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Pernilla Glader - One of the best experts on this subject based on the ideXlab platform.
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discovery of the oral Leukotriene C4 Synthase inhibitor 1s 2s 2 5 5 chloro 2 4 difluorophenyl 2 fluoro 2 methylpropyl amino 3 methoxypyrazin 2 yl carbonyl cyclopropanecarboxylic acid azd9898 as a new treatment for asthma
Journal of Medicinal Chemistry, 2019Co-Authors: Magnus Munck Af Rosenschöld, Petra Johannesson, Antonios Nikitidis, Christian Tyrchan, Hui-fang Chang, Robert Ronn, Dave Chapman, Victoria Ullah, Grigorios Nikitidis, Pernilla GladerAbstract:While bronchodilators and inhaled corticosteroids are the mainstay of asthma treatment, up to 50% of asthmatics remain uncontrolled. Many studies show that the cysteinyl Leukotriene cascade remains highly activated in some asthmatics, even those on high-dose inhaled or oral corticosteroids. Hence, inhibition of the Leukotriene C4 Synthase (LTC4S) enzyme could provide a new and differentiated core treatment for patients with a highly activated cysteinyl Leukotriene cascade. Starting from a screening hit (3), a program to discover oral inhibitors of LTC4S led to (1S,2S)-2-({5-[(5-chloro-2,4-difluorophenyl)(2-fluoro-2-methylpropyl)amino]-3-methoxypyrazin-2-yl}carbonyl)cyclopropanecarboxylic acid (AZD9898) (36), a picomolar LTC4S inhibitor (IC50 = 0.28 nM) with high lipophilic ligand efficiency (LLE = 8.5), which displays nanomolar potency in cells (peripheral blood mononuclear cell, IC50,free = 6.2 nM) and good in vivo pharmacodynamics in a calcium ionophore-stimulated rat model after oral dosing (in vivo, I...
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discovery of the oral Leukotriene C4 Synthase inhibitor 1s 2s 2 5 5 chloro 2 4 difluorophenyl 2 fluoro 2 methylpropyl amino 3 methoxypyrazin 2 yl carbonyl cyclopropanecarboxylic acid azd9898 as a new treatment for asthma
Journal of Medicinal Chemistry, 2019Co-Authors: Magnus Munck Af Rosenschöld, Petra Johannesson, Antonios Nikitidis, Christian Tyrchan, Hui-fang Chang, Robert Ronn, Dave Chapman, Victoria Ullah, Grigorios Nikitidis, Pernilla GladerAbstract:While bronchodilators and inhaled corticosteroids are the mainstay of asthma treatment, up to 50% of asthmatics remain uncontrolled. Many studies show that the cysteinyl Leukotriene cascade remains highly activated in some asthmatics, even those on high-dose inhaled or oral corticosteroids. Hence, inhibition of the Leukotriene C4 Synthase (LTC4S) enzyme could provide a new and differentiated core treatment for patients with a highly activated cysteinyl Leukotriene cascade. Starting from a screening hit (3), a program to discover oral inhibitors of LTC4S led to (1S,2S)-2-({5-[(5-chloro-2,4-difluorophenyl)(2-fluoro-2-methylpropyl)amino]-3-methoxypyrazin-2-yl}carbonyl)cyclopropanecarboxylic acid (AZD9898) (36), a picomolar LTC4S inhibitor (IC50 = 0.28 nM) with high lipophilic ligand efficiency (LLE = 8.5), which displays nanomolar potency in cells (peripheral blood mononuclear cell, IC50,free = 6.2 nM) and good in vivo pharmacodynamics in a calcium ionophore-stimulated rat model after oral dosing (in vivo, IC50,free = 34 nM). Compound 36 mitigates the GABA binding, hepatic toxicity signal, and in vivo toxicology findings of an early lead compound 7 with a human dose predicted to be 30 mg once daily.
Magnus Munck Af Rosenschöld - One of the best experts on this subject based on the ideXlab platform.
