David R. Derksen

Short Hydrophobic Peptides with Cyclic Constraints Are Potent Glucagon-like Peptide-1 Receptor (GLP-1R) Agonists.

Cyclic constraints are incorporated into an 11-residue analogue of the N-terminus of glucagon-like peptide-1 (GLP-1) to investigate effects of structure on agonist activity. Cyclization through linking side chains of residues 2 and 5 or 5 and 9 produced agonists at nM concentrations in a cAMP assay. 2D NMR and CD spectra revealed an N-terminal β-turn and a C-terminal helix that differentially influenced affinity and agonist potency. These structures can inform development of small molecule agonists of the GLP-1 receptor to treat type 2 diabetes.

Discovery of (S)-6-(3-Cyclopentyl-2-(4-(trifluoromethyl)-1H-imidazol-1-yl)propanamido)nicotinic Acid as a Hepatoselective Glucokinase Activator Clinical Candidate for Treating Type 2 Diabetes Mellitus

Glucokinase is a key regulator of glucose homeostasis, and small molecule allosteric activators of this enzyme represent a promising opportunity for the treatment of type 2 diabetes. Systemically acting glucokinase activators (liver and pancreas) have been reported to be efficacious but in many cases present hypoglycaemia risk due to activation of the enzyme at low glucose levels in the pancreas, leading to inappropriately excessive insulin secretion. It was therefore postulated that a liver selective activator may offer effective glycemic control with reduced hypoglycemia risk. Herein, we report structure-activity studies on a carboxylic acid containing series of glucokinase activators with preferential activity in hepatocytes versus pancreatic β-cells. These activators were designed to have low passive permeability thereby minimizing distribution into extrahepatic tissues; concurrently, they were also optimized as substrates for active liver uptake via members of the organic anion transporting polypeptide (OATP) family. These studies lead to the identification of 19 as a potent glucokinase activator with a greater than 50-fold liver-to-pancreas ratio of tissue distribution in rodent and non-rodent species. In preclinical diabetic animals, 19 was found to robustly lower fasting and postprandial glucose with no hypoglycemia, leading to its selection as a clinical development candidate for treating type 2 diabetes.

Designing glucokinase activators with reduced hypoglycemia risk: discovery of N,N-dimethyl-5-(2-methyl-6-((5-methylpyrazin-2-yl)-carbamoyl)benzofuran-4-yloxy)pyrimidine-2-carboxamide as a clinical candidate for the treatment of type 2 diabetes mellitus

Glucokinase is a key regulator of glucose homeostasis and small molecule activators of this enzyme represent a promising opportunity for the treatment of Type 2 diabetes. Several glucokinase activators have advanced to clinical studies and demonstrated promising efficacy; however, many of these early candidates also revealed hypoglycemia as a key risk. In an effort to mitigate this hypoglycemia risk while maintaining the promising efficacy of this mechanism, we have investigated a series of substituted 2-methylbenzofurans as “partial activators” of the glucokinase enzyme leading to the identification of N,N-dimethyl-5-(2-methyl-6-((5-methylpyrazin-2-yl)-carbamoyl)benzofuran-4-yloxy)pyrimidine-2-carboxamide as an early development candidate.

The design and synthesis of indazole and pyrazolopyridine based glucokinase activators for the treatment of type 2 diabetes mellitus.

Glucokinase activators represent a promising potential treatment for patients with Type 2 diabetes. Herein, we report the identification and optimization of a series of novel indazole and pyrazolopyridine based activators leading to the identification of 4-(6-(azetidine-1-carbonyl)-5-fluoropyridin-3-yloxy)-2-ethyl-N-(5-methylpyrazin-2-yl)-2H-indazole-6-carboxamide (42) as a potent activator with favorable preclinical pharmacokinetic properties and in vivo efficacy.

Pyrimidone-based series of glucokinase activators with alternative donor-acceptor motif.

Glucokinase activators are a class of experimental agents under investigation as a therapy for Type 2 diabetes mellitus. An X-ray crystal structure of a modestly potent agent revealed the potential to substitute the common heterocyclic amide donor-acceptor motif for a pyridone moiety. We have successfully demonstrated that both pyridone and pyrimidone heterocycles can be used as a potent donor-acceptor substituent. Several sub-micromolar analogs that possess the desired partial activator profile were synthesized and characterized. Unfortunately, the most potent activators suffered from sub-optimal pharmacokinetic properties. Nonetheless, these donor-acceptor motifs may find utility in other glucokinase activator series or beyond.

Discovery of an intravenous hepatoselective glucokinase activator for the treatment of inpatient hyperglycemia.

Glucokinase (hexokinase IV) continues to be a compelling target for the treatment of type 2 diabetes given the wealth of supporting human genetics data and numerous reports of robust clinical glucose lowering in patients treated with small molecule allosteric activators. Recent work has demonstrated the ability of hepatoselective activators to deliver glucose lowering efficacy with minimal risk of hypoglycemia. While orally administered agents require a considerable degree of passive permeability to promote suitable exposures, there is no such restriction on intravenously delivered drugs. Therefore, minimization of membrane diffusion in the context of an intravenously agent should ensure optimal hepatic targeting and therapeutic index. This work details the identification a hepatoselective GKA exhibiting the aforementioned properties.

