Dual Glucagon-like Peptide 1 (GLP-1)/Glucagon Receptor Agonists Specifically Optimized for Multidose Formulations.

Novel peptidic dual agonists of the glucagon-like peptide 1 (GLP-1) and glucagon receptor are reported to have enhanced efficacy over pure GLP-1 receptor agonists with regard to treatment of obesity and diabetes. We describe novel exendin-4 based dual agonists designed with an activity ratio favoring the GLP-1 versus the glucagon receptor. As result of an iterative optimization procedure that included molecular modeling, structural biological studies (X-ray, NMR), peptide design and synthesis, experimental activity, and solubility profiling, a candidate molecule was identified. Novel SAR points are reported that allowed us to fine-tune the desired receptor activity ratio and increased solubility in the presence of antimicrobial preservatives, findings that can be of general applicability for any peptide discovery project. The peptide was evaluated in chronic in vivo studies in obese diabetic monkeys as translational model for the human situation and demonstrated favorable blood glucose and body weight lowering effects.

Design of Novel Exendin-Based Dual Glucagon-like Peptide 1 (GLP-1)/Glucagon Receptor Agonists.

Dual activation of the glucagon-like peptide 1 (GLP-1) and glucagon receptor has the potential to lead to a novel therapy principle for the treatment of diabesity. Here, we report a series of novel peptides with dual activity on these receptors that were discovered by rational design. On the basis of sequence analysis and structure-based design, structural elements of glucagon were engineered into the selective GLP-1 receptor agonist exendin-4, resulting in hybrid peptides with potent dual GLP-1/glucagon receptor activity. Detailed structure-activity relationship data are shown. Further modifications with unnatural and modified amino acids resulted in novel metabolically stable peptides that demonstrated a significant dose-dependent decrease in blood glucose in chronic studies in diabetic db/db mice and reduced body weight in diet-induced obese (DIO) mice. Structural analysis by NMR spectroscopy confirmed that the peptides maintain an exendin-4-like structure with its characteristic tryptophan-cage fold motif that is responsible for favorable chemical and physical stability.

Discovery of a potent, selective, and orally bioavailable histamine H3 receptor antagonist SAR110068 for the treatment of sleep-wake disorders.

Previous studies have shown that compound 1 displayed high affinity towards histamine H3 receptor (H3R), (human (h-H3R), K(i)=8.6 nM, rhesus monkey (rh-H3R), K(i)=1.2 nM, and rat (r-H3R), K(i)=16.5 nM), but exhibited high affinity for hERG channel. Herein, we report the discovery of a novel, potent, and highly selective H3R antagonist/inverse agonist 5a(SS) (SAR110068) with acceptable hERG channel selectivity and desirable pharmacological and pharmacokinetic properties through lead optimization sequence. The significant awakening effects of 5a(SS) on sleep-wake cycles studied by using EEG recording in rats during their light phase support its potential therapeutic utility in human sleep-wake disorders.

Discovery of aryl ureas and aryl amides as potent and selective histamine H3 receptor antagonists for the treatment of obesity (part I).

A series of structurally novel aryl ureas was derived from optimization of the HTS lead as selective histamine H3 receptor (H3R) antagonists. The SAR was explored and the data obtained set up the starting point and foundation for further optimization. The most potent tool compounds, as exemplified by compounds 2l, 5b, 5d, and 5e, displayed antagonism potencies in the subnanomolar range in in vitro human-H3R FLIPR assays and rhesus monkey H3R binding assays.

Discovery of aryl ureas and aryl amides as potent and selective histamine H3 receptor antagonists for the treatment of obesity (part II).

A novel series of histamine H3 receptor (H3R) antagonists was derived from an arylurea lead series (1) via bioisosteric replacement of the urea functionality by an amide linkage. The arylamide series was optimized through SAR studies by a broad variation of substituents in the left-hand side benzoyl residue (analogs 2a-2ag) or replacement of the benzoyl moiety by heteroarylcarbonyl residues (analogs 5a-5n). Compounds 2p and 2q were identified within the series as potent and selective H3R antagonists/inverse agonists with acceptable overall profile. Compound 2q was orally active in food intake inhibition in diet-induced obese (DIO) mice. Compound 2q represents a novel H3R antagonist template with improved in vitro potency and oral efficacy and has its merits as a new lead for further optimization.