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discovery of the oral Leukotriene C4 Synthase inhibitor 1s 2s 2 5 5 chloro 2 4 difluorophenyl 2 fluoro 2 methylpropyl amino 3 methoxypyrazin 2 yl carbonyl cyclopropanecarboxylic acid azd9898 as a new treatment for asthma
Journal of Medicinal Chemistry, 2019Co-Authors: Magnus Munck Af Rosenschöld, Petra Johannesson, Antonios Nikitidis, Christian Tyrchan, Hui-fang Chang, Robert Ronn, Dave Chapman, Victoria Ullah, Grigorios Nikitidis, Pernilla GladerAbstract:While bronchodilators and inhaled corticosteroids are the mainstay of asthma treatment, up to 50% of asthmatics remain uncontrolled. Many studies show that the cysteinyl Leukotriene cascade remains highly activated in some asthmatics, even those on high-dose inhaled or oral corticosteroids. Hence, inhibition of the Leukotriene C4 Synthase (LTC4S) enzyme could provide a new and differentiated core treatment for patients with a highly activated cysteinyl Leukotriene cascade. Starting from a screening hit (3), a program to discover oral inhibitors of LTC4S led to (1S,2S)-2-({5-[(5-chloro-2,4-difluorophenyl)(2-fluoro-2-methylpropyl)amino]-3-methoxypyrazin-2-yl}carbonyl)cyclopropanecarboxylic acid (AZD9898) (36), a picomolar LTC4S inhibitor (IC50 = 0.28 nM) with high lipophilic ligand efficiency (LLE = 8.5), which displays nanomolar potency in cells (peripheral blood mononuclear cell, IC50,free = 6.2 nM) and good in vivo pharmacodynamics in a calcium ionophore-stimulated rat model after oral dosing (in vivo, I...
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discovery of the oral Leukotriene C4 Synthase inhibitor 1s 2s 2 5 5 chloro 2 4 difluorophenyl 2 fluoro 2 methylpropyl amino 3 methoxypyrazin 2 yl carbonyl cyclopropanecarboxylic acid azd9898 as a new treatment for asthma
Journal of Medicinal Chemistry, 2019Co-Authors: Magnus Munck Af Rosenschöld, Petra Johannesson, Antonios Nikitidis, Christian Tyrchan, Hui-fang Chang, Robert Ronn, Dave Chapman, Victoria Ullah, Grigorios Nikitidis, Pernilla GladerAbstract:While bronchodilators and inhaled corticosteroids are the mainstay of asthma treatment, up to 50% of asthmatics remain uncontrolled. Many studies show that the cysteinyl Leukotriene cascade remains highly activated in some asthmatics, even those on high-dose inhaled or oral corticosteroids. Hence, inhibition of the Leukotriene C4 Synthase (LTC4S) enzyme could provide a new and differentiated core treatment for patients with a highly activated cysteinyl Leukotriene cascade. Starting from a screening hit (3), a program to discover oral inhibitors of LTC4S led to (1S,2S)-2-({5-[(5-chloro-2,4-difluorophenyl)(2-fluoro-2-methylpropyl)amino]-3-methoxypyrazin-2-yl}carbonyl)cyclopropanecarboxylic acid (AZD9898) (36), a picomolar LTC4S inhibitor (IC50 = 0.28 nM) with high lipophilic ligand efficiency (LLE = 8.5), which displays nanomolar potency in cells (peripheral blood mononuclear cell, IC50,free = 6.2 nM) and good in vivo pharmacodynamics in a calcium ionophore-stimulated rat model after oral dosing (in vivo, IC50,free = 34 nM). Compound 36 mitigates the GABA binding, hepatic toxicity signal, and in vivo toxicology findings of an early lead compound 7 with a human dose predicted to be 30 mg once daily.
Bing K Lam - One of the best experts on this subject based on the ideXlab platform.
