The transient receptor potential (TRP) family of ion channels comprises nonselective cation channels that respond to a wide range of chemical and thermal stimuli. TRPM8, a member of the melastatin subfamily, is activated by cold temperatures (<28 °C), and antagonists of this channel have the potential to treat cold induced allodynia and hyperalgesia. However, TRPM8 has also been implicated in mammalian thermoregulation and antagonists have the potential to induce hypothermia in patients. We report herein the identification and optimization of a series of TRPM8 antagonists that ultimately led to the discovery of PF-05105679. The clinical finding with this compound will be discussed, including both efficacy and its ability to affect thermoregulation processes in humans.

In ongoing studies towards novel hepatitis C virus (HCV) therapeutics, inhibitors of nonstructural protein 5A (NS5A) were evaluated. Specifically, starting from previously reported lead compounds, peripheral substitution patterns of a series of biaryl-linked pyrrolidine NS5A replication complex inhibitors were probed and structure-activity relationships were elucidated. Using molecular modelling and a supercritical fluid chromatographic (SFC) technique, intramolecular H-bonding and peripheral functional group topology were evaluated as key determinants of activity and membrane permeability. The novel compounds exhibited retained potency as compared with the lead compounds, and also showed promising results against a panel of resistance viruses. Together, the results of the study take us a step closer towards understanding the potency of daclatasvir, a clinical candidate upon which the compounds were based, and to designing improved analogues as second-generation antiviral agents targeting NS5A.

Nonstructural protein 5A (NS5A) represents a novel target for the treatment of hepatitis C virus (HCV). Daclatasvir, recently reported by Bristol-Myers-Squibb, is a potent NS5A inhibitor currently under investigation in phase 3 clinical trials. While the performance of daclatasvir has been impressive, the emergence of resistance could prove problematic and as such, improved analogues are being sought. By varying the biphenyl-imidazole unit of daclatasvir, novel inhibitors of HCV NS5A were identified with an improved resistance profile against mutant strains of the virus while retaining the picomolar potency of daclatasvir. One compound in particular, methyl ((S)-1-((S)-2-(4-(4-(6-(2-((S)-1-((methoxycarbonyl)-L-valyl)pyrrolidin-2-yl)-1H-imidazol-5-yl)quinoxalin-2-yl)phenyl)-1H-imidazol-2-yl)pyrrolidin-1-yl)-3-methyl-1-oxobutan-2-yl)carbamate (17), exhibited very promising activity and showed good absorption and a long predicted human pharmacokinetic half-life. This compound represents a promising lead that warrants further evaluation.

Several non-benzimidazole containing inhibitors of respiratory syncytial virus are described. Core template modification, analysis of antiviral activity, physicochemistry and optimisation of properties led to the thiazole-imidazole 13, that showed a good potency and pharmacokinetic profile in the rat.

A high throughput screen of the Pfizer compound collection was carried out using a hepatitis C virus (HCV) genotype 1b subgenomic replicon cell line. Those confirmed hits that demonstrated broad spectrum activity without overt cytotoxicity were further evaluated, leading to the identification of a series of pyrrolopyridines with excellent antiviral activity in a fully infectious HCV cell-based assay and pharmacokinetic properties.

Aldehyde oxidase (AO) is a molybdenum-containing enzyme distributed throughout the animal kingdom and capable of metabolising a wide range of aldehydes and N-heterocyclic compounds. Although metabolism by this enzyme in man is recognised to have significant clinical impact where human AO activity was not predicted by screening in preclinical species, there is very little reported literature offering real examples where drug discoverers have successfully designed away from AO oxidation. This article reports on some strategies adopted in the Pfizer TLR7 agonist programme to successfully switch off AO metabolism that was seen principally in the rat.

The development and implementation of a scalable process for the manufacture of the Toll-like receptor (TLR7) agonist PF-4171455 (1) is described. Initial routes used to synthesise 1 in milligram quantities were unsuitable for large-scale synthesis to provide bulk material. As part of the transfer between Medicinal Chemistry and Research-API, collaboration provided a fit for purpose route for the kilo-scale synthesis of 1. Key aspects of the synthesis included (i) a safe and practical synthesis of a key nitropyridone intermediate 7 over four steps, (ii) a sequential regioselective chlorination to selectively functionalise 7 and (iii) use of a carbamate as a tethered carbonyl group, allowing an efficient regiospecific synthesis of 1.

Cette invention concerne des composes de formule (I) et leurs sels pharmaceutiquement acceptables; des compositions les contenant; et l'utilisation desdits composes a titre d'inhibiteurs de replication du VHC.

