Bruce A. Ellsworth

Discovery of Dapagliflozin: A Potent, Selective Renal Sodium-Dependent Glucose Cotransporter 2 (SGLT2) Inhibitor for the Treatment of Type 2 Diabetes

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.

In vitro characterization and pharmacokinetics of dapagliflozin (BMS-512148), a potent sodium-glucose cotransporter type II inhibitor, in animals and humans.

(2S,3R,4R,5S,6R)-2-(3-(4-Ethoxybenzyl)-4-chlorophenyl)-6-hydroxymethyl-tetrahydro-2H-pyran-3,4,5-triol (dapagliflozin; BMS-512148) is a potent sodium-glucose cotransporter type II inhibitor in animals and humans and is currently under development for the treatment of type 2 diabetes. The preclinical characterization of dapagliflozin, to allow compound selection and prediction of pharmacological and dispositional behavior in the clinic, involved Caco-2 cell permeability studies, cytochrome P450 (P450) inhibition and induction studies, P450 reaction phenotyping, metabolite identification in hepatocytes, and pharmacokinetics in rats, dogs, and monkeys. Dapagliflozin was found to have good permeability across Caco-2 cell membranes. It was found to be a substrate for P-glycoprotein (P-gp) but not a significant P-gp inhibitor. Dapagliflozin was not found to be an inhibitor or an inducer of human P450 enzymes. The in vitro metabolic profiles of dapagliflozin after incubation with hepatocytes from mice, rats, dogs, monkeys, and humans were qualitatively similar. Rat hepatocyte incubations showed the highest turnover, and dapagliflozin was most stable in human hepatocytes. Prominent in vitro metabolic pathways observed were glucuronidation, hydroxylation, and O-deethylation. Pharmacokinetic parameters for dapagliflozin in preclinical species revealed a compound with adequate oral exposure, clearance, and elimination half-life, consistent with the potential for single daily dosing in humans. The pharmacokinetics in humans after a single dose of 50 mg of [14C]dapagliflozin showed good exposure, low clearance, adequate half-life, and no metabolites with significant pharmacological activity or toxicological concern.

Reductions in log P improved protein binding and clearance predictions enabling the prospective design of cannabinoid receptor (CB1) antagonists with desired pharmacokinetic properties.

Several strategies have been employed to reduce the long in vivo half-life of our lead CB1 antagonist, triazolopyridazinone 3, to differentiate the pharmacokinetic profile versus the lead clinical compounds. An in vitro and in vivo clearance data set revealed a lack of correlation; however, when compounds with <5% free fraction were excluded, a more predictable correlation was observed. Compounds with log P between 3 and 4 were likely to have significant free fraction, so we designed compounds in this range to give more predictable clearance values. This strategy produced compounds with desirable in vivo half-lives, ultimately leading to the discovery of compound 46. The progression of compound 46 was halted due to the contemporaneous marketing and clinical withdrawal of other centrally acting CB1 antagonists; however, the design strategy successfully delivered a potent CB1 antagonist with the desired pharmacokinetic properties and a clean off-target profile.

Chapter Twenty-Eight – To Market, To Market—2012

Abstract This year’s To-Market, To-Market chapter provides summaries for 30 compounds that received first-time approval worldwide in 2012. Anticancer treatments dominated the new entries with 14 approvals, including 12 small molecules and 2 monoclonal antibodies. Three new diabetes treatments were introduced, along with three agents for gastrointestinal disorders. The remaining therapeutic areas had 1–2 approvals each. The summaries include indication, information about the disease treated, mechanism of action, selected preclinical data, key steps in the synthesis, pharmacokinetic and metabolism profile, clinical efficacy and safety data, and other key information about the approval.

Cannabinoid CB1 receptor ligand binding and function examined through mutagenesis studies of F200 and S383

The cannabinoid CB(1) G protein-coupled receptor has been shown to be a regulator of food consumption and has been studied extensively as a drug target for the treatment of obesity. To advance understanding of the receptors three-dimensional structure, we performed mutagenesis studies at human cannabinoid CB(1) receptor residues F200 and S383 and measured changes in activity and binding affinity of compounds from two recently discovered active chemotypes, arylsulfonamide agonists and tetrahydroquinoline-based inverse agonists, as well as literature compounds. Our results add support to previous findings that both agonists and inverse agonists show varied patterns of binding at the two mutated residue sites, suggesting multiple subsites for binding to the cannabinoid CB(1) receptor for both functional types of ligands. We additionally find that an F200L mutation in the receptor largely restores binding affinity to ligands and significantly decreases constitutive activity when compared to F200A, resulting in a receptor phenotype that is closer to the wild-type receptor. The results downplay the importance of aromatic stacking interactions at F200 and suggest that a bulky hydrophobic contact is largely sufficient to provide significant receptor function and binding affinity to cannabinoid CB(1) receptor ligands.