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the catalytic architecture of Leukotriene C4 Synthase with two arginine residues
Journal of Biological Chemistry, 2011Co-Authors: Hiromichi Saino, Yoshihide Kanaoka, Bing K Lam, Hideo Ago, Daisuke Irikura, Yoko Ukita, Atsushi Nisawa, Go Ueno, Masaki Yamamoto, Frank K AustenAbstract:Leukotriene (LT) C4 and its metabolites, LTD4 and LTE4, are involved in the pathobiology of bronchial asthma. LTC4 Synthase is the nuclear membrane-embedded enzyme responsible for LTC4 biosynthesis, catalyzing the conjugation of two substrates that have considerably different water solubility; that amphipathic LTA4 as a derivative of arachidonic acid and a water-soluble glutathione (GSH). A previous crystal structure revealed important details of GSH binding and implied a GSH activating function for Arg-104. In addition, Arg-31 was also proposed to participate in the catalysis based on the putative LTA4 binding model. In this study enzymatic assay with mutant enzymes demonstrates that Arg-104 is required for the binding and activation of GSH and that Arg-31 is needed for catalysis probably by activating the epoxide group of LTA4.
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crystal structure of a human membrane protein involved in cysteinyl Leukotriene biosynthesis
Nature, 2007Co-Authors: Hideo Ago, Yoshihide Kanaoka, Frank K Austen, Bing K Lam, Daisuke Irikura, Tatsuro Shimamura, Masashi MiyanoAbstract:This paper reports the X-ray crystal structure of Leukotriene C4 Synthase (the pivotal enzyme for the biosynthesis of Leukotriene C4), in a complex with glutathione at 3.3 Angstrom. This work may provide a structural basis for the development of new Leukotriene C4 Synthase inhibitors. The cysteinyl Leukotrienes, namely Leukotriene (LT)C4 and its metabolites LTD4 and LTE4, the components of slow-reacting substance of anaphylaxis1,2, are lipid mediators of smooth muscle constriction3,4,5 and inflammation6,7, particularly implicated in bronchial asthma8,9. LTC4 Synthase (LTC4S), the pivotal enzyme for the biosynthesis of LTC4 (ref. 10), is an 18-kDa integral nuclear membrane protein11,12 that belongs to a superfamily of membrane-associated proteins in eicosanoid and glutathione metabolism that includes 5-lipoxygenase-activating protein, microsomal glutathione S-transferases (MGSTs), and microsomal prostaglandin E Synthase 1 (ref. 13). LTC4S conjugates glutathione to LTA4, the endogenous substrate derived from arachidonic acid through the 5-lipoxygenase pathway14. In contrast with MGST2 and MGST3 (refs 15, 16), LTC4S does not conjugate glutathione to xenobiotics17. Here we show the atomic structure of human LTC4S in a complex with glutathione at 3.3 A resolution by X-ray crystallography and provide insights into the high substrate specificity for glutathione and LTA4 that distinguishes LTC4S from other MGSTs. The LTC4S monomer has four transmembrane α-helices and forms a threefold symmetric trimer as a unit with functional domains across each interface. Glutathione resides in a U-shaped conformation within an interface between adjacent monomers, and this binding is stabilized by a loop structure at the top of the interface. LTA4 would fit into the interface so that Arg 104 of one monomer activates glutathione to provide the thiolate anion that attacks C6 of LTA4 to form a thioether bond, and Arg 31 in the neighbouring monomer donates a proton to form a hydroxyl group at C5, resulting in 5(S)-hydroxy-6(R)-S-glutathionyl-7,9-trans-11,14-cis-eicosatetraenoic acid (LTC4). These findings provide a structural basis for the development of LTC4S inhibitors for a proinflammatory pathway mediated by three cysteinyl Leukotriene ligands whose stability and potency are different and by multiple cysteinyl Leukotriene receptors whose functions may be non-redundant.
