bisphosphatase

A library of novel allosteric inhibitors against fructose 1,6-bisphosphatase

MB06322 (CS-917): A potent and selective inhibitor of fructose 1,6-bisphosphatase for controlling gluconeogenesis in type 2 diabetes

AMPD1 regulates mTORC1-p70 S6 kinase axis in the control of insulin sensitivity in skeletal muscle

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