Odile Esther Levy

Novel Exenatide Analogs with Peptidic Albumin Binding Domains: Potent Anti-Diabetic Agents with Extended Duration of Action

The design, synthesis and pharmacology of novel long-acting exenatide analogs for the treatment of metabolic diseases are described. These molecules display enhanced pharmacokinetic profile and potent glucoregulatory and weight lowering actions compared to native exenatide. [Leu14]exenatide-ABD is an 88 residue peptide amide incorporating an Albumin Binding Domain (ABD) scaffold. [Leu14]exenatide-ABP is a 53 residue peptide incorporating a short Albumin Binding Peptide (ABP). [Leu14]exenatide-ABD and [Leu14]exenatide-ABP exhibited nanomolar functional GLP-1 receptor potency and were metabolically stable in vitro in human plasma and in a pancreatic digestive enzyme mixture. Both molecules displayed picomolar and nanomolar binding association with albumin across multiple species and circulating half lives of 16 and 11 hours, respectively, post a single IV dose in rats. Unlike exenatide, both molecules elicited robust glucose lowering when injected 1 day prior to an oral glucose tolerance test, indicative of their extended duration of action. [Leu14]exenatide-ABD was compared to exenatide in a Lep ob/ob mouse model of diabetes. Twice-weekly subcutaneously dosed [Leu14]exenatide-ABD displayed superior glucose lowering and weight loss in diabetic mice when compared to continuously infused exenatide at the same total weekly dose. A single oral administration of each molecule via an enteric coated capsule to cynomolgus monkeys showed superior pharmacokinetics for [Leu14]exenatide-ABD as compared to [Leu14]exenatide-ABP with detectable exposure longer than 14 days. These studies support the potential use of these novel long acting exenatide analogs with different routes of administration for the treatment of type 2 diabetes.

A novel long‐acting glucose‐dependent insulinotropic peptide analogue: enhanced efficacy in normal and diabetic rodents

Glucose‐dependent insulinotropic peptide (GIP) is an incretin hormone that is released from intestinal K cells in response to nutrient ingestion. We aimed to investigate the therapeutic potential of the novel N‐ and C‐terminally modified GIP analogue AC163794.

Improved Glucose Control and Reduced Body Weight in Rodents with Dual Mechanism of Action Peptide Hybrids

Combination therapy is being increasingly used as a treatment paradigm for metabolic diseases such as diabetes and obesity. In the peptide therapeutics realm, recent work has highlighted the therapeutic potential of chimeric peptides that act on two distinct receptors, thereby harnessing parallel complementary mechanisms to induce additive or synergistic benefit compared to monotherapy. Here, we extend this hypothesis by linking a known anti-diabetic peptide with an anti-obesity peptide into a novel peptide hybrid, which we termed a phybrid. We report on the synthesis and biological activity of two such phybrids (AC164204 and AC164209), comprised of a glucagon-like peptide-1 receptor (GLP1-R) agonist, and exenatide analog, AC3082, covalently linked to a second generation amylin analog, davalintide. Both molecules acted as full agonists at their cognate receptors in vitro, albeit with reduced potency at the calcitonin receptor indicating slightly perturbed amylin agonism. In obese diabetic Lepob/Lep ob mice sustained infusion of AC164204 and AC164209 reduced glucose and glycated haemoglobin (HbA1c) equivalently but induced greater weight loss relative to exenatide administration alone. Weight loss was similar to that induced by combined administration of exenatide and davalintide. In diet-induced obese rats, both phybrids dose-dependently reduced food intake and body weight to a greater extent than exenatide or davalintide alone, and equal to co-infusion of exenatide and davalintide. Phybrid-mediated and exenatide + davalintide-mediated weight loss was associated with reduced adiposity and preservation of lean mass. These data are the first to provide in vivo proof-of-concept for multi-pathway targeting in metabolic disease via a peptide hybrid, demonstrating that this approach is as effective as co-administration of individual peptides.

Synthesis & biological evaluation of PYY(3-36) analogs substituted with alanine.

Introduction PYY is a 36-residue peptide first isolated from porcine intestine (Figure 1) [1,2]. Two endogenous forms of PYY, PYY(1-36) and the post-DPPIV [3,4] activated PYY(3-36), are released into the circulation following a meal [1,5]. PYY(3-36) appears to be the predominant secreted form. PYY has been known to inhibit gastric, [6] pancreatic and intestinal secretions [7]. PYY binds and activates at least four receptor subtypes (Y1, Y2, Y4 and Y5) [8-10] in rats and humans. These Y receptor subtypes display different patterns of affinity and activation for PYY, PYY(3-36) and synthetically modified PYY analogs. PYY(3-36) is a selective ligand for Y2 and Y5 receptors, implicated in food intake and feeding behavior, respectively [11]. In this study, PYY(3-36) analogs, where each residue of the natural sequence is replaced by L-alanine, and analogs with multiple alanine substitutions were synthesized. The three alanines at positions 7, 12 and 22 were replaced by a D-alanine. The affinity of each analog to the Y family of receptors and the ability of the analogs to decrease acute food intake in mouse are presented. Additionally, the efficacy of a single equimolar dose of PYY(3-36) and a subset of the analogs to reduce body weight in the chronic weight loss assay in the mouse is summarized.

Optimization of a continuous assay for obtaining sensitive kinetic data on the inhibition of the HCV NS3 protease

Hepatitis C, which is caused by HCV, affects about 2% of the world’s population and is responsible for more than 50 million cases of hepatocellular carcinomas worldwide. HCV uses hepacivirin, a serine protease at the N-terminal third of the protein product of its NS3 gene, to process its NS4a-NS4b-NS5a-NS5b polyprotein. In addition, hepacivirin cleaves internally the NS3-NS4a site, releasing its cofactor NS4a. Inhibition of hepacivirin’s activity is one strategy for developing drugs to treat hepatitis C. Methods to measure the activity and inhibition of hepacivirin have included the use of polyproteins, peptides, and depsipeptides as substrates in PAGE, ELISA, BIAcore, and RP-HPLC-based assays [1-5]. More preferable are continuous chromogenic and fluorogenic assays, which use commercially available peptide-pNAs and peptide-AMCs, respectively. Additionally, an assay using peptide-p-phenylazophenyl ester as substrate has been developed [6], which is limiting due to a requirement for low pH and inherent high background and stability problems. Most recently, a continuous assay based on fluorescence resonance energy transfer (FRET) depsipeptide substrates has been presented [7]. Using the same FRET principle, we designed depsipeptide substrates for greater ease of chemical synthesis, lower molecular weight, and enhancing assay sensitivity with lower background, while maintaining the advantages of compatibility and stability at physiologic pH.