CB1 receptor antagonist AVE1625 affects primarily metabolic parameters independently of reduced food intake in Wistar rats.

The objective of the present study was to investigate in fed Wistar rats whether the cannabinoid-1 (CB1) receptor antagonist AVE1625 causes primary effects on metabolic blood and tissue parameters as well as metabolic rate, which are independent of reduced caloric intake. After single administration to rats postprandially, AVE1625 caused a slight dose-dependent increase in basal lipolysis. Six hours after single administration, liver glycogen content was dose-dependently reduced to approximately 60% of that of untreated controls. These findings demonstrate a primary acute effect of AVE1625 on induction of 1) lipolysis from fat tissue (increased FFA) and 2) glycogenolysis from the liver (reduced hepatic glycogen). Measured by indirect calorimetry, AVE1625 caused an immediate increase in total energy expenditure, a long-lasting increase of fat oxidation, and a transient increase of glucose oxidation, which were consistent with the acute findings on metabolic blood and tissue parameters. We conclude that, in addition to the well-investigated effects of CB1 receptor antagonists to reduce caloric intake and subsequently body weight, this pharmacological approach is additionally linked to inherently increased lipid oxidation. This oxidation is driven by persistently increased lipolysis from fat tissues, independently of reduced caloric intake, and might significantly contribute to the weight-reducing effect.

Insights into the bile acid transportation system: the human ileal lipid-binding protein-cholyltaurine complex and its comparison with homologous structures.

Bile acids are generated in vivo from cholesterol in the liver, and they undergo an enterohepatic circulation involving the small intestine, liver, and kidney. To understand the molecular mechanism of this transportation, it is essential to gain insight into the three-dimensional (3D) structures of proteins involved in the bile acid recycling in free and complexed form and to compare them with homologous members of this protein family. Here we report the solution structure of the human ileal lipid-binding protein (ILBP) in free form and in complex with cholyltaurine. Both structures are compared with a previously published structure of the porcine ILBP-cholylglycine complex and with related lipid-binding proteins. Protein structures were determined in solution by using two-dimensional (2D)- and 3D-homo and heteronuclear NMR techniques, leading to an almost complete resonance assignment and a significant number of distance constraints for distance geometry and restrained molecular dynamics simulations. The identification of several intermolecular distance constraints unambiguously determines the cholyltaurine-binding site. The bile acid is deeply buried within ILBP with its flexible side-chain situated close to the fatty acid portal as entry region into the inner ILBP core. This binding mode differs significantly from the orientation of cholylglycine in porcine ILBP. A detailed analysis using the GRID/CPCA strategy reveals differences in favorable interactions between protein-binding sites and potential ligands. This characterization will allow for the rational design of potential inhibitors for this relevant system.

Cyclosporin a and enterohepatic circulation of bile salts in rats: decreased cholate synthesis but increased intestinal reabsorption.

Cyclosporin A (CsA) has been shown to inhibit synthesis and hepatobiliary transport of bile salts. However, effects of CsA on the enterohepatic circulation of bile salts in vivo are largely unknown. We characterized the effects of CsA on the enterohepatic circulation of cholate, with respect to synthesis rate, pool size, cycling time, intestinal absorption, and the expression of relevant transporters in liver and intestine in rats. CsA (1 mg. 100 g(-1). day(-1) s.c.) or its solvent was administered daily to male rats for 10 days. Cholate synthesis rate and pool size were determined by a 2H4-cholate dilution technique. Bile and feces were collected for determination of cholate and total bile salts, respectively. Cycling time and intestinal absorption of cholate were calculated. The mRNA levels and corresponding transporter protein levels in liver and intestine were assessed by real-time polymerase chain reaction and Western analysis, respectively. CsA treatment decreased cholate synthesis rate by 71%, but did not affect pool size or cycling time. CsA reduced the amount of cholate lost per enterohepatic cycle by approximately 70%. Protein levels of the apical sodium-dependent bile salt transporter (Asbt) were 2-fold increased in distal ileum of CsA-treated rats, due to post-transcriptional events. In conclusion, chronic CsA treatment markedly reduces cholate synthesis rate in rats, but does not affect cholate pool size or cycling time. Our results strongly suggest that CsA enhances efficacy of intestinal cholate reabsorption through increased Asbt protein expression in the distal ileum, which contributes to maintenance of cholate pool size in CsA-treated rats.