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PF-06651600 was developed as an irreversible inhibitor of JAK3 with selectivity over the other three JAK isoforms. A high level of selectivity toward JAK3 is achieved by the covalent interaction of PF-06651600 with a unique cysteine residue (Cys-909) in the catalytic domain of JAK3, which is replaced by a serine residue in the other JAK isoforms. Importantly, 10 other kinases in the kinome have a cysteine at the equivalent position of Cys-909 in JAK3. Five of those kinases belong to the TEC kinase family including BTK, BMX, ITK, RLK, and TEC and are also inhibited by PF-06651600. Preclinical data demonstrate that inhibition of the cytolytic function of CD8+ T cells and NK cells by PF-06651600 is driven by the inhibition of TEC kinases. On the basis of the underlying pathophysiology of inflammatory diseases such as rheumatoid arthritis, inflammatory bowel disease, alopecia areata, and vitiligo, the dual activity of PF-06651600 toward JAK3 and the TEC kinase family may provide a beneficial inhibitory profile for therapeutic intervention.

Recent years have seen a resurgence in drug discovery efforts aimed at the identification of covalent inhibitors which has led to an explosion of literature reports in this area and most importantly new approved therapies. These reports and breakthroughs highlight the significant investments made across the industry in SAR campaigns to optimize inhibitors. The potency of covalent inhibitors is generally considered to be more accurately described by the time-independent kinetic parameter k

Biochemical screening is a major source of lead generation for novel targets. However, during the process of small molecule lead optimization, compounds with excellent biochemical activity may show poor cellular potency, making structure-activity relationships difficult to decipher. This may be due to low membrane permeability of the molecule, resulting in insufficient intracellular drug concentration. The Cell Squeeze platform increases permeability regardless of compound structure by mechanically disrupting the membrane, which can overcome permeability limitations and bridge the gap between biochemical and cellular studies. In this study, we show that poorly permeable Janus kinase (JAK) inhibitors are delivered into primary cells using Cell Squeeze, inhibiting up to 90% of the JAK pathway, while incubation of JAK inhibitors with or without electroporation had no significant effect. We believe this robust intracellular delivery approach could enable more effective lead optimization and deepen our understanding of target engagement by small molecules and functional probes.

Recent years have seen a resurgence in drug discovery efforts aimed at the identification of covalent inhibitors which has led to an explosion of literature reports in this area and most importantly new approved therapies. These reports and breakthroughs highlight the significant investments made across the industry in SAR campaigns to optimize inhibitors. The potency of covalent inhibitors is generally considered to be more accurately described by the time-independent kinetic parameter kinact/Ki rather than a by a simple IC50 since the latter is a time-dependent parameter. Enzyme substrate concentrations are an additional important factor to consider when attempting to translate parameters derived from enzymology experiments to phenotypic behavior in a physiologically relevant cell-based system. Theoretical and experimental investigations into the relationship between IC50, time, substrate concentration and Kinact/Ki provided us with an effective approach to provide meaningful data for SAR optimization. The data we generated for our JAK3 irreversible covalent inhibitor program using IC50 values provided by enzyme assays with long incubations (>1h) coupled with physiological substrate concentration provided the medicinal chemist with optimal information in a rapid and efficient manner. We further document the wide applicability of this method by applying it to other enzymes systems where we have run covalent inhibitor programs.

The concept of target-specific covalent enzyme inhibitors appears attractive from both an efficacy and a selectivity viewpoint considering the potential for enhanced biochemical efficiency associated with an irreversible mechanism. Aside from potential safety concerns, clearance prediction of covalent inhibitors represents a unique challenge due to the inclusion of nontraditional metabolic pathways of direct conjugation with glutathione (GSH) or via GSH S-transferase-mediated processes. In this article, a novel pharmacokinetic algorithm was developed using a series of Pfizer kinase selective acrylamide covalent inhibitors based on their in vitro-in vivo extrapolation of systemic clearance in rats. The algorithm encompasses the use of hepatocytes as an in vitro model for hepatic clearance due to oxidative metabolism and GSH conjugation, and the use of whole blood as an in vitro surrogate for GSH conjugation in extrahepatic tissues. Initial evaluations with clinical covalent inhibitors suggested that the scaling algorithm developed from rats may also be useful for human clearance prediction when species-specific parameters, such as hepatocyte and blood stability and blood binding, were considered. With careful consideration of clearance mechanisms, the described in vitro-in vivo extrapolation approach may be useful to facilitate candidate optimization, selection, and prediction of human pharmacokinetic clearance during the discovery and development of targeted covalent inhibitors.

The proteins of the Bcl-2 family play key roles in the regulation of programmed cell death by controlling the integrity of the outer mitochondrial membrane and the initiation of the apoptosis process. We performed extensive molecular dynamics simulations to investigate the conformational flexibility of the Bcl-xL protein in both the apo and holo (with Bad peptide and ABT-737) states. The accelerated molecular dynamics method implemented in Amber 14 was used to produce broader conformational sampling of 200 ns simulations. The pocket mining method based on the variational implicit-solvent model tracks the dynamic evolution of the ligand binding site with a druggability score characterizing the maximal affinity achievable by a drug-like molecule. Major movements were observed around the α3-helical domain and the loop region connecting the α1 and α2 helices, reshaping the ligand interaction in the BH3 binding groove. Starting with the apo crystal structure, which is recognized as "closed" and undruggable, the BH3 groove transitioned between the "open" and "closed" states during equilibrium simulation. Further analysis revealed a small percentage of the trajectory frames (∼10%) with a moderate degree of druggability that mimic the ligand-bound states. The ability to attain and detect by computer simulation the most suitable conformational states for ligand binding in advance of compound synthesis and crystal structure solution is of immense value to the application and success of structure-based drug design.

The nuclear hormone receptor retinoic acid receptor-related orphan C2 (RORC2, also known as RORγt) is a promising target for the treatment of autoimmune diseases. A small molecule, inverse agonist of the receptor is anticipated to reduce production of IL-17, a key proinflammatory cytokine. Through a high-throughput screening approach, we identified a molecule displaying promising binding affinity for RORC2, inhibition of IL-17 production in Th17 cells, and selectivity against the related RORA and RORB receptor isoforms. Lead optimization to improve the potency and metabolic stability of this hit focused on two key design strategies, namely, iterative optimization driven by increasing lipophilic efficiency and structure-guided conformational restriction to achieve optimal ground state energetics and maximize receptor residence time. This approach successfully identified 3-cyano- N-(3-(1-isobutyrylpiperidin-4-yl)-1-methyl-4-(trifluoromethyl)-1 H-pyrrolo[2,3- b]pyridin-5-yl)benzamide as a potent and selective RORC2 inverse agonist, demonstrating good metabolic stability, oral bioavailability, and the ability to reduce IL-17 levels and skin inflammation in a preclinical in vivo animal model upon oral administration.

There has been significant interest in spleen tyrosine kinase (Syk) owing to its role in a number of disease states, including autoimmunity, inflammation, and cancer. Ongoing therapeutic programs have resulted in several compounds that are now in clinical use. Herein we report our optimization of the imidazopyrazine core scaffold of Syk inhibitors through the use of empirical and computational approaches. Free-energy perturbation (FEP) methods with MCPRO+ were undertaken to calculate the relative binding free energies for several alternate scaffolds. FEP was first applied retrospectively to determine if there is any predictive value; this resulted in 12 of 13 transformations being predicted in a directionally correct manner. FEP was then applied in a prospective manner to evaluate 17 potential targets, resulting in the realization of imidazotriazine 17 (3-(4-(3,4-dimethoxyphenylamino)imidazo[1,2-f][1,2,4]triazin-2-yl)benzamide), which shows a tenfold improvement in activity relative to the parent compound and no increase in atom count. Optimization of 17 led to compounds with nanomolar cellular activity.

Significant work has been dedicated to the discovery of JAK kinase inhibitors resulting in several compounds entering clinical development and two FDA approved NMEs. However, despite significant effort during the past 2 decades, identification of highly selective JAK3 inhibitors has eluded the scientific community. A significant effort within our research organization has resulted in the identification of the first orally active JAK3 specific inhibitor, which achieves JAK isoform specificity through covalent interaction with a unique JAK3 residue Cys-909. The relatively rapid resynthesis rate of the JAK3 enzyme presented a unique challenge in the design of covalent inhibitors with appropriate pharmacodynamics properties coupled with limited unwanted off-target reactivity. This effort resulted in the identification of 11 (PF-06651600), a potent and low clearance compound with demonstrated in vivo efficacy. The favorable efficacy and safety profile of this JAK3-specific inhibitor 11 led to its evaluation in several human clinical studies.

Interleukin-17A (IL-17A) is a principal driver of multiple inflammatory and immune disorders. Antibodies that neutralize IL-17A or its receptor (IL-17RA) deliver efficacy in autoimmune diseases, but no small-molecule IL-17A antagonists have yet progressed into clinical trials. Investigation of a series of linear peptide ligands to IL-17A and characterization of their binding site has enabled the design of novel macrocyclic ligands that are themselves potent IL-17A antagonists.

IL-17A is a pro-inflammatory cytokine that has been implicated in autoimmune and inflammatory diseases. Monoclonal antibodies inhibiting IL-17A signaling have demonstrated remarkable efficacy, but an oral therapy is still lacking. A high affinity IL-17A peptide antagonist (HAP) of 15 residues was identified through phage-display screening followed by saturation mutagenesis optimization and amino acid substitutions. HAP binds specifically to IL-17A and inhibits the interaction of the cytokine with its receptor, IL-17RA. Tested in primary human cells, HAP blocked the production of multiple inflammatory cytokines. Crystal structure studies revealed that two HAP molecules bind to one IL-17A dimer symmetrically. The N-terminal portions of HAP form a β-strand that inserts between two IL-17A monomers while the C-terminal section forms an α helix that directly blocks IL-17RA from binding to the same region of IL-17A. This mode of inhibition suggests opportunities for developing peptide antagonists against this challenging target.

Spleen tyrosine kinase is a non-receptor tyrosine kinase, overactivation of which is thought to contribute to autoimmune diseases as well as allergy and asthma. Protein kinases have a highly conserved ATP binding site, thus making challenging the design of selective small molecule inhibitors. It has been well documented that some protein kinases can be stabilized in their inactive conformations (Type-II inhibitors). Herein, we describe a protein structure/ligand-based approach to successfully identify ligands that bind to novel conformations of spleen tyrosine kinase. By utilizing kinase protein crystal structures both in the public domain (RCSB) and within Pfizer's protein crystal database, we report the discovery of the first spleen tyrosine kinase Type-II ligands. Compounds 1 and 3 were found to bind to the DFG-out conformation of spleen tyrosine kinase, while compound 2 binds to a DFG-in, C-Helix-out conformation. In this instance, the C-helix moved significantly to create a large hydrophobic pocket rarely seen in kinase protein crystal structures.

Hematopoietic prostaglandin D synthase (HPGDS) is primarly expressed in mast cells, antigen-presenting cells, and Th-2 cells. HPGDS converts PGH2 into PGD2, a mediator thought to play a pivotal role in airway allergy and inflammatory processes. In this letter, we report the discovery of an orally potent and selective inhibitor of HPGDS that reduces the antigen-induced response in allergic sheep.

A regioselective and efficient procedure for the N-alkylation of 3-iodoindazole with chlorodifluoromethane is described. The reaction was extensively optimized with regard to combinations of base, solvent, addition order as well as stoichiometry. A key reaction parameter was the generation of a 2-M stock solution of the gaseous reagent chlorodifluoromethane in acetonitrile. This allowed control of the quantity of carbene generated in the reaction mixture, providing good N1-regioselectivity between the two reactive indazole nitrogens. Upon optimization of this protocol 1-difluoromethyl-3-iodoindazole was synthesized on a preparative scale providing 14.2 grams of product in 59% yield.

In this paper, we describe a fluorescent antibacterial analog, 6, with utility as a competition probe to determine affinities of other antibacterial analogs for human serum albumin (HSA). Analog 6 bound to HSA with an affinity of 400+/-100 nM and the fluorescence was environmentally sensitive. With 370 nm excitation, environmental sensitivity was indicated by a quenching of the 530 nm emission when the probe bound to HSA. Displacement of dansylsarcosine from HSA by 6 indicated it competed with compounds that bound at site II (ibuprofen binding site) on HSA. Analog 6 also shifted the NMR peaks of an HSA bound oleic acid molecule that itself was affected by compounds that bound at site II. In addition to binding at site II, 6 interacted at site I (warfarin binding site) as indicated by displacement of dansylamide and the shifting of NMR peaks of an HSA bound oleic acid molecule affected by warfarin site binding. Additional evidence for multiple site interaction was discovered when a percentage of 6 could be displaced by either ibuprofen or phenylbutazone. A competition assay was established using 6 to determine relative affinities of other antibacterial inhibitors for HSA.

In the past few years a significant effort has been devoted by Pharmacia toward the discovery of novel antibiotics. We have recently described the identification of an anthranilic acid lead 1 and the optimization resulting in the advanced lead 2. In this report, we describe the preparation of several selected analogs to probe the dependency of this template for antibacterial activity and the affinity these compounds have for human serum albumin (HSA). These analogs illustrate that decreased affinity for HSA can be achieved while retaining relevant antibacterial activity. The most important factor for reduced HSA affinity is decrease in logP rather than a structural change.

In the past few years, a significant effort has been devoted by Pharmacia toward the discovery of novel antibiotics. We have recently described the identification of an anthranilic acid lead 1 and the optimization resulting in the advanced lead 2. In this report, we describe the preparation of several selected amide bioisosteres connecting the A- and the B-rings. The E-alkene provided a rigid analog with equal potency to the corresponding amide. This indicates that the amide is not a recognition element rather acts as an appropriate spatial linker of the two important aryl A and B rings. The work here clearly demonstrates that the amide linker can be replaced with several functionalities without significant deterioration in the MIC activity.

Discovery of novel antibacterial agents is a significant challenge. We have recently reported on our discovery of novel antibacterial agents in which we have rapidly optimized potency utilizing a parallel chemistry approach. These advanced leads suffer from high affinity for human serum albumin (HSA). In an effort to decrease the affinity for HSA we have prepared a series of heterocyclic analogs, which retained antibacterial activity and demonstrated reduced affinity for HSA.

The development of bacterial resistance is a significant problem in the treatment of infection, and the importance of research directed toward the discovery of novel agents to trea 2213t infections cannot be underestimated. In the past, discovery programs have focused on modification of natural products or existing classes of marketed antibacterial agents. A significant period of time lapsed between the introduction of the nalidixic acid-based quinolones and the next novel antibacterial agent (Zyvox). However, the advent of the genomics era has provided a wealth of new targets that afford the opportunity to discover novel antibacterial agents. This review reports on the state of antibacterial research directed toward the development of novel antibacterial agents with novel mechanisms of action for the calendar year, 2002. While variations on existing drug classes continue to appear, we have chosen to limit our discussion to novel classes of antibacterial agents which have not yet been marketed.

Screening of our compound collection identified PNU-92560, a 2-[1,3,4]thiadiazolo[3,2-a]pyrimidine-6-carboxamide, as a novel antibacterial agent. Extensive analogue development identified that the 2-position of the thiadiazole could be functionalized with a linker that would allow the compound to be attached to a solid support. The extreme insolubility of the analogues prevented the mechanism of action for these compounds to be determined utilizing traditional methodology. The solid-supported compounds were utilized as affinity columns to identify elongation factor Tu (EF-Tu) as a putative target for this class of compounds. The activity of the compounds in a metabolic labeling experiments and in translation assay supports the identity of the target for these compounds to be EF-Tu.

Spleen tyrosine kinase (SYK) is one of the more advanced small-molecule targets with regard to clinical development for treatment of inflammatory diseases. In this review we continue our analysis of the patent literature covering the time period 2011–2013. The analysis relates to any organization that has filed applications that explicitly discloses SYK as the intended target. In the last 2 years there has been a surge of application with a few new entries in a crowded field with the structural theme of compounds in these applications being a traditional type I ATP competitive inhibitor [ 1 ]. This overview of the SYK patent literature and the learning's of the inhibitors substitution patterns would be an important reading for anyone working in the area of SYK inhibitors.

Important resistance patterns in Gram-negative pathogens include active efflux of antibiotics out of the cell via a cellular pump and decreased membrane permeability. A 3-arylpiperidine derivative (1) has been identified by high-throughput assay as a potentiator with an IC(50) approximately 90 microM. This report details the evaluation of the tether length, aryl substitution and the importance of the fluorine on antibiotic accumulation. Evaluation of various tether lengths demonstrated that the two-carbon tethered analogues are optimal. Removal of the fluorine has a modest effect on antibiotic accumulation and the defluorinated analogue 17 is equally potent to the original lead 1.

Peptidyl deformylase (PDF) is a metallo protease that catalyzes the removal of a formyl group from the N-termini of prokaryotic prepared polypeptides, an essential step in bacterial protein synthesis. Screening of our compound collection using Staphylococcus aureus PDF afforded a very potent inhibitor with an IC 50 in the low nanomolar range. Unfortunately, the compound that contains a hydroxamic acid did not exhibit antibacterial activity (MIC). In order to address the lack of activity in the MIC assay and to determine what portion of the molecule was responsible for binding to PDF, we prepared several analogues. This paper describes our findings that the hydroxamic acid functionality found in 1 is mainly responsible for the high affinity to PDF. In addition, we identified an alternative class of PDF inhibitors, the N- hydroxy urea 18 , which has both PDF and antibacterial activity.

Two efficient and scaleable synthetic routes to previously unknown bifunctional tert-butyl 6-oxo-2-azaspiro[3.3]heptane-2-carboxylate are described. This compound and related intermediates described herein are useful for further selective derivation on the azetidine and cyclobutane rings providing a convenient entry point to novel compounds accessing chemical space complementary to piperidine ring systems.

The immobilized 1,3-regiospecific Rhizomucor miehei lipase (Lipozyme™) was employed to catalyze the transesterification reaction (acidolysis) of 1,2-diacyl-sn-glycero-3-phosphatidylcholine with n-3 polyunsaturated fatty acids under nonaqueous solvent-free conditions. With a concentrate of 55% eicosapentaenoic acid (EPA) and 30% docosahexaenoic acid (DHA) and pure phosphatidylcholine from egg yolk, phospholipids of 32% EPA and 16% DHA content were obtained, presumably as a mixture of phosphatidylcholine and lysophosphatidylcholine. 31P nuclear magnetic resonance (NMR) analysis turned out to be a valuable technique to study the details of the reactions involved. It revealed that when 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine was transesterified with 98% pure EPA, a substantial amount of hydrolysis side reaction took place (39%), leading to a product mixture of 39% phosphatidylcholine, 44% lysophosphatidylcholine, and 17% sn-glycerol-3-phosphatidylcholine. The lysophosphatidylcholine constituent comprised 70% EPA, whereas the phosphatidylcholine component contained 58% EPA. The 31P NMR technique provided valid information about the mechanism of the reaction. It became evident that a high dosage of lipase containing 5% water afforded optimal conditions for the optimal extent of EPA incorporation into the phospholipids, under which the extent of hydrolysis side reaction remained relatively high.

(+)-Crotanecine (1) is the necine base component of a number of pyrrolizidine alkaloids. This necine subunit is an amino triol bearing a primary allylic alcohol characterized by an all-cis relationship of its stereocenters. The synthesis of (+)-crotanecine has been accomplished in 10 steps and 10.2% overall yield. The key step in the asymmetric synthesis is a Lewis acid-promoted, tandem inter[4 + 2]/intra[3 + 2] cycloaddition between a (fumaroyloxy)nitroalkene 14 and chiral β-silylvinyl ether (−)-26. This synthesis serves to illustrate the synthetic versatility of the tandem cycloaddition to incorporate additional functionality.

The structure of the title compound, C31H39NO7Si, was determined and found to be a fused tricyclic nitroso acetal. Remarkable features include a twist-boat conformation of the tetrahydro-1,2-oxazine ring and a highly pyramidalized N atom [Sigma (angles) = 310.6(6) degrees]. Three of the contiguous stereocenters in the nitroso acetal are of the same correct relative and absolute configuration as is found in (+)-crotanecine.

Insofar as one of the fundamental objectives of organic synthesis is the construction of complex molecules from simpler ones, the importance of synthetic efficiency becomes immediately apparent and has been well-recognized.1 The increase in molecular complexity2 that necessarily accompanies the course of a synthesis provides a guide (and a measure) of synthetic efficiency. As a goal, one would like to optimally match the change in molecular complexity at each step with reactions of comparable synthetic complexity. Thus, the creation of many bonds, rings, and stereocenters in a single transformation is a necessary (though not sufficient) condition for high synthetic efficiency. The ultimate, perfect match would constitute single-step syntheses. More realistically, especially in view of the desire for general synthetic methods, the combination of mulScott E. Denmark was born in New York in 1953. He obtained an S.B. degree from MIT in 1975 and carried out research with Daniel Kemp and Richard Holm. His graduate studies at the ETH-Zurich with Albert Eschenmoser culminated with the D. Sc. Tech. degree in 1980. That same year he joined the faculty of the University of Illinois at UrbanasChampaign and was promoted to full professor in 1987. In 1991 he was named the Reynold C. Fuson Professor of Chemistry. His research interests are primarily in the invention of new synthetic reactions, structure and reactivity of organoelement reagents, and the origin of stereocontrol in fundamental carbon−carbon bond forming reactions.

