Decreasing the Rate of Metabolic Ketone Reduction in the Discovery of a Clinical Acetyl-CoA Carboxylase Inhibitor for the Treatment of Diabetes

Acetyl-CoA carboxylase (ACC) inhibitors offer significant potential for the treatment of type 2 diabetes mellitus (T2DM), hepatic steatosis, and cancer. However, the identification of tool compounds suitable to test the hypothesis in human trials has been challenging. An advanced series of spirocyclic ketone-containing ACC inhibitors recently reported by Pfizer were metabolized in vivo by ketone reduction, which complicated human pharmacology projections. We disclose that this metabolic reduction can be greatly attenuated through introduction of steric hindrance adjacent to the ketone carbonyl. Incorporation of weakly basic functionality improved solubility and led to the identification of <b>9</b> as a clinical candidate for the treatment of T2DM. Phase I clinical studies demonstrated dose-proportional increases in exposure, single-dose inhibition of de novo lipogenesis (DNL), and changes in indirect calorimetry consistent with increased whole-body fatty acid oxidation. This demonstration of target engagement validates the use of compound <b>9</b> to evaluate the role of DNL in human disease

The Hepatoselective Glucokinase Activator PF-04991532 Ameliorates Hyperglycemia without Causing Hepatic Steatosis in Diabetic Rats

<div><p>Hyperglycemia resulting from type 2 diabetes mellitus (T2DM) is the main cause of diabetic complications such as retinopathy and neuropathy. A reduction in hyperglycemia has been shown to prevent these associated complications supporting the importance of glucose control. Glucokinase converts glucose to glucose-6-phosphate and determines glucose flux into the β-cells and hepatocytes. Since activation of glucokinase in β-cells is associated with increased risk of hypoglycemia, we hypothesized that selectively activating hepatic glucokinase would reduce fasting and postprandial glucose with minimal risk of hypoglycemia. Previous studies have shown that hepatic glucokinase overexpression is able to restore glucose homeostasis in diabetic models; however, these overexpression experiments have also revealed that excessive increases in hepatic glucokinase activity may also cause hepatosteatosis. Herein we sought to evaluate whether liver specific pharmacological activation of hepatic glucokinase is an effective strategy to reduce hyperglycemia without causing adverse hepatic lipids changes. To test this hypothesis, we evaluated a hepatoselective glucokinase activator, PF-04991532, in Goto-Kakizaki rats. In these studies, PF-04991532 reduced plasma glucose concentrations independent of changes in insulin concentrations in a dose-dependent manner both acutely and after 28 days of sub-chronic treatment. During a hyperglycemic clamp in Goto-Kakizaki rats, the glucose infusion rate was increased approximately 5-fold with PF-04991532. This increase in glucose infusion can be partially attributed to the 60% reduction in endogenous glucose production. While PF-04991532 induced dose-dependent increases in plasma triglyceride concentrations it had no effect on hepatic triglyceride concentrations in Goto-Kakizaki rats. Interestingly, PF-04991532 decreased intracellular AMP concentrations and increased hepatic futile cycling. These data suggest that hepatoselective glucokinase activation may offer glycemic control without inducing hepatic steatosis supporting the evaluation of tissue specific activators in clinical trials.</p></div

PF-04991532 improves glucose metabolism in rats.

<p>PF-04991532 increased the rate of glucose infusion in order to maintain hyperglycemia in Goto-Kakizaki rats (n = 6/group) (<b>A</b>) which can be attributed to the increased glucose disposal and decreased glucose production (n = 6/group) during steady state (<b>B</b>). PF-04991532 decreased plasma glucose in Goto-Kakizaki rats over 28 days of dosing (n = 6–8/group) [P<0.05 for 30, 60, and 100 mg/kg compared to vehicle] (<b>C</b>) which was accompanied by an increase in plasma triglycerides at the highest dose (n = 6–8/group) [P<0.05 for 100 mg/kg] (<b>D</b>). These plasma changes were not associated with any changes in liver triglycerides compared to vehicle treated animals (<b>E</b>). **P<0.01 One-way ANOVA and Tukey’s Multiple Comparison Test were used for A,C,D, & E. Student’s t-test was used for B.</p

PF-04991532 effects on lipid metabolism and downstream hepatic metabolites.

<p>Expression of key lipid, carbohydrate, and cholesterol metabolism genes in rats treated with an acute 100/kg dose of PF-04991532 relative to vehicle (<b>A</b>). The increase in expression of the lipogenic genes can be explained by the increased nuclear ChREBP (n = 3/4/group) (<b>B</b>). Lastly, a metabolic profile of key metabolites using ³¹P-NMR and ¹H-NMR in rats treated with an acute dose of PF-04991532 (n = 5–6/group) (<b>C</b>). *P<0.05; #P<0.1 for Vehicle vs. 100 mg/kg PF-04991532. Student’s t-test was used.</p

PF-04991532 increased hepatic futile cycling.

<p>The effect of hepatic glucokinase activation on G6Pase (<b>A</b>) and the loss of positional labeling (<b>B</b>) in primary rat hepatocytes. PF-04991532 increased total hepatic substrate cycling due to increased substrate cycling between glucose and glucose-6-phosphate (n = 5–7/group) (<b>C</b>). The increased substrate cycling decreased hepatic ATP concentrations as assessed by NMR (n = 6/group) (<b>D</b>) which in turn increased hepatic pAMPK/AMPK ratio (<b>E</b>) (n = 5/group). *P<0.05, **P<0.01, ***P<0.005 for Vehicle vs. 100 mg/kg PF-04991532. Student’s t-test was used.</p