Design and Synthesis of Metabolically Stable tRNA Synthetase Inhibitors Derived from Cladosporin.

Selective and specific inhibitors of Plasmodium falciparum lysyl-tRNA synthetase represent promising therapeutic antimalarial avenues. Cladosporin was identified as a potent P. falciparum lysyl-tRNA synthetase inhibitor, with an activity against parasite lysyl-tRNA synthetase >100-fold more potent than that of the activity registered against the human enzyme. Despite its compelling activity, cladosporin exhibits poor oral bioavailability; a critical requirement for antimalarial drugs. Thus, the quest to develop metabolically stable cladosporin-derived analogues, while retaining similar selectivity and potency to that of the natural compound, has begun. Chemogenomic profiling of a designed library allowed an entirely innovative structure-activity relationship study to be initiated; this shed light on structural evidence of a privileged scaffold with a unique activity against tRNA synthetases.

Oxysterols direct immune cell migration via EBI2.

Epstein-Barr virus-induced gene 2 (EBI2, also known as GPR183) is a G-protein-coupled receptor that is required for humoral immune responses; polymorphisms in the receptor have been associated with inflammatory autoimmune diseases. The natural ligand for EBI2 has been unknown. Here we describe the identification of 7α,25-dihydroxycholesterol (also called 7α,25-OHC or 5-cholesten-3ß,7α,25-triol) as a potent and selective agonist of EBI2. Functional activation of human EBI2 by 7α,25-OHC and closely related oxysterols was verified by monitoring second messenger readouts and saturable, high-affinity radioligand binding. Furthermore, we find that 7α,25-OHC and closely related oxysterols act as chemoattractants for immune cells expressing EBI2 by directing cell migration in vitro and in vivo. A critical enzyme required for the generation of 7α,25-OHC is cholesterol 25-hydroxylase (CH25H). Similar to EBI2 receptor knockout mice, mice deficient in CH25H fail to position activated B cells within the spleen to the outer follicle and mount a reduced plasma cell response after an immune challenge. This demonstrates that CH25H generates EBI2 biological activity in vivo and indicates that the EBI2-oxysterol signalling pathway has an important role in the adaptive immune response.

The discovery of potent, orally bioavailable pyrazolo and triazolopyrimidine CXCR2 receptor antagonists.

A hit-to-lead optimisation programme was carried out on the Novartis archive screening hit, pyrazolopyrimidine 2-methyl-5-((phenylthio)methyl)pyrazolo[1,5-a]pyrimidin-7-ol 1, resulting in the discovery of CXCR2 receptor antagonist 2-benzyl-5-(((2,3-difluorophenyl)thio)methyl)-[1,2,4]triazolo[1,5-a]pyrimidin-7-ol 14. The SAR was investigated by systematic variation of the pendant thiol, alkyl and pyrimidinol groups. Replacement of the pyrazolopyrimidine core with a triazolo alternative led to a dual series of antagonists with favourable biological and pharmacokinetic properties.

Discovery of Amino Alcohols as Highly Potent, Selective, and Orally Efficacious Inhibitors of Leukotriene A4 Hydrolase.

The discovery of chiral amino alcohols derived from our previously disclosed clinical LTA4H inhibitor LYS006 is described. In a biochemical assay, their optical antipodes showed similar potencies, which could be rationalized by the cocrystal structures of these compounds bound to LTA4H. Despite comparable stabilities in liver microsomes, they showed distinct in vivo PK properties. Selective O-phosphorylation of the (R)-enantiomers in blood led to clearance values above the hepatic blood flow, whereas the (S)-enantiomers were unaffected and exhibited satisfactory metabolic stabilities in vivo. Introduction of two pyrazole rings led to compound (S)-2 with a more balanced distribution of polarity across the molecule, exhibiting high selectivity and excellent potency in vitro and in vivo. Furthermore, compound (S)-2 showed favorable profiles in 16-week IND-enabling toxicology studies in dogs and rats. Based on allometric scaling and potency in whole blood, compound (S)-2 has the potential for a low oral efficacious dose administered once daily.

Piperidyl amides as novel, potent and orally active mGlu5 receptor antagonists with anxiolytic-like activity.

High throughput screening led to the identification of nicotinamide derivative 2 as a structurally novel mGluR5 antagonist. Optimization of the modular scaffold led to the discovery of 16m, a compound with high affinity for mGluR5 and excellent selectivity over other glutamate receptors. Compound 16m exhibits a favorable PK profile in rats, robust anxiolytic-like effects in three different animal models of fear and anxiety, as well as a good PK/PD correlation.

A segmented flow platform for on-demand medicinal chemistry and compound synthesis in oscillating droplets.

We report an automated flow chemistry platform that can efficiently perform a wide range of chemistries, including single/multi-phase and single/multi-step, with a reaction volume of just 14 µL. The breadth of compatible chemistries is successfully demonstrated and the desired products are characterized, isolated, and collected online by preparative HPLC/MS/ELSD.

Developing soluble polymers for high-throughput synthetic chemistry.

Soluble polymers have emerged as viable alternatives to resin supports across the broad spectrum of high-throughput organic chemistry. As the application of these supports become more widespread, issues such as broad-spectrum solubility and loading are becoming limiting factors and therefore new polymers are required to overcome such limitations. This article details the approach made within our group to new soluble polymer supports and specifically focuses on parallel libraries of block copolymers, de novo poly(styrene-co-chloromethylstyrene), PEG- stealth stars, and substituted poly(norbornylene)s.

