PGF2α signaling drives fibrotic remodeling and fibroblast population dynamics in mice.

Idiopathic pulmonary fibrosis (IPF) is a chronic parenchymal lung disease characterized by repetitive alveolar cell injury, myofibroblast proliferation, and excessive extracellular matrix deposition for which unmet need persists for effective therapeutics. The bioactive eicosanoid, prostaglandin F2α, and its cognate receptor FPr (Ptgfr) are implicated as a TGF-β1-independent signaling hub for IPF. To assess this, we leveraged our published murine PF model (IER-SftpcI73T) expressing a disease-associated missense mutation in the surfactant protein C (Sftpc) gene.

Disruption of Prostaglandin F Receptor Signaling Attenuates Fibrotic Remodeling and Alters Fibroblast Population Dynamics in A Preclinical Murine Model of Idiopathic Pulmonary Fibrosis.

Idiopathic Pulmonary Fibrosis (IPF) is a chronic parenchymal lung disease characterized by repetitive alveolar cell injury, myofibroblast proliferation, and excessive extracellular matrix deposition for which unmet need persists for effective therapeutics. The bioactive eicosanoid, prostaglandin F2, and its cognate receptor FPr () are implicated as a TGF1 independent signaling hub for IPF. To assess this, we leveraged our published murine PF model (I - ) expressing a disease-associated missense mutation in the surfactant protein C () gene.

Discovery of a new class of integrin antibodies for fibrosis.

Lung fibrosis, or the scarring of the lung, is a devastating disease with huge unmet medical need. There are limited treatment options and its prognosis is worse than most types of cancer. We previously discovered that MK-0429 is an equipotent pan-inhibitor of αv integrins that reduces proteinuria and kidney fibrosis in a preclinical model.

Molecular Profiling Reveals a Common Metabolic Signature of Tissue Fibrosis.

Fibrosis, or the accumulation of extracellular matrix, is a common feature of many chronic diseases. To interrogate core molecular pathways underlying fibrosis, we cross-examine human primary cells from various tissues treated with TGF-β, as well as kidney and liver fibrosis models. Transcriptome analyses reveal that genes involved in fatty acid oxidation are significantly perturbed.

Differential anti-thrombotic benefit and bleeding risk profiles of antagonists of protease-activated receptor 1 and 4 in Cynomolgus Macaques.

Platelet activation plays a crucial role in hemostasis and thrombosis. Thrombin, the most potent stimulus of platelet activation, mediates platelet activation via the protease activated receptors (PARs). The platelet PAR repertoire in mediating thrombin's action differs across species.

Discovery of Triazole CYP11B2 Inhibitors with in Vivo Activity in Rhesus Monkeys.

Hit-to-lead efforts resulted in the discovery of compound 19, a potent CYP11B2 inhibitor that displays high selectivity vs related CYPs, good pharmacokinetic properties in rat and rhesus, and lead-like physical properties. In a rhesus pharmacodynamic model, compound 19 displays robust, dose-dependent aldosterone lowering efficacy, with no apparent effect on cortisol levels.

Factor XII full and partial null in rat confers robust antithrombotic efficacy with no bleeding.

This report aims at exploring quantitatively the relationship between FXII inhibition and thromboprotection. FXII full and partial null in rats were established via zinc finger nuclease-mediated knockout and siRNA-mediated knockdown, respectively. The rats were subsequently characterized in thrombosis and hemostasis models.

Discovery of Benzimidazole CYP11B2 Inhibitors with in Vivo Activity in Rhesus Monkeys.

We report the discovery of a benzimidazole series of CYP11B2 inhibitors. Hit-to-lead and lead optimization studies identified compounds such as 32, which displays potent CYP11B2 inhibition, high selectivity versus related CYP targets, and good pharmacokinetic properties in rat and rhesus. In a rhesus pharmacodynamic model, 32 produces dose-dependent aldosterone lowering efficacy, with no apparent effect on cortisol levels.

Platelet transfusion reverses bleeding evoked by triple anti-platelet therapy including vorapaxar, a novel platelet thrombin receptor antagonist.

Vorapaxar is a novel protease-activated receptor-1 (PAR-1) antagonist recently approved for the reduction of thrombotic cardiovascular events in patients with a history of myocardial infarction or with peripheral arterial disease. Patients who received vorapaxar in addition to standard of care antiplatelet therapy had an increased incidence of major bleeding events compared with placebo. To assess whether platelet transfusion can restore hemostasis in primates on triple antiplatelet therapy, template bleeding times were assessed concurrently in the buccal mucosa, finger pad, and distolateral tail of anesthetized cynomolgus macaques to evaluate bleeding with vorapaxar as either monotherapy or in combination with aspirin or aspirin and clopidogrel.