Background Janus kinase (JAK) inhibitors targeting multiple JAK isoforms, JAK1 and 2, or selective against JAK1 are currently utilized in clinical practice or being developed for the treatment of various inflammatory and oncological diseases. No truly JAK3-selective inhibitor has reached the clinic. JAK3 signal in pairs with JAK1 to transduce signal elicited from six known cytokines (IL-2, IL-4, IL-7, IL-9, IL-15 & IL-21) binding to the gamma-common (g-c) chain cytokine receptors. JAK1 in addition to be required for g-c chain cytokine receptors signaling can also signal in pair with JAK2 or TYK2 and is required for other cytokine receptors signaling such as type I and II interferons and IL-6 and IL-10 families of cytokines. Other cytokines such as IL-12 and IL-23 are signaling via JAK2 and TYK2. Additionally, hematopoietic cytokines such as EPO and TPO as well as cytokines such as IL-3 and IL-5 signal via homodimers of JAK2. Objectives Develop a JAK3 selective inhibitor that will inhibit signaling from the g-c chain cytokine receptors at therapeutic dosing without inhibiting signaling by other cytokines such as hematopoietic cytokines, type I and II interferons and IL-6, IL-12 and IL-10 families of cytokines. Methods A JAK3-specific covalent ATP competitive inhibitor was assessed functionally in vitro and in vivo . Functional differentiation of JAK3 selective inhibition as compare to pan-JAK inhibition (tofacitinib) or JAK1 selective inhibition was also assessed. Results The inhibitor showed JAK3 selective inhibition in biochemical and cellular assays. It inhibits Th1 and Th17 cell differentiation and function. Importantly, sparing JAK1 inhibition, through the selectivity of this inhibitor, preserved anti-inflammatory functions such as the differentiation of alternatively activated M2a macrophages. Unlike pan-JAK or JAK1-selective inhibitors, JAK3 selective inhibition also preserved IL-10-dependent suppression of the production of pro-inflammatory cytokines following LPS treatment in macrophages while maintaining the suppression of TNF and IL-1 responses in IL-27-primed macrophages. Further characterization of this compound in the rat adjuvant-induced arthritis (AIA) demonstrated efficacy at reducing disease pathology. Conclusions We have identified the first trully selective JAK3 inhibitor with potency and properties that make it suitable for further clinical development. Disclosure of Interest J.-B. Telliez Employee of: Pfizer, L. Wang Employee of: Pfizer, J. Jussif Employee of: Pfizer, T. Lin Employee of: Pfizer, L. Li Employee of: Pfizer, E. Moy Employee of: Pfizer, W. Li Employee of: Pfizer, Y. Zhao Employee of: Pfizer, K. Crouse Employee of: Pfizer, P. Symanowicz Employee of: Pfizer, M. Hegen Employee of: Pfizer, M. E. Banker Employee of: Pfizer, F. Vincent Employee of: Pfizer, J. Clark Employee of: Pfizer, A. Thorarensen Employee of: Pfizer

Diseases such as rheumatoid arthritis, asthma and chronic allergies are thought to be the result of chronic inflammatory processes and are the subject of intense research by academia and pharmaceutical companies. One of the major proteins targeted by this research is Syk kinase. Since Syk is uniquely positioned at the origins of multiple inflammatory pathways in many cell types, inhibition of the activity of this kinase promises to slow the debilitating effects of chronic inflammation. This review will present a discussion of what has been learned through the efforts on this field of research and report on the recent progress toward clinical drug development. Initially, the biologic role of Syk in generating inflammation will be considered and some of the major genetic and animal studies that have been performed to validate Syk as a target will be described. In the second section, the structural features of the enzyme and approaches taken for the design of inhibitors will be presented. The third section will review the inhibitors that have been designed and chemotypes that have proved effective at inhibiting Syk. And finally, the results of clinical trials are described.

Spleen tyrosine kinase is a non-receptor tyrosine kinase, overactivation of which is thought to contribute to autoimmune diseases as well as allergy and asthma. Protein kinases have a highly conserved ATP binding site, thus making challenging the design of selective small molecule inhibitors. It has been well documented that some protein kinases can be stabilized in their inactive conformations (Type-II inhibitors). Herein, we describe a protein structure/ligand-based approach to successfully identify ligands that bind to novel conformations of spleen tyrosine kinase. By utilizing kinase protein crystal structures both in the public domain (RCSB) and within Pfizer's protein crystal database, we report the discovery of the first spleen tyrosine kinase Type-II ligands. Compounds 1 and 3 were found to bind to the DFG-out conformation of spleen tyrosine kinase, while compound 2 binds to a DFG-in, C-Helix-out conformation. In this instance, the C-helix moved significantly to create a large hydrophobic pocket rarely seen in kinase protein crystal structures.

The inhibition of hH-PGDS has been proposed as a potential target for the development of anti-allergic and anti-inflammatory drugs. Herein we describe our investigation of the binding pocket of this important enzyme and our observation that two water molecules bind to our inhibitors and the enzyme. A series of compounds were prepared to the probe the importance of the water molecules in determining the binding affinity of the inhibitors to the enzyme. The study provides insight into the binding requirements for the design of potent hH-PGDS inhibitors.

Methods for generating non-polar glyceryl ether lipids highly enriched with EPA or DHA or preparing them highly efficiently homogeneous with EPA, 1, or DHA, 2, by immobilized lipases is described.

A short and efficient asymmetric synthesis of the pyrrolizidine necine base (-)-hastanecine is described. The key reaction in the synthesis is a sequential inter [4+2]/inter [3+2] cycloaddition. The Lewis acid promoted [4+2] cycloaddition between 2-acyloxy nitroalkene 16 and chiral vinyl ether 17 afforded a nitronate which underwent facile [3+2] cycloaddition with dimethyl maleate. The resulting nitroso acetal 19 had all the required stereocenters for (-)-hastanecine. The critical unmasking of the nitroso acetal 19 employed a hydrogenolytic cleavage to give the 1-azabicyclo[3.3.0]octane skeleton of (-)-hastanecine

Bbstract; Nitroalkenes have proven to be extremely versatile 4x-components in heterodiene [4+2]-cycloadditions. Our efforts in this area have been intensively methodological with focus on the reaction scope, mechanism, stereoselectivity and transformations of the cycloadducts. The three modes of tandem inter [4+2]/ intra [3+2] cycloaddition, fused, spiro and bridged have been documented. An efficient and highly selective synthesis of (-)-hastanwine has been completed.

Discovery of small molecular inhibitors for treatment of rheumatoid arthritis is a major ongoing effort within the pharmaceutical industry. Spleen tyrosine kinase (SYK) is one of leading small molecular targets with regard to clinical development primarlly due to efforts by Rigel and Portola. In this review, we provide a comprehensive overview of the SYK patent landscape. The patent literature we evaluated relates to any organization that has filed applications that imply that SYK is the intended target. The interest in SYK was initiated in the early 2000’s with many organizations, including several large pharmaceutical companies, and has been active for years. In general, the structural theme of most of the compounds in these applications is a traditional ATP competitive inhibitor with each organization having a different hinge binding element. In general, the attachment to the hinge is an aryl amine that is decorated with a solubilizing group. The other substituents are broadly variable across the numerous companies indicating that SYK has significant flexibility in its interactions in that portion of the kinase. This overview of the SYK patent literature and the learnings of the inhibitors’ substitution patterns would be an important reference for anyone working in this area.

Herein we report the identification of two new fatty acid amide hydrolase (FAAH) inhibitor lead series with FAAH k(inact)/K(i) potency values greater than 1500M(-1)s(-1). The two novel spirocyclic cores, 7-azaspiro[3.5]nonane and 1-oxa-8-azaspiro[4.5]decane, clearly distinguished themselves from the other spirocyclic cores on the basis of their superior potency for FAAH. Lead compounds from these two series have suitable FAAH potency and selectivity for additional medicinal chemistry optimization.

Fatty acid amide hydrolase (FAAH) is an integral membrane serine hydrolase responsible for the degradation of fatty acid amide signaling molecules such as endocannabinoid anandamide (AEA), which has been shown to possess cannabinoid-like analgesic properties. Herein we report the optimization of spirocyclic 7-azaspiro[3.5]nonane and 1-oxa-8-azaspiro[4.5]decane urea covalent inhibitors of FAAH. Using an iterative design and optimization strategy, lead compounds were identified with a remarkable reduction in molecular weight and favorable CNS drug like properties. 3,4-Dimethylisoxazole and 1-methyltetrazole were identified as superior urea moieties for this inhibitor class. A dual purpose in vivo efficacy and pharmacokinetic screen was designed to be the key decision enabling experiment affording the ability to move quickly from compound synthesis to selection of preclinical candidates. On the basis of the remarkable potency, selectivity, pharmacokinetic properties and in vivo efficacy, PF-04862853 (15p) was advanced as a clinical candidate.

Discovery of small molecule inhibitors for treatment of rheumatoid arthritis is a major ongoing effort within the pharmaceutical industry. Spleen tyrosine kinase (SYK) is one of the leading small molecular targets with regard to clinical developments due to the efforts of Rigel and Portola.Diaminopyrimidines are one of the most prominent structural elements incorporated into the design of kinase inhibitors. This review provides an extensive overview of the patent estates for the leading discovery programs at Portola and Rigel on diaminopyrimidines and how their patent estates are in relationship with the competition and prior art.An overview of the patent landscape for diaminopyrimidines. In addition, the reader will be updated on what modifications in these scaffolds lead to very potent SYK inhibitors as judged by the applications. Finally, the authors will provide their best guess on what the structure is of Portola's recently announced clinical candidate.The Rigel and the Portola research organizations have filed a series of patent applications on diaminopyrimidines as inhibitors of SYK. The scope of these applications is broad in a crowded chemical space. These applications contain very broad claims and the future will tell how much of the generic space claimed will be allowed in granted patents.

A short and efficient asymmetric synthesis of the pyrrolizidine necine base (-)-hastanecine is described. The key reaction in the synthesis is a sequential inter [4+2]/inter [3+2] cycloaddition. The Lewis acid promoted [4+2] cycloaddition between 2-acyloxy nitroalkene 16 and chiral vinyl ether 17 afforded a nitronate which underwent facile [3+2] cycloaddition with dimethyl maleate. The resulting nitroso acetal 19 had all the required stereocenters for (-)-hastanecine. The critical unmasking of the nitroso acetal 19 employed a hydrogenolytic cleavage to give the 1-azabicyclo[3.3.0]octane skeleton of (-)-hastanecine

Bbstract; Nitroalkenes have proven to be extremely versatile 4x-components in heterodiene [4+2]-cycloadditions. Our efforts in this area have been intensively methodological with focus on the reaction scope, mechanism, stereoselectivity and transformations of the cycloadducts. The three modes of tandem inter [4+2]/ intra [3+2] cycloaddition, fused, spiro and bridged have been documented. An efficient and highly selective synthesis of (-)-hastanwine has been completed.

Insofar as one of the fundamental objectives of organic synthesis is the construction of complex molecules from simpler ones, the importance of synthetic efficiency becomes immediately apparent and has been well-recognized.1 The increase in molecular complexity2 that necessarily accompanies the course of a synthesis provides a guide (and a measure) of synthetic efficiency. As a goal, one would like to optimally match the change in molecular complexity at each step with reactions of comparable synthetic complexity. Thus, the creation of many bonds, rings, and stereocenters in a single transformation is a necessary (though not sufficient) condition for high synthetic efficiency. The ultimate, perfect match would constitute single-step syntheses. More realistically, especially in view of the desire for general synthetic methods, the combination of mulScott E. Denmark was born in New York in 1953. He obtained an S.B. degree from MIT in 1975 and carried out research with Daniel Kemp and Richard Holm. His graduate studies at the ETH-Zurich with Albert Eschenmoser culminated with the D. Sc. Tech. degree in 1980. That same year he joined the faculty of the University of Illinois at UrbanasChampaign and was promoted to full professor in 1987. In 1991 he was named the Reynold C. Fuson Professor of Chemistry. His research interests are primarily in the invention of new synthetic reactions, structure and reactivity of organoelement reagents, and the origin of stereocontrol in fundamental carbon−carbon bond forming reactions.

Important resistance patterns in Gram-negative pathogens include active efflux of antibiotics out of the cell via a cellular pump and decreased membrane permeability. A 3-arylpiperidine derivative (1) has been identified by high-throughput assay as a potentiator with an IC50 ∼90 μM. This report details the evaluation of the tether length, aryl substitution and the importance of the fluorine on antibiotic accumulation. Evaluation of various tether lengths demonstrated that the two-carbon tethered analogues are optimal. Removal of the fluorine has a modest effect on antibiotic accumulation and the defluorinated analogue 17 is equally potent to the original lead 1.

This paper describes the discovery of α-trifluoroketoacetamides as potent antibacterial agents against Gram-positive organisms. The initial SAR indicates that the aryl ethyl side chain is essential in maintaining antibacterial activity. The SAR observations have been utilized to design a bioisostere for the α-trifluoroketoacetamide with good activity against Gram-positive organisms.

Autotaxin is the enzyme responsible for the production of lysophosphatidic acid (LPA) from lysophosphatidyl choline (LPC), and it is up-regulated in many inflammatory conditions, including but not limited to cancer, arthritis, and multiple sclerosis. LPA signaling causes angiogenesis, mitosis, cell proliferation, and cytokine secretion. Inhibition of autotaxin may have anti-inflammatory properties in a variety of diseases; however, this hypothesis has not been tested pharmacologically because of the lack of potent inhibitors. Here, we report the development of a potent autotaxin inhibitor, PF-8380 [6-(3-(piperazin-1-yl)propanoyl)benzo[d]oxazol-2(3H)-one] with an IC(50) of 2.8 nM in isolated enzyme assay and 101 nM in human whole blood. PF-8380 has adequate oral bioavailability and exposures required for in vivo testing of autotaxin inhibition. Autotaxin's role in producing LPA in plasma and at the site of inflammation was tested in a rat air pouch model. The specific inhibitor PF-8380, dosed orally at 30 mg/kg, provided >95% reduction in both plasma and air pouch LPA within 3 h, indicating autotaxin is a major source of LPA during inflammation. At 30 mg/kg PF-8380 reduced inflammatory hyperalgesia with the same efficacy as 30 mg/kg naproxen. Inhibition of plasma autotaxin activity correlated with inhibition of autotaxin at the site of inflammation and in ex vivo whole blood. Furthermore, a close pharmacokinetic/pharmacodynamic relationship was observed, which suggests that LPA is rapidly formed and degraded in vivo. PF-8380 can serve as a tool compound for elucidating LPA's role in inflammation.