Discovery of 3-(2,6-dichloro-3,5-dimethoxy-phenyl)-1-{6-[4-(4-ethyl-piperazin-1-yl)-phenylamino]-pyrimidin-4-yl}-1-methyl-urea (NVP-BGJ398), a potent and selective inhibitor of the fibroblast growth factor receptor family of receptor tyrosine kinase.

A novel series of N-aryl-N'-pyrimidin-4-yl ureas has been optimized to afford potent and selective inhibitors of the fibroblast growth factor receptor tyrosine kinases 1, 2, and 3 by rationally designing the substitution pattern of the aryl ring. On the basis of its in vitro profile, compound 1h (NVP-BGJ398) was selected for in vivo evaluation and showed significant antitumor activity in RT112 bladder cancer xenografts models overexpressing wild-type FGFR3. These results support the potential therapeutic use of 1h as a new anticancer agent.

N-sulfonylanthranilic acid derivatives as allosteric inhibitors of dengue viral RNA-dependent RNA polymerase.

A novel class of compounds containing N-sulfonylanthranilic acid was found to specifically inhibit dengue viral polymerase. The structural requirements for inhibition and a preliminary structure-activity relationship are described. A UV cross-linking experiment was used to map the allosteric binding site of the compound on the viral polymerase.

Preparation of new microgel polymers and their application as supports in organic synthesis.

A series of soluble microgel polymers have been synthesized using solution-phase polymerization reactions. In a systematic manner, several variables such as monomer concentration, cross-linker content, reaction solvent and reaction time were examined, and this provided an optimal polymer with both solubility and precipitation characteristics suitable for synthetic applications. Thus, a chemically functionalized microgel polymer was synthesized, and the utility of this polymer in the synthesis of a small array of oxazole compounds has been demonstrated. The advantage of the microgel polymers produced was that they exhibited solution viscosities lower than those of conventional linear polymers even at higher concentrations, and this was found to be beneficial for their precipitation properties. Compounds prepared using the described microgel polymer supports were obtained in similar yields and purity when compared with insoluble resins, and more importantly, the soluble polymer bound intermediates could be analyzed at each step using standard NMR techniques.

A parallel approach to the discovery of carrier delivery vehicles to enhance antigen immunogenicity.

As part of an ongoing effort to generate human and murine monoclonal antibodies against poorly immunogenic tumor-associated antigens we have merged the rapidly expanding disciplines of parallel polymer synthesis and controlled-release technology with immunology to produce a rapid and generic approach to improve the immunogenicity of carrier-bound antigens. The process involves three stages: An array of cross-linked hydrogel materials containing a carrier protein (at various concentrations) is prepared in parallel in one step. The array is then screened in mice to determine the most effective hydrogel at enhancing the immunogenicity of the encapsulated versus nonencapsulated carrier. Finally, the most efficient hydrogel is prepared containing the critical carrier-antigen conjugate and is used for immunization protocols. The strategy was successful for the BSA-glycoconjugate of the tumor-associated antigen GM3 analogue 4. When encapsulated within the hydrogel array member most efficient at elevating BSA immunogenicity, the BSA-4 glycoconjugate was significantly more immunogenic that when administered as a free antigen.

A tunable hydrogel for encapsulation and controlled release of bioactive proteins.

A N,N-dimethylacrylamide-based hydrogel (2) with the new cross-linker (ethylenedioxy) bis[2,2'-(N-acryloylamino)ethane] (1) has been prepared, and its physicochemical properties in aqueous solution were studied. Three different native proteins (lysozyme, bovine serum albumin, and rabbit IgG) were encapsulated within the polymeric matrix 2, and the kinetics of their release from the swollen hydrogel were determined. The rate of protein release exhibits a clear dependence on both the molecular weight of the protein and the amount of cross-linker utilized to prepare the hydrogel. This is reflected by the fact that the low molecular weight proteins are released at an increased rate versus higher molecular weight proteins. In addition a greater amount of protein is released from the hydrogels with a lower percentage of cross-linker. The polymerization procedure used in this study is sufficiently mild to safeguard the functional integrity of attendant biomolecules as determined by the retention of catalytic activity of encapsulated alpha-chymotrypsin and aldolase catalytic antibody 38C2. The potential utility of these hydrogels for the controlled release of bioactive agents in vivo is strengthened by both their lack of toxicity against human dermal fibroblasts and their lack of immunogenicity in mice.

Application of microgels as polymer supports for organic synthesis: preparation of a small phthalide library, a scavenger, and a borohydride reagent.

Microgel polymers containing a series of functional groups have been prepared. These microgels were composed of cross-linked poly(styrene) and were prepared by radical polymerization in solution. The microgel polymers exhibit good solubility in an array of different organic solvents, and in addition, they can be efficiently precipitated by the addition of methanol and isolated by filtration. A nine-member phthalide library was synthesized using an aminomethyl-functionalized microgel 5. To further demonstrate the versatility of these microgel polymers, tris(2-aminoethyl)amino microgel 11 was examined as a scavenger reagent to remove unreacted isocyanate after a urea synthesis. Finally, a microgel-supported ammonium borohydride reagent 14 was successfully prepared and used as a reducing agent. Notable features of these microgels are that in all applications the progress of the reaction could be monitored by standard NMR techniques and their preparation is performed using common glassware and techniques found in all organic laboratories.