A flexible route which enables access to derivatives of 4-amino-1,3-dihydroimidazo[4,5-c]pyridin-2-ones is described. Issues of selectivity, reaction safety, and low yields in original routes are overcome with the key improvements to the route, including a Negishi cross-coupling and use of a carbamate as a protecting group and intrinsic carbonyl source. The new route enables variation of C-6 and N-1 substituents.

The synthesis and structure–activity relationships of a series of novel interferon inducers are described. Pharmacokinetic studies and efficacy assessment of a series of 8-oxo-3-deazapurine analogues led to the identification of compound 33, a potent and selective agonist of the TLR7 receptor with an excellent in vivo efficacy profile in a mouse model.

The discovery of a series of highly potent and novel TLR7 agonist interferon inducers is described. Structure- activity relationships are presented, along with pharmacokinetic studies of a lead molecule from this series of N9-pyridylmethyl-8-oxo-3-deazapurine analogues. A rationale for the very high potency observed is offered. An investigation of the clearance mechanism of this class of compounds in rat was carried out, resulting in aldehyde oxidase mediated oxidation being identified as a key component of the high clearance observed. A possible solution to this problem is discussed. (C) 2011 Elsevier Ltd. All rights reserved.

A series of heterocycle analogues of an adenine template were explored for TLR7 agonist potency and pharmacokinetics. One compound was identified with an excellent pharmacokinetic, in vitro potency and in vivo interferon induction profile in a mouse model, and was selected for further pre-clinical evaluation as a potential treatment for hepatitis C viral infection.

The current standard of care for hepatitis C virus (HCV)-infected patients consists of lengthy treatment with interferon and ribavirin. To increase the effectiveness of HCV therapy, future regimens will incorporate multiple direct-acting antiviral (DAA) drugs. Recently, the HCV-encoded NS5A protein has emerged as a promising DAA target. Compounds targeting NS5A exhibit remarkable potency in vitro and demonstrate early clinical promise, suggesting that NS5A inhibitors could feature in future DAA combination therapies. Since the mechanisms through which these molecules operate are unknown, we have used NS5A inhibitors as tools to investigate their modes of action. Analysis of replicon-containing cells revealed dramatic phenotypic alterations in NS5A localization following treatment with NS5A inhibitors; NS5A was redistributed from the endoplasmic reticulum to lipid droplets. The NS5A relocalization did not occur in cells treated with other classes of HCV inhibitors, and NS5A-targeting molecules did not cause similar alterations in the localization of other HCV-encoded proteins. Time course analysis of the redistribution of NS5A revealed that the transfer of protein to lipid droplets was concomitant with the onset of inhibition, as judged by the kinetic profiles for these compounds. Furthermore, analysis of the kinetic profile of inhibition for a panel of test molecules permitted the separation of compounds into different kinetic classes based on their modes of action. Results from this approach suggested that NS5A inhibitors perturbed the function of new replication complexes, rather than acting on preformed complexes. Taken together, our data reveal novel biological consequences of NS5A inhibition, which may help enable the development of future assay platforms for the identification of new and/or different NS5A inhibitors.

David C. Pryde,* Deepak Dalvie, Qiyue Hu, Peter Jones, R. Scott Obach, ) and Thien-Duc Tran WorldWide Medicinal Chemistry, Pfizer Global Research and Development, Sandwich, Kent, CT13 9NJ, England, Pharmacokinetics, Dynamics andMetabolism, Pfizer Global Research andDevelopment, 10628 ScienceCenterDrive, La Jolla, California 92121, WorldWide Medicinal Chemistry, Pfizer Global Research and Development, 10628 Science Center Drive, La Jolla, California 92121, and ) Pharmacokinetics, Dynamics and Metabolism, Pfizer Global Research and Development, Eastern Point Road, Groton, Connecticut 06340

Small molecule fluorometric boron dipyrromethene probes were developed to bind hepatitis C virus-encoded NS5A protein and aid subcellular distribution studies. These molecules did not co-locate with NS5A, therefore alternative ‘silent’ azide reporters were used to obtain a more relevant picture of their distribution. Following pre-incubation with replicon cells, click chemistry was used to append a fluorophore to the azide that confirmed the co-localisation of the small molecule with the NS5A protein, thus providing greater insight into the antiviral mode of action of this chemotype.

A series of piperazine derivatives were designed and synthesised as gp120-CD4 inhibitors. SAR studies led to the discovery of potent inhibitors in a cell based anti viral assay represented by compounds 9 and 28. The rat pharmacokinetic and antiviral profiles of selected compounds are also presented. Crown Copyright (C) 2009 Published by Elsevier Ltd. All rights reserved.

The synthesis and structure-activity relationship of a series of novel gp120-CD4 inhibitors are described. Pharmacokinetic studies and antiviral spectrum assessment of lead compounds led to the identification of compound 36, a potent gp120-CD4 inhibitor which exhibited antiviral potency across a spectrum of 25 clade B isolates.