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human Leukotriene C4 Synthase at 4 5 a resolution in projection
Structure, 2004Co-Authors: Ingeborg Schmidtkrey, Yoshihide Kanaoka, Frank K Austen, Bing K Lam, Daisuke Irikura, Winfried Haase, Deryck J Mills, Werner KuhlbrandtAbstract:Abstract Leukotriene (LT) C 4 Synthase, an 18 kDa integral membrane enzyme, conjugates LTA 4 with reduced glutathione to form LTC 4 , the parent compound of all cysteinyl Leukotrienes that play a crucial role in the pathobiology of bronchial asthma. We have calculated a projection map of recombinant human LTC 4 Synthase at a resolution of 4.5 A by electron crystallography, which shows that the enzyme is a trimer. A map truncated at 7.5 A visualizes four transmembrane α helices per protein monomer. The densities in projection indicate that most of the α helices run nearly perpendicular to the plane of the membrane. At this resolution, LTC 4 Synthase is strikingly similar to microsomal glutathione S-transferase 1, which belongs to the same gene family but bears little sequence identity and no resemblance in substrate specificity to the LTC 4 Synthase. These results provide new insight into the structure and function of membrane proteins involved in eicosanoid and glutathione metabolism.
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Leukotriene C4 Synthase
Prostaglandins Leukotrienes and Essential Fatty Acids, 2003Co-Authors: Bing K LamAbstract:LTC(4) Synthase conjugates LTA(4) with glutathione (GSH) to form LTC(4), the parent compound of the cysteinyl Leukotrienes. LTC(4) Synthase is a membrane protein that functions as a non-covalent homodimer of two 18-kDa polypeptides. The enzymatic activity of LTC(4) Synthase is augmented by Mg(2+) and inhibited by Co(2+) and the FLAP inhibitor MK-886. The K(m) and V(max) values of human LTC(4) Synthase are 3.6 microM and 1.3 micromol/mg/min for LTA(4) and 1.6 mM and 2.7 micromol/mg/min for GSH, respectively. The deduced amino acid sequence and the predicted secondary structure of LTC(4) Synthase share significant homology to FLAP, mGST-2, and mGST-3. Site-directed mutagenesis of LTC(4) Synthase suggests that Arg-51 is involved in opening the epoxide ring of LTA(4) and Tyr-93 in GSH thiolate anion formation during catalytic conjugation. LTC(4) Synthase is a TATA-less gene whose transcription involved both cell- and non-specific regulatory elements. LTC(4) Synthase gene disrupted mice grow normally, and are attenuated for innate and adaptive immune inflammatory permeability responses.
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Leukotriene C4 Synthase a pivotal enzyme in cellular biosynthesis of the cysteinyl Leukotrienes
Prostaglandins & Other Lipid Mediators, 2002Co-Authors: Bing K Lam, Frank K AustenAbstract:Leukotriene C4 Synthase (LTC4S) conjugates LTA4 with glutathione (GSH) to form LTC4, the parent compound of the cysteinyl LTs. LTC4S is an 18 kDa membrane protein and functions as a noncovalent homodimer. The enzyme activity of LTC4S is augmented by Mg2+ and inhibited by Co2+ and the function of 5-lipoxygenase (LO) activating protein (FLAP) inhibitor MK-886. The Km and Vmax values are 3.6 microM and 1.3 micromol/mg/min for LTA4 and 1.6 mM and 2.7 micromol/mg/min for GSH, respectively. The deduced amino acid sequence and the predicted secondary of LTC4S shares significant homology to FLAP, mGST-2 and mGST-3 which are all members of MAPEG protein superfamily. LTC4S and FLAP exhibited identical genomic organization of five exons and four introns. Site-directed mutagenesis suggests that Arg-51 is involved in opening the epoxide ring of LTA4 and Tyr-93 in GSH thiolate anion formation during catalytic conjugation. LTC4S is a TATA-less gene whose transcription assessed in a reporter construct involved both cell-specific and nonspecific regulatory elements. LTC4S-/- mice grow normally, and are attenuated for innate and adaptive immune inflammatory permeability responses.
Jesper Z. Haeggström - One of the best experts on this subject based on the ideXlab platform.