Optimizing glucokinase activator binding kinetics to lower in vivo hypoglycemia risk

Activation of glucokinase represents a promising strategy for the treatment of type 2 diabetes; however, drug candidates have failed in clinical trials due to narrow therapeutic index between glucose-lowering efficacy and hypoglycemia. Described herein is a novel strategy for the design of next generation glucokinase activators with increased therapeutic index, which involves the optimization of activator-enzyme binding kinetics (kon, koff). This approach is based on the idea that activator binding kinetics are relevant to pharmacodynamics since the affinity of activator binding to glucokinase is cooperative with glucose such that, the rate at which an activator dissociates may influence the enzymes sensitivity to changes in physiological glucose concentrations. This study provides a compelling example of using fast-off binding kinetics for developing safe and effective activator drugs targeting glucokinase.

Cyclic alpha-conotoxin peptidomimetic chimeras as potent GLP-1R agonists

Type 2 diabetes mellitus (T2DM) results from compromised pancreatic β-cell function, reduced insulin production, and lowered insulin sensitivity in target organs resulting in hyperglycemia. The GLP-1 hormone has two biologically active forms, GLP-1-(7-37) and GLP-1-(7-36)amide, which are equipotent at the glucagon-like peptide-1 receptor (GLP-1R). These peptides are central both to normal glucose metabolism and dysregulation in T2DM. Several structurally modified GLP-1 analogues are now approved drugs, and a number of other analogues are in clinical trials. None of these compounds is orally bioavailable and all require parenteral delivery. Recently, a number of smaller peptidomimetics containing 11-12 natural and unnatural amino acids have been identified that have similar insulin regulating profiles as GLP-1. The α-conotoxins are a class of disulfide rich peptide venoms isolated from cone snails, and are known for their highly constrained structures and resistance to enzymatic degradation. In this study, we examined whether 11-residue peptidomimetics incorporated into α-conotoxin scaffolds, forming monocyclic or bicyclic compounds constrained by disulfide bonds and/or backbone cyclization, could activate the GLP-1 receptor (GLP-1R). Several compounds showed potent (nanomolar) agonist activity at GLP-1R, as evaluated via cAMP signaling. In addition, HPLC retention times and in silico calculations suggested that mono- and bicyclic compounds had more favorable n-octanol/water partition coefficients according to the virtual partition coefficient model (vLogP), while maintaining a smaller radius of gyration compared to corresponding uncyclized peptidomimetics. Our findings suggest that cyclic peptidomimetics provide a potential avenue for future design of potent, compact ligands targeting GLP-1R and possessing improved physicochemical properties.

Truncated Glucagon-like Peptide-1 and Exendin-4 α-Conotoxin pl14a Peptide Chimeras Maintain Potency and α-Helicity and Reveal Interactions Vital for cAMP Signaling in Vitro.

Glucagon-like peptide-1 (GLP-1) signaling through the glucagon-like peptide 1 receptor (GLP-1R) is a key regulator of normal glucose metabolism, and exogenous GLP-1R agonist therapy is a promising avenue for the treatment of type 2 diabetes mellitus. To date, the development of therapeutic GLP-1R agonists has focused on producing drugs with an extended serum half-life. This has been achieved by engineering synthetic analogs of GLP-1 or the more stable exogenous GLP-1R agonist exendin-4 (Ex-4). These synthetic peptide hormones share the overall structure of GLP-1 and Ex-4, with a C-terminal helical segment and a flexible N-terminal tail. Although numerous studies have investigated the molecular determinants underpinning GLP-1 and Ex-4 binding and signaling through the GLP-1R, these have primarily focused on the length and composition of the N-terminal tail or on how to modulate the helicity of the full-length peptides. Here, we investigate the effect of C-terminal truncation in GLP-1 and Ex-4 on the cAMP pathway. To ensure helical C-terminal regions in the truncated peptides, we produced a series of chimeric peptides combining the N-terminal portion of GLP-1 or Ex-4 and the C-terminal segment of the helix-promoting peptide α-conotoxin pl14a. The helicity and structures of the chimeric peptides were confirmed using circular dichroism and NMR, respectively. We found no direct correlation between the fractional helicity and potency in signaling via the cAMP pathway. Rather, the most important feature for efficient receptor binding and signaling was the C-terminal helical segment (residues 22–27) directing the binding of Phe22 into a hydrophobic pocket on the GLP-1R.

Helixconstraints and amino acid substitution in GLP-1 increase cAMP and insulin secretion but not beta-arrestin 2 signaling

Glucagon-like peptide (GLP-1) is an endogenous hormone that induces insulin secretion from pancreatic islets and modified forms are used to treat diabetes mellitus type 2. Understanding how GLP-1 interacts with its receptor (GLP-1R) can potentially lead to more effective drugs. Modeling and NMR studies of the N-terminus of GLP-1 suggest a β-turn between residues Glu9-Phe12 and a kinked alpha helix between Val16-Gly37. N-terminal turn constraints attenuated binding affinity and activity (compounds 1-8). Lys-Asp (i, i+4) crosslinks in the middle and at the C-terminus increased alpha helicity and cAMP stimulation without much effect on binding affinity or beta-arrestin 2 recruitment (compounds 9-18). Strategic positioning of helix-inducing constraints and amino acid substitutions (Tyr16, Ala22) increased peptide helicity and produced ten-fold higher cAMP potency (compounds 19-28) over GLP-1(7-37)-NH2. The most potent cAMP activator (compound 23) was also the most potent inducer of insulin secretion.