Stimulation of Hepatocyte Glucose Metabolism by Novel Small Molecule Glucokinase Activators

Abstract

Glucokinase (GK) has a major role in the control of blood glucose homeostasis and is a strong potential target for the pharmacological treatment of type 2 diabetes. We report here the mechanism of action of two novel and potent direct activators of GK: 6-[(3-isobutoxy-5-isopropoxybenzoyl)amino]nicotinic acid (GKA1) and 5-({3-isopropoxy-5-[2-(3-thienyl)ethoxy] benzoyl}amino)-1,3,4-thiadiazole-2-carboxylic acid (GKA2), which increase the affinity of GK for glucose by 4-and 11-fold, respectively. GKA1 increased the affinity of GK for the competitive inhibitor mannoheptulose but did not affect the affinity for the inhibitors palmitoylCoA and the endogenous 68-kDa regulator (GK regulatory protein [GKRP]), which bind to allosteric sites or to N-acetylglucosamine, which binds to the catalytic site. In hepatocytes, GKA1 and GKA2 stimulated glucose phosphorylation, glycolysis, and glycogen synthesis to a similar extent as sorbitol, a precursor of fructose 1-phosphate, which indirectly activates GK through promoting its dissociation from GKRP. Consistent with their effects on isolated GK, these compounds also increased the affinity of hepatocyte metabolism for glucose. GKA1 and GKA2 caused translocation of GK from the nucleus to the cytoplasm. This effect was additive with the effect of sorbitol and is best explained by a "glucose-like" effect of the GK activators in translocating GK to the cytoplasm. In conclusion, GK activators are potential antihyperglycemic agents for the treatment of type 2 diabetes through the stimulation of hepatic glucose metabolism by a mechanism independent of GKRP. Diabetes 53:535-541, 2004

Upregulation of β-cell genes and improved function in rodent islets following chronic glucokinase activation

“…Although we did not measure GK activity of islets, we would predict that chronic treatment with GKA would increase enzyme activity. This would be consistent with the effects of GKAs measured in experiments assessing direct activation of the isolated GK enzyme, albeit in hepatocytes (Brocklehurst et al 2004).…”

“…GK was assayed at its S 0 . 5 glucose concentration as described previously (Brocklehurst et al 2004).…”

“…Each was expressed in Escherichia coli with an NH 2 -terminal 6His tag and purified using Ni-NTA chromatography. Recombinant proteins were O95% pure as assessed by SDS/PAGE analysis (Brocklehurst et al 2004). GK was assayed at its S 0 .…”

“…A number of small molecule GK activators (GKAs) have been described. These agents have effects on hepatic glucose metabolism and insulin secretion, both in vitro and in normal and diabetic animal models (Grimsby et al 2003, Brocklehurst et al 2004, Kamata et al 2004, Efanov et al 2005, Coope et al 2006, Futamura et al 2006, Matschinsky et al 2006, Johnson et al 2007, Coghlan & Leighton 2008.…”

5-Aminoimidazole-4-Carboxamide-1-β-d-Ribofuranoside and Metformin Inhibit Hepatic Glucose Phosphorylation by an AMP-Activated Protein Kinase–Independent Effect on Glucokinase Translocation

“…Finally, it has been recently proposed that specific glucokinase activators could be potent tools to reduce hyperglycemia in type 2 diabetic patients, partly by stimulating liver glucose uptake (46,47). However, the role of glucokinase in the control of glucose homeostasis is not only quantitative but also qualitative due to the glucoseinduced transcriptional effects that result from the intrahepatic accumulation of Glc-6-P (48) and should also be taken into account.…”