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development of smart cell free and cell based assay systems for investigation of Leukotriene C4 Synthase activity and evaluation of inhibitors
Biochimica et Biophysica Acta, 2016Co-Authors: Stefanie Liening, Jesper Z. Haeggström, Thea Kleinschmidt, Gerhard K E Scriba, Silke Rummler, Christina Weinigel, Oliver Werz, Ulrike GarschaAbstract:Cysteinyl Leukotrienes (cys-LTs) cause bronchoconstriction in anaphylaxis and asthma. They are formed by 5-lipoxygenase (5-LOX) from arachidonic acid (AA) yielding the unstable Leukotriene A4 (LTA4) that is subsequently conjugated with glutathione (GSH) by LTC4 Synthase (LTC4S). Cys-LT receptor antagonists and LTC4S inhibitors have been developed, but only the former have reached the market. High structural homology to related enzymes and lack of convenient test systems due to instability of added LTA4 have hampered the development of LTC4S inhibitors. We present smart cell-free and cell-based assay systems based on in situ-generated LTA4 that allow studying LTC4S activity and investigating LTC4S inhibitors. Co-incubations of microsomes from HEK293 cells expressing LTC4S with isolated 5-LOX efficiently converted exogenous AA to LTC4 (~1.3μg/200μg protein). Stimulation of HEK293 cells co-expressing 5-LOX and LTC4S with Ca2+-ionophore A23187 and 20μM AA resulted in strong LTC4 formation (~250ng/106 cells). MK-886, a well-known 5-LOX activating protein (FLAP) inhibitor that also acts on LTC4S, consistently inhibited LTC4 formation in all assay types (IC50=3.1-3.5μM) and we successfully confirmed TK04a as potent LTC4S inhibitor in these assay systems (IC50=17 and 300nM, respectively). We demonstrated transcellular LTC4 biosynthesis between neutrophils or 5-LOX-expressing HEK293 cells that produce LTA4 from AA and HEK293 cells expressing LTC4S that transform LTA4 to LTC4. In conclusion, our assay approaches are advantageous as the substrate LTA4 is generated in situ and are suitable for studying enzymatic functionality of LTC4S including site-directed mutations and evaluation of LTC4S inhibitors.
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phosphorylation of Leukotriene C4 Synthase at serine 36 impairs catalytic activity
Journal of Biological Chemistry, 2016Co-Authors: Shabbir Ahmad, Agnes Rinaldomatthis, Anders Wetterholm, Jimmy A Ytterberg, Madhuranayaki Thulasingam, Fredrik Tholander, Tomas Bergman, Roman A Zubarev, Jesper Z. HaeggströmAbstract:Leukotriene C4 Synthase (LTC4S) catalyzes the formation of the proinflammatory lipid mediator Leukotriene C4 (LTC4). LTC4 is the parent molecule of the cysteinyl Leukotrienes, which are recognized for their pathogenic role in asthma and allergic diseases. Cellular LTC4S activity is suppressed by PKC-mediated phosphorylation, and recently a downstream p70S6k was shown to play an important role in this process. Here, we identified Ser36 as the major p70S6k phosphorylation site, along with a low frequency site at Thr40, using an in vitro phosphorylation assay combined with mass spectrometry. The functional consequences of p70S6k phosphorylation were tested with the phosphomimetic mutant S36E, which displayed only about 20% (20 μmol/min/mg) of the activity of WT enzyme (95 μmol/min/mg), whereas the enzyme activity of T40E was not significantly affected. The enzyme activity of S36E increased linearly with increasing LTA4 concentrations during the steady-state kinetics analysis, indicating poor lipid substrate binding. The Ser36 is located in a loop region close to the entrance of the proposed substrate binding pocket. Comparative molecular dynamics indicated that Ser36 upon phosphorylation will pull the first luminal loop of LTC4S toward the neighboring subunit of the functional homotrimer, thereby forming hydrogen bonds with Arg104 in the adjacent subunit. Because Arg104 is a key catalytic residue responsible for stabilization of the glutathione thiolate anion, this phosphorylation-induced interaction leads to a reduction of the catalytic activity. In addition, the positional shift of the loop and its interaction with the neighboring subunit affect active site access. Thus, our mutational and kinetic data, together with molecular simulations, suggest that phosphorylation of Ser36 inhibits the catalytic function of LTC4S by interference with the catalytic machinery.
