The C-aryl glucoside 6 (dapagliflozin) was identified as a potent and selective hSGLT2 inhibitor which reduced blood glucose levels in a dose-dependent manner by as much as 55% in hyperglycemic streptozotocin (STZ) rats. These findings, combined with a favorable ADME profile, have prompted clinical evaluation of dapagliflozin for the treatment of type 2 diabetes.

OBJECTIVE-The inhibition of gut and renal sodium-glucose cotransporters (SGLTs) has been proposed as a novel therapeutic approach to the treatment of diabetes. We have identified dapagliflozin as a potent and selective inhibitor of the renal sodium-glucose cotransporter SGLT2 in vitro and characterized its in vitro and in vivo pharmacology.RESEARCH DESIGN AND METHODS-Cell-based assays measuring glucose analog uptake were used to assess dapagliflozin's ability to inhibit sodium-dependent and facilitative glucose transport activity. Acute and multi-dose studies in normal and diabetic rats were performed to assess the ability of dapagliflozin to improve fed and fasting plasma glucose levels. A hyperinsulinemic-euglycemic clamp study was performed to assess the ability of dapagliflozin to improve glucose utilization after multi-dose treatment.RESULTS-Dapagliflozin potently and selectively inhibited human SGLT2 versus human SGLT1, the major cotransporter of glucose in the gut, and did not significantly inhibit facilitative glucose transport in human adipocytes. In vivo, dapagliflozin acutely induced renal glucose excretion in normal and diabetic rats, improved glucose tolerance in normal rats, and reduced hyperglycemia in Zucker diabetic fatty (ZDF) rats after single oral doses ranging from 0.1 to 1.0 mg/kg. Once-daily dapagliflozin treatment over 2 weeks significantly lowered fasting and fed glucose levels at doses ranging from 0.1 to 1.0 mg/kg and resulted in a significant increase in glucose utilization rate accompanied by a significant reduction in glucose production.CONCLUSIONS-These data suggest that dapagliflozin has the potential to be an efficacious treatment for type 2 diabetes.

SGLT2 (for “Sodium GLucose coTransporter” protein 2) is the major protein responsible for glucose reabsorption in the kidney and its inhibition has been the focus of drug discovery efforts to treat type 2 diabetes. In order to better clarify the human tissue distribution of expression of SGLT2 and related members of this cotransporter class, we performed TaqMan™ (Applied Biosystems, Foster City, CA, USA) quantitative polymerase chain reaction (PCR) analysis of SGLT2 and other sodium/glucose transporter genes on RNAs from 72 normal tissues from three different individuals. We consistently observe that SGLT2 is highly kidney specific while SGLT5 is highly kidney abundant; SGLT1, sodium-dependent amino acid transporter (SAAT1), and SGLT4 are highly abundant in small intestine and skeletal muscle; SGLT6 is expressed in the central nervous system; and sodium myoinositol cotransporter is ubiquitously expressed across all human tissues.Electronic Supplementary MaterialSupplementary material is available for this article at 10.1007/s13300-010-0006-4 and is accessible for authorized users.

The importance of glucokinase (GK; EC 2.7.1.12) in glucose homeostasis has been demonstrated by the association of GK mutations with diabetes mellitus in humans and by alterations in glucose metabolism in transgenic and gene knockout mice. Liver GK activity in humans and rodents is allosterically inhibited by GK regulatory protein (GKRP). To further understand the role of GKRP in GK regulation, the mouse GKRP gene was inactivated. With the knockout of the GKRP gene, there was a parallel loss of GK protein and activity in mutant mouse liver. The loss was primarily because of posttranscriptional regulation of GK, indicating a positive regulatory role for GKRP in maintaining GK levels and activity. As in rat hepatocytes, both GK and GKRP were localized in the nuclei of mouse hepatocytes cultured in low-glucose-containing medium. In the presence of fructose or high concentrations of glucose, conditions known to relieve GK inhibition by GKRP in vitro, only GK was translocated into the cytoplasm. In the GKRP-mutant hepatocytes, GK was not found in the nucleus under any tested conditions. We propose that GKRP functions as an anchor to sequester and inhibit GK in the hepatocyte nucleus, where it is protected from degradation. This ensures that glucose phosphorylation is minimal when the liver is in the fasting, glucose-producing phase. This also enables the hepatocytes to rapidly mobilize GK into the cytoplasm to phosphorylate and store or metabolize glucose after the ingestion of dietary glucose. In GKRP-mutant mice, the disruption of this regulation and the subsequent decrease in GK activity leads to altered glucose metabolism and impaired glycemic control. G lucokinase (GK; EC 2.7.1.12), the principal hexokinase in liver parenchymal cells and in pancreatic ␤ cells, is a critical component of the physiological glucose-sensing apparatus (1). In humans, GK heterozygous mutations lead to the autosomal, dominant, maturity-onset diabetes of the young (MODY) (2, 3) phenotype. In mice, changes in GK activity by gene knockout and by overexpression resulted in altered glucose homeostasis and demonstrated that modest changes in liver GK activity alone were sufficient to cause significant alterations in blood glucose levels (4-9). Liver GK activity has been reported to be lower in some obese humans with type 2 diabetes mellitus when compared with nondiabetic normal weight or obese individuals, suggesting a role for liver GK in glucose homeostasis in type 2 diabetes (10).In the liver, GK activity is allosterically inhibited by GK regulatory protein (GKRP) (11-13). This protein, found in the livers of all animal species where GK is present, shows no inhibitory effect on other known hexokinases (11-13). In rodents, GKRP inhibition of GK is relieved by high concentrations of glucose and by fructose 1-phosphate, and is potentiated by fructose 6-phosphate (11-13). Recent studies have demonstrated that both GK and GKRP are in the nucleus of rat hepatocytes cultured in a medium containing 5.5 mM glucose (14,15). When the growth medium w...

Dapagliflozin, a first-in-class, selective inhibitor of sodium-glucose cotransporter 2 (SGLT2), promotes urinary glucose excretion to reduce hyperglycemia for the treatment of type 2 diabetes. A series of nonclinical studies were undertaken to evaluate dapagliflozin in species where it was shown to have pharmacologic activity comparable with that in humans at doses that resulted in supratherapeutic exposures. In vitro screening (>300 targets; 10 μmol/L) indicated no significant off-target activities for dapagliflozin or its primary human metabolite. Once daily, orally administered dapagliflozin was evaluated in Sprague-Dawley rats (≤6 months) and in beagle dogs (≤1 year) at exposures >5000-fold those observed at the maximum recommended human clinical dose (MRHD; 10 mg). Anticipated, pharmacologically mediated effects of glucosuria, osmotic diuresis, and mild electrolyte loss were observed, but there were no adverse effects at clinically relevant exposures, including in the kidneys or urogenital tract. The SGLT2-/- mice, which show chronic glucosuria, and dapagliflozin-treated, wild-type mice exhibited similar safety profiles. In rats but not dogs, dapagliflozin at >2000-fold MRHD exposures resulted in tissue mineralization and trabecular bone accretion. Investigative studies suggested that the effect was not relevant to human safety, since it was partially related to off-target inhibition of SGLT1, which was observed only at high doses of dapagliflozin and resulted in intestinal glucose malabsorption and increased intestinal calcium absorption. The rigorous assessment of supra- and off-target dapagliflozin pharmacology in nonclinical species allowed for a thorough evaluation of potential toxicity, providing us with confidence in its safety in patients with diabetes.