Fructose-1, 6-bisphosphatase inhibitors for reducing excessive endogenous glucose production in type 2 diabetes.

Fructose-1,6-bisphosphatase (FBPase), a rate-controlling enzyme of gluconeogenesis, has emerged as an important target for the treatment of type 2 diabetes due to the well-recognized role of excessive endogenous glucose production (EGP) in the hyperglycemia characteristic of the disease. Inhibitors of FBPase are expected to fulfill an unmet medical need because the majority of current antidiabetic medications act primarily on insulin resistance or insulin insufficiency and do not reduce gluconeogenesis effectively or in a direct manner. Despite significant challenges, potent and selective inhibitors of FBPase targeting the allosteric site of the enzyme were identified by means of a structure-guided design strategy that used the natural inhibitor, adenosine monophosphate (AMP), as the starting point. Oral delivery of these anionic FBPase inhibitors was enabled by a novel diamide prodrug class. Treatment of diabetic rodents with CS-917, the best characterized of these prodrugs, resulted in a reduced rate of gluconeogenesis and EGP. Of note, inhibition of gluconeogenesis by CS-917 led to the amelioration of both fasting and postprandial hyperglycemia without weight gain, incidence of hypoglycemia, or major perturbation of lactate or lipid homeostasis. Furthermore, the combination of CS-917 with representatives of the insulin sensitizer or insulin secretagogue drug classes provided enhanced glycemic control. Subsequent clinical evaluations of CS-917 revealed a favorable safety profile as well as clinically meaningful reductions in fasting glucose levels in patients with T2DM. Future trials of MB07803, a second generation FBPase inhibitor with improved pharmacokinetics, will address whether this novel class of antidiabetic agents can provide safe and long-term glycemic control.

Discovery of potent and specific fructose-1,6-bisphosphatase inhibitors and a series of orally-bioavailable phosphoramidase-sensitive prodrugs for the treatment of type 2 diabetes.

Excessive glucose production by the liver coupled with decreased glucose uptake and metabolism by muscle, fat, and liver results in chronically elevated blood glucose levels in patients with type 2 diabetes. Efforts to treat diabetes by reducing glucose production have largely focused on the gluconeogenesis pathway and rate-limiting enzymes within this pathway such as fructose-1,6-bisphosphatase (FBPase). The first potent FBPase inhibitors were identified using a structure-guided drug design strategy (Erion, M. D.; et al. J. Am. Chem. Soc. 2007, 129, 15480-15490) but proved difficult to deliver orally. Herein, we report the synthesis and characterization of a series of orally bioavailable FBPase inhibitors identified following the combined discoveries of a low molecular weight inhibitor series with increased potency and a phosphonate prodrug class suitable for their oral delivery. The lead inhibitor, 10A, was designed with the aid of X-ray crystallography and molecular modeling to bind to the allosteric AMP binding site of FBPase. High potency (IC50 = 16 nM) and FBPase specificity were achieved by linking a 2-aminothiazole with a phosphonic acid. Free-energy perturbation calculations provided insight into the factors that contributed to the high binding affinity. 10A and standard phosphonate prodrugs of 10A exhibited poor oral bioavailability (0.2-11%). Improved oral bioavailability (22-47%) was achieved using phosphonate diamides that convert to the corresponding phosphonic acid by sequential action of an esterase and a phosphoramidase. Oral administration of the lead prodrug, MB06322 (30, CS-917), to Zucker Diabetic Fatty rats led to dose-dependent inhibition of gluconeogenesis and endogenous glucose production and consequently to significant blood glucose reduction.

Inhibition of fructose 1,6-bisphosphatase reduces excessive endogenous glucose production and attenuates hyperglycemia in Zucker diabetic fatty rats.

Gluconeogenesis is increased in type 2 diabetes and contributes significantly to fasting and postprandial hyperglycemia. We recently reported the discovery of the first potent and selective inhibitors of fructose 1,6-bisphosphatase (FBPase), a rate-controlling enzyme of gluconeogenesis. Herein we describe acute and chronic effects of the lead inhibitor, MB06322 (CS-917), in rodent models of type 2 diabetes. In fasting male ZDF rats with overt diabetes, a single dose of MB06322 inhibited gluconeogenesis by 70% and overall endogenous glucose production by 46%, leading to a reduction in blood glucose of >200 mg/dl. Chronic treatment of freely feeding 6-week-old male Zucker diabetic fatty (ZDF) rats delayed the development of hyperglycemia and preserved pancreatic function. Elevation of lactate ( approximately 1.5-fold) occurred after 4 weeks of treatment, as did the apparent shunting of precursors into triglycerides. Profound glucose lowering ( approximately 44%) and similar metabolic ramifications were associated with 2-week intervention therapy of 10-week-old male ZDF rats. In high-fat diet-fed female ZDF rats, MB06322 treatment for 2 weeks fully attenuated hyperglycemia without evidence of metabolic perturbation other than a modest reduction in glycogen stores ( approximately 20%). The studies confirm that excessive gluconeogenesis plays an integral role in the pathophysiology of type 2 diabetes and suggest that FBPase inhibitors may provide a future treatment option.

Discovery of a series of phosphonic acid-containing thiazoles and orally bioavailable diamide prodrugs that lower glucose in diabetic animals through inhibition of fructose-1,6-bisphosphatase.

Oral delivery of previously disclosed purine and benzimidazole fructose-1,6-bisphosphatase (FBPase) inhibitors via prodrugs failed, which was likely due to their high molecular weight (>600). Therefore, a smaller scaffold was desired, and a series of phosphonic acid-containing thiazoles, which exhibited high potency against human liver FBPase (IC(50) of 10-30 nM) and high selectivity relative to other 5'-adenosinemonophosphate (AMP)-binding enzymes, were discovered using a structure-guided drug design approach. The initial lead compound (30j) produced profound glucose lowering in rodent models of type 2 diabetes mellitus (T2DM) after parenteral administration. Various phosphonate prodrugs were explored without success, until a novel phosphonic diamide prodrug approach was implemented, which delivered compound 30j with good oral bioavailability (OBAV) (22-47%). Extensive lead optimization of both the thiazole FBPase inhibitors and their prodrugs culminated in the discovery of compound 35n (MB06322) as the first oral FBPase inhibitor advancing to human clinical trials as a potential treatment for T2DM.

Fructose-1,6-bisphosphatase Inhibitors. 2. Design, synthesis, and structure-activity relationship of a series of phosphonic acid containing benzimidazoles that function as 5'-adenosinemonophosphate (AMP) mimics.

Efforts to enhance the inhibitory potency of the initial purine series of fructose-1,6-bisphosphatase (FBPase) inhibitors led to the discovery of a series of benzimidazole analogues with human FBPase IC(50)s < 100 nM. Inhibitor 4.4 emerged as a lead compound based on its potent inhibition of human liver FBPase (IC(50) = 55 nM) and significant glucose lowering in normal fasted rats. Intravenous administration of 4.4 to Zucker diabetic fatty rats led to rapid and robust glucose lowering, thereby providing the first evidence that FBPase inhibitors could improve glycemia in animal models of type 2 diabetes.

Discovery of phosphonic diamide prodrugs and their use for the oral delivery of a series of fructose 1,6-bisphosphatase inhibitors.

Like most phosphonic acids, the recently discovered potent and selective thiazole phosphonic acid inhibitors of fructose 1,6-bisphosphatase (FBPase) exhibited low oral bioavailability (OBAV) and therefore required a prodrug to achieve oral efficacy. Syntheses of known phosphonate prodrugs did not afford the desired OBAV; hence, a new class of prodrugs was sought. Phosphonic diamides derived from amino acid esters were discovered as viable prodrugs, which met our preset goals: excellent aqueous stability over a wide pH range, benign byproducts (amino acids and low molecular weight alcohols), and most importantly good OBAV leading to robust oral glucose lowering effects. These desirable properties of phosphonic diamides represent significant improvements over existing prodrug classes. Optimization of the diamide prodrugs of phosphonic acid 2a (MB05032) led to the identification of diamide 8 (MB06322), the first reported orally efficacious FBPase inhibitor.

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

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.