Titrating haemophilia B phenotypes using siRNA strategy: evidence that antithrombotic activity is separated from bleeding liability.

Haemophilia A and B are characterised by a life-long bleeding predisposition, and several lines of evidence suggest that risks of atherothrombotic events may also be reduced. Establishing a direct correlation between coagulation factor levels, thrombotic risks and bleeding propensity has long been hampered by an inability to selectively and specifically inhibit coagulation factor levels. Here, the exquisite selectivity of gene silencing combined with a gene knockout (KO) approach was used to define the relative contribution of factor IX (fIX) to thrombosis and primary haemostasis in the rat.

Proof-of-concept Studies for siRNA-mediated Gene Silencing for Coagulation Factors in Rat and Rabbit.

The present study aimed at establishing feasibility of delivering short interfering RNA (siRNA) to target the coagulation cascade in rat and rabbit, two commonly used species for studying thrombosis and hemostasis. siRNAs that produced over 90% mRNA knockdown of rat plasma prekallikrein and rabbit Factor X (FX) were identified from in vitro screens. An ionizable amino lipid based lipid nanoparticle (LNP) formulation for siRNA in vivo delivery was characterized as tolerable and exerting no appreciable effect on coagulability at day 7 postdosing in both species.

Rhesus monkey model for concurrent analyses of in vivo selectivity, pharmacokinetics and pharmacodynamics of aldosterone synthase inhibitors.

Introduction: In vivo profiles of aldosterone synthase inhibitors (ASIs) have been investigated utilizing various rodent models. Due to lack of CYP17 activity, rodents produce corticosterone rather than cortisol as that of humans, which raised concern to their effectiveness in translational pharmacological characterization of ASI.

Methods: A rhesus monkey model that combines a low sodium diet with adrenocorticotropin (ACTH) treatment was developed.

Factor XIIa inhibition by Infestin-4: in vitro mode of action and in vivo antithrombotic benefit.

Coagulation factor XII (FXII) plays a central role in initiating the intrinsic cascade of blood coagulation. Purified recombinant Human Albumin-tagged Infestin-4 (rHA-Infestin-4) is a recently described FXIIa inhibitor that displayed strong anticoagulant activity without compromising haemostasis in several animal models. We pursued detailed in vitro characterisation of rHA-Infestin-4 and demonstrated that it is a competitive inhibitor of FXIIa with slow on and off rate constants for binding (kon=5x10⁵ M⁻¹s⁻¹, koff=6x10⁻⁴ s⁻¹), it can block FXIIa activation of its physiological substrates (plasma prekallikrein and FXI), and it can inhibit ellagic acid-triggered thrombin generation in plasma.

Discovery of pyrazole carboxylic acids as potent inhibitors of rat long chain L-2-hydroxy acid oxidase.

Long chain L-2-hydroxy acid oxidase 2 (Hao2) is a peroxisomal enzyme expressed in the kidney and the liver. Hao2 was identified as a candidate gene for blood pressure (BP) quantitative trait locus (QTL) but the identity of its physiological substrate and its role in vivo remains largely unknown. To define a pharmacological role of this gene product, we report the development of selective inhibitors of Hao2.

Potent and Selective Inhibitors of Long Chain l-2-Hydroxy Acid Oxidase Reduced Blood Pressure in DOCA Salt-Treated Rats.

l-2-Hydroxy acid oxidase (Hao2) is a peroxisomal enzyme with predominant expression in the liver and kidney. Hao2 was recently identified as a candidate gene for blood pressure quantitative trait locus in rats. To investigate a pharmacological role of Hao2 in the management of blood pressure, selective Hao2 inhibitors were developed.

Discovery of a potent and selective small molecule hGPR91 antagonist.

GPR91, a 7TM G-Protein-Coupled Receptor, has been recently deorphanized with succinic acid as its endogenous ligand. Current literature indicates that GPR91 plays role in various pathophysiology including renal hypertension, autoimmune disease and retinal angiogenesis. Starting from a small molecule high-throughput screening hit 1 (hGPR91 IC(50): 0.

Transcriptome profiling and network analysis of genetically hypertensive mice identifies potential pharmacological targets of hypertension.

Hypertension is a condition with major cardiovascular and renal complications, affecting nearly a billion patients worldwide. Few validated gene targets are available for pharmacological intervention, so there is a need to identify new biological pathways regulating blood pressure and containing novel targets for treatment. The genetically hypertensive "blood pressure high" (BPH), normotensive "blood pressure normal" (BPN), and hypotensive "blood pressure low" (BPL) inbred mouse strains are an ideal system to study differences in gene expression patterns that may represent such biological pathways.

Discovery of pyrazolyl propionyl cyclohexenamide derivatives as full agonists for the high affinity niacin receptor GPR109A.

A series of pyrazolyl propionyl cyclohexenamides were discovered as full agonists for the high affinity niacin receptor GPR109A. The structure-activity relationship (SAR) studies were aimed to improve activity on GPR109A, reduce Cytochrome P450 2C8 (CYP2C8) and Cytochrome P450 2C9 (CYP2C9) inhibition, reduce serum shift and improve pharmacokinetic (PK) profiles.

Discovery of a biaryl cyclohexene carboxylic acid (MK-6892): a potent and selective high affinity niacin receptor full agonist with reduced flushing profiles in animals as a preclinical candidate.

Biaryl cyclohexene carboxylic acids were discovered as full and potent niacin receptor (GPR109A) agonists. Compound 1e (MK-6892) displayed excellent receptor activity, good PK across species, remarkably clean off-target profiles, good ancillary pharmacology, and superior therapeutic window over niacin regarding the FFA reduction versus vasodilation in rats and dogs.

Discovery of novel tricyclic full agonists for the G-protein-coupled niacin receptor 109A with minimized flushing in rats.

Tricyclic analogues were rationally designed as the high affinity niacin receptor G-protein-coupled receptor 109A (GPR109A) agonists by overlapping three lead structures. Various tricyclic anthranilide and cycloalkene carboxylic acid full agonists were discovered with excellent in vitro activity. Compound 2g displayed a good therapeutic index regarding free fatty acids (FFA) reduction and vasodilation effects in rats, with very weak cytochrome P450 2C8 (CYP2C8) and cytochrome P450 2C9 (CYP2C9) inhibition, and a good mouse pharmacokinetics (PK) profile.

Phenolic acids suppress adipocyte lipolysis via activation of the nicotinic acid receptor GPR109A (HM74a/PUMA-G).

Phenolic acids are found in abundance throughout the plant kingdom. Consumption of wine or other rich sources of phenolic acids, such as the "Mediterranean diet," has been associated with a lower risk of cardiovascular disease. The underlying mechanism(s), however, has remained unclear.

Role of GPR81 in lactate-mediated reduction of adipose lipolysis.

Heavy exercise or oxygen deficit often links with higher levels of arterial lactate and lower levels of plasma free fatty acids (FFA). Treatment with lactate reduces circulating levels of FFA in vivo and lipolysis in adipose tissues in vitro. However, the underlying mechanism has remained unclear.

Molecular modeling aided design of nicotinic acid receptor GPR109A agonists.

A homology model of the nicotinic acid receptor GPR109A was constructed based on the X-ray crystal structure of bovine rhodopsin. An HTS hit was docked into the homology model. Characterization of the binding pocket by a grid-based surface calculation of the docking model suggested that a larger hydrophobic body plus a polar tail would improve interaction between the ligand and the receptor.

3-(1H-tetrazol-5-yl)-1,4,5,6-tetrahydro-cyclopentapyrazole (MK-0354): a partial agonist of the nicotinic acid receptor, G-protein coupled receptor 109a, with antilipolytic but no vasodilatory activity in mice.

The discovery and profiling of 3-(1H-tetrazol-5-yl)-1,4,5,6-tetrahydro-cyclopentapyrazole (5a, MK-0354), a partial agonist of GPR109a, is described. Compound 5a retained the plasma free fatty acid lowering effects in mice associated with GPR109a agonism, but did not induce vasodilation at the maximum feasible dose. Moreover, preadministration of 5a blocked the flushing effect induced by nicotinic acid but not that induced by PGD2.

Discovery of orally bioavailable and novel urea agonists of the high affinity niacin receptor GPR109A.

A urea class of high affinity niacin receptor agonists was discovered. Compound 1a displayed good PK, better in vivo efficacy in reducing FFA in mouse than niacin, and no vasodilation in a mouse model. Compound 1q demonstrated equal affinity to GPR109A as niacin.