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Leukotriene A4 Hydrolase and Leukotriene C4 Synthase
Lipoxygenases in Inflammation, 2016Co-Authors: Agnes Rinaldo-matthis, Jesper Z. HaeggströmAbstract:Leukotrienes are potent proinflammatory and immune modulating lipid mediators synthesized along the 5-lipoxygenase pathway of arachidonic acid metabolism. Leukotriene B4 is one of the most potent chemotactic agents known while Leukotriene C4, D4, and E4 are a powerful smooth muscle contracting agents, particularly in the respiratory tract and microcirculation. The committed steps in the biosynthesis of Leukotriene B4 and C4 are catalyzed by the key enzymes Leukotriene A 4 hydrolase and Leukotriene C 4 Synthase, respectively. In this chapter we discuss the most recent advances in the understanding of these two enzymes at a structural, functional, and biological level.
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structure and inhibition of mouse Leukotriene C4 Synthase
PLOS ONE, 2014Co-Authors: Damian Niegowski, Agnes Rinaldomatthis, Shabbir Ahmad, Thea Kleinschmidt, Abdul Aziz Qureshi, Michaela Marback, Jesper Z. HaeggströmAbstract:Leukotriene (LT) C4 Synthase (LTC4S) is an integral membrane protein that catalyzes the conjugation reaction between the fatty acid LTA4 and GSH to form the pro-inflammatory LTC4, an important mediator of asthma. Mouse models of inflammatory disorders such as asthma are key to improve our understanding of pathogenesis and potential therapeutic targets. Here, we solved the crystal structure of mouse LTC4S in complex with GSH and a product analog, S-hexyl-GSH. Furthermore, we synthesized a nM inhibitor and compared its efficiency and binding mode against the purified mouse and human isoenzymes, along with the enzymes’ steady-state kinetics. Although structural differences near the active site and along the C-terminal α-helix V suggest that the mouse and human LTC4S may function differently in vivo, our data indicate that mouse LTC4S will be a useful tool in future pharmacological research and drug development.
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crystal structures of Leukotriene C4 Synthase in complex with product analogs implications for the enzyme mechanism
Journal of Biological Chemistry, 2014Co-Authors: Damian Niegowski, Agnes Rinaldomatthis, Shabbir Ahmad, Thea Kleinschmidt, Ulrika Olsson, Jesper Z. HaeggströmAbstract:Leukotriene (LT) C4 Synthase (LTC4S) catalyzes the conjugation of the fatty acid LTA4 with the tripeptide GSH to produce LTC4, the parent compound of the cysteinyl Leukotrienes, important mediators of asthma. Here we mutated Trp-116 in human LTC4S, a residue proposed to play a key role in substrate binding, into an Ala or Phe. Biochemical and structural characterization of these mutants along with crystal structures of the wild type and mutated enzymes in complex with three product analogs, viz. S-hexyl-, 4-phenyl-butyl-, and 2-hydroxy-4-phenyl-butyl-glutathione, provide new insights to binding of substrates and product, identify a new conformation of the GSH moiety at the active site, and suggest a route for product release, aided by Trp-116.
Victoria Ullah - One of the best experts on this subject based on the ideXlab platform.
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discovery of the oral Leukotriene C4 Synthase inhibitor 1s 2s 2 5 5 chloro 2 4 difluorophenyl 2 fluoro 2 methylpropyl amino 3 methoxypyrazin 2 yl carbonyl cyclopropanecarboxylic acid azd9898 as a new treatment for asthma
Journal of Medicinal Chemistry, 2019Co-Authors: Magnus Munck Af Rosenschöld, Petra Johannesson, Antonios Nikitidis, Christian Tyrchan, Hui-fang Chang, Robert Ronn, Dave Chapman, Victