Small-molecule allosteric activators of PDE4 long form cyclic AMP phosphodiesterases.

Cyclic AMP (cAMP) phosphodiesterase-4 (PDE4) enzymes degrade cAMP and underpin the compartmentalization of cAMP signaling through their targeting to particular protein complexes and intracellular locales. We describe the discovery and characterization of a small-molecule compound that allosterically activates PDE4 long isoforms. This PDE4-specific activator displays reversible, noncompetitive kinetics of activation (increased Vmax with unchanged Km), phenocopies the ability of protein kinase A (PKA) to activate PDE4 long isoforms endogenously, and requires a dimeric enzyme assembly, as adopted by long, but not by short (monomeric), PDE4 isoforms. Abnormally elevated levels of cAMP provide a critical driver of the underpinning molecular pathology of autosomal dominant polycystic kidney disease (ADPKD) by promoting cyst formation that, ultimately, culminates in renal failure. Using both animal and human cell models of ADPKD, including ADPKD patient-derived primary cell cultures, we demonstrate that treatment with the prototypical PDE4 activator compound lowers intracellular cAMP levels, restrains cAMP-mediated signaling events, and profoundly inhibits cyst formation. PDE4 activator compounds thus have potential as therapeutics for treating disease driven by elevated cAMP signaling as well as providing a tool for evaluating the action of long PDE4 isoforms in regulating cAMP-mediated cellular processes.

Targeting the P2X7 receptor in rheumatoid arthritis: biological rationale for P2X7 antagonism.

OBJECTIVES: This paper aims to explore the functional significance of the P2X7 receptor in preclinical models of rheumatoid arthritis. METHODS: Preclinical studies in vivo were performed using the rat streptococcal cell wall (SCW) arthritis model. Ex vivo cultures of lipopolysaccharide (LPS)/benzoylbenzoyl adenosine triphosphate (BzATP)-stimulated human monocytes were generated to test the activities of a novel, highly specific inhibitor of human P2X7, AZD9056, on interleukin (IL)-1 and IL-18 release. RESULTS: P2X7 receptor expression was detected in inflamed synovial tissue after onset of SCW-induced arthritis in rats. Inhibition of P2X7 therein led to reduced articular inflammation and erosive progression. No effect was noted on acute-phase responses. Ex vivo, AZD9056 inhibited IL-1 and IL-18 release to BzATP in LPS-primed human monocytes. CONCLUSIONS: P2X7 receptor inhibition could represent a novel approach to the treatment of inflammatory arthritis. However, confirmatory clinical studies are warranted to further explore this possibility.

The design of a novel series of muscarinic receptor antagonists leading to AZD8683, a potential inhaled treatment for COPD.

A novel series of muscarinic receptor antagonists was developed, with the aim of identifying a compound with high M3 receptor potency and a reduced risk of dose-limiting side effects with potential for the treatment of COPD. Initial compound modifications led to a novel cycloheptyl series, which was improved by focusing on a quinuclidine sub-series. A wide range of N-substituents was evaluated to determine the optimal substituent providing a high M3 receptor potency, high intrinsic clearance and high human plasma protein binding. Compounds achieving in vitro study criteria were selected for in vivo evaluation. Pharmacokinetic half-lives, inhibition of bronchoconstriction and duration of action, as well as systemic side effects, induced by the compounds were assessed in guinea-pig models. Compounds with a long duration of action and good therapeutic index were identified and AZD8683 was selected for progression to the clinic.

Discovery and evolution of phenoxypiperidine hydroxyamide dual CCR3/H1 antagonists. Part I.

The discovery of potent small molecule dual antagonists of the human CCR3 and H(1) receptors is described for the treatment of allergic diseases, for example, asthma and allergic rhinitis. Optimizing in vitro potency and metabolic stability, starting from a CCR1 lead compound, led to compound 20 with potent dual CCR3/H(1) activity and in vitro metabolic stability.

Discovery and evolution of phenoxypiperidine hydroxyamide dual CCR3/H1 antagonists. Part II: optimising in vivo clearance.

The second part of this communication focuses on the resolution of issues surrounding the series of hydroxyamide phenoxypiperidine CCR3/H(1) dual antagonists described in Part I. This involved further structural exploration directed at reducing metabolism and leading to the identification of compound 60 with a greatly improved in vivo pharmacokinetic profile.

The discovery of CCR3/H1 dual antagonists with reduced hERG risk.

A series of dual CCR3/H(1) antagonists based on a bispiperidine scaffold were discovered. Introduction of an acidic group overcame hERG liability. Bioavailability was optimised by modulation of physico-chemical properties and physical form to deliver a compound suitable for clinical evaluation.

Scaffold-hopping with zwitterionic CCR3 antagonists: identification and optimisation of a series with good potency and pharmacokinetics leading to the discovery of AZ12436092.

The discovery and optimisation of a series of zwitterionic CCR3 antagonists is described. Optimisation of the structure led to AZ12436092, a compound with excellent selectivity over activity at hERG and outstanding pharmacokinetics in preclinical species.

The discovery of AZD9164, a novel muscarinic M3 antagonist.

The optimization of a new series of muscarinic M(3) antagonists is described, leading to the identification of AZD9164 which was progressed into the clinic for evaluation of its potential as a treatment for COPD.

The effects of P2X7 receptor antagonists on the formation and function of human osteoclasts in vitro.

The P2X7 receptor (P2X7R) has been implicated in the process of multinucleation and cell fusion. We have previously demonstrated that blockade of P2X7Rs on osteoclast precursors using a blocking antibody inhibited multinucleated osteoclast formation in vitro, but that P2X7R KO mice maintain the ability to form multinucleated osteoclasts. This apparent contradiction of the role the P2X7R plays in multinucleation has prompted us to examine the effect of the most commonly used and recently available P2X7R antagonists on osteoclast formation and function. When added to recombinant RANKL and M-CSF human blood monocytes cultures, all but one compound, decreased the formation and function of multinucleated TRAP-positive osteoclasts in a concentration-dependent manner. These data provide further evidence for the role of the P2X7R in the formation of functional human multinucleated osteoclasts and highlight the importance of selection of antagonists for use in long-term experiments.

The discovery of new spirocyclic muscarinic M3 antagonists.

The optimisation of a new series of high potency muscarinic M3 antagonists, derived from high throughput screening library hit is described.

Releasing phosphorus from calcium for struvite fertilizer production from anaerobically digested dairy effluent.

Being a non-renewable resource and a source of potential water pollution, phosphorus could be recovered from animal manure in the form of struvite (MgNH4PO4.6H2O) to be used as a slow-release fertilizer. It was found recently that the majority of phosphorus in anaerobically digested dairy effluent is tied up in a fine suspended calcium-phosphate solid, thus becoming unavailable for struvite formation. Acidification and use of a chelating agent were investigated for converting the calcium-associated phosphorus in the digested effluent to dissolved phosphate ions, so that struvite can be produced. The results demonstrated that the phosphorus in the effluent was released into the solution by lowering the pH. In addition, the phosphorus concentration in the solution increased significantly with increased ethylenediaminetetraacetic acid (EDTA) concentration, as EDTA has a high stability constant with calcium. Most of the phosphorus (91%) was released into the solution after adding EDTA. Further, the freed phosphorus ion precipitated out as struvite provided that sufficient magnesium ions (Mg2+) were present in the solution. Furthermore, the phase structure of the solid precipitate obtained from the EDTA treatment matched well with standard struvite, based on the data from X-ray diffraction analysis. These results provide methods for altering the forms of phosphorus for the design and application of phosphorus-removal technologies for dairy wastewater management.

Discovery of potent and selective adamantane-based small-molecule P2X(7) receptor antagonists/interleukin-1beta inhibitors.

A novel series of antagonists of the human P2X7 receptor is described. Modification of substituents enabled identification of compounds selective for the rat P2X7 receptor and provides useful pharmacological tools for evaluation of the role of P2X7 in disease.

Coordinating ecological restoration options analysis and risk assessment to improve environmental outcomes.

Ecological risk assessment as currently practiced has hindered consideration of ecosystem services endpoints and restoration goals in the environmental management process. Practitioners have created barriers between procedures to clean up contaminated areas and efforts to restore ecosystem functions. In this article, we examine linkages between contaminant risk assessment approaches and restoration efforts with the aim of identifying ways to improve environmental outcomes. We advocate that project managers and other stakeholders use an ecological planning framework, with restoration options included upfront in the risk assessment. We also considered the opportunities to incorporate ecosystem services as potential assessment endpoints in the Problem Formulation stages of a risk assessment. Indeed, diverse perspectives of stakeholders are central to understand the relevance of social, cultural, economic, and regional ecology as influences on future use options for the landscape being restored. The measurement endpoints used to characterize the existing ecological conditions for selected ecosystem services can also be used to evaluate restoration success. A regional, landscape, or seascape focus is needed throughout the risk assessment process, so that restoration efforts play a more prominent role in enhancing ecosystem services. In short, we suggest that practitioners begin with the question of "how can the ecological risk assessment inform the decision on how best to restore the ecosystem?"

Hit-to-Lead studies: the discovery of potent adamantane amide P2X7 receptor antagonists.

A Hit-to-Lead optimisation programme was carried out on the adamantane high throughput screening hit 1 resulting in the discovery of a number of potent P2X(7) antagonists.