Discovery of Pyrrolidine-Containing GPR40 Agonists: Stereochemistry Effects a Change in Binding Mode

A novel series of pyrrolidine-containing GPR40 agonists is described as a potential treatment for type 2 diabetes. The initial pyrrolidine hit was modified by moving the position of the carboxylic acid, a key pharmacophore for GPR40. Addition of a 4-cis-CF3 to the pyrrolidine improves the human GPR40 binding Ki and agonist efficacy. After further optimization, the discovery of a minor enantiomeric impurity with agonist activity led to the finding that enantiomers (R,R)-68 and (S,S)-68 have differential effects on the radioligand used for the binding assay, with (R,R)-68 potentiating the radioligand and (S,S)-68 displacing the radioligand. Compound (R,R)-68 activates both Gq-coupled intracellular Ca2+ flux and Gs-coupled cAMP accumulation. This signaling bias results in a dual mechanism of action for compound (R,R)-68, demonstrating glucose-dependent insulin and GLP-1 secretion in vitro. In vivo, compound (R,R)-68 significantly lowers plasma glucose levels in mice during an oral glucose challenge, encouraging further development of the series.

Chapter Twenty-Seven – To Market, To Market—2013

Abstract This years To Market, To Market chapter provides summaries for 25 small molecules and two monoclonal antibodies that received approval for the first time in any country in 2013. The summaries include indication, information about the disease treated, mechanism of action, selected preclinical data, key steps in the synthesis, pharmacokinetic and metabolism profile, clinical efficacy and safety data, and other key information about the approval.

Discovery of Potent and Orally Bioavailable Dihydropyrazole GPR40 Agonists

G protein-coupled receptor 40 (GPR40) has become an attractive target for the treatment of diabetes since it was shown clinically to promote glucose-stimulated insulin secretion. Herein, we report our efforts to develop highly selective and potent GPR40 agonists with a dual mechanism of action, promoting both glucose-dependent insulin and incretin secretion. Employing strategies to increase polarity and the ratio of sp3/sp2 character of the chemotype, we identified BMS-986118 (compound 4), which showed potent and selective GPR40 agonist activity in vitro. In vivo, compound 4 demonstrated insulinotropic efficacy and GLP-1 secretory effects resulting in improved glucose control in acute animal models.

Discovery and Development of Selective Renal Sodium-Dependent Glucose Cotransporter 2 (SGLT2) Dapagliflozin for the Treatment of Type 2 Diabetes

When blood flows through the renal capillaries, glucose is one of the many substances filtered by the kidney. However, glucose is subsequently recovered primarily by the sodium-dependent glucose transporter 2 (SGLT2) as the glomerular filtrate flows down the renal tubules. SGLT2 inhibitors inhibit this transporter leading to the loss of a significant fraction of the filtered glucose. The resulting glucosuria is of sufficient magnitude to reduce diabetes-related hyperglycemia and ameliorate-associated complications of diabetes. A systematic study was conducted to identify superior SGLT2 inhibitors based on a β- 1C-arylglucoside with substituted diarylmethane moieties. Such compounds are potent and selective SGLT2 inhibitors with metabolic stability that promote glucosuria when administered in vivo. Through this investigation, the β- 1C-arylglucoside dapagliflozin was identified as a potent and selective hSGLT2 inhibitor with an EC50 for hSGLT2 of 1.0 nM and 1,200-fold selectivity over hSGLT1. Dapagliflozin produced glucosuria in normal Sprague Dawley rats in a dose-dependent fashion. Moreover, a 0.1 mg/kg oral dose reduced blood glucose levels by as much as 55% in rats that had been made hyperglycemic by streptozotocin, a pancreatic toxin. These findings, combined with a favorable ADME profile and vivo data, led to nomination of dapagliflozin as a drug for the treatment of type 2 diabetes. The structural architecture of β- 1C-arylglucosides and their amphiphilic nature presented significant obstacles to the synthesis of dapagliflozin and similar candidates for toxicological and clinical testing, prompting the development of a new, safe, efficient, and economical process for the synthesis of C-4′ and C-4 substituted β- 1C-arylglucosides. A key element of the process was a remarkable discovery of novel crystalline complexes that enabled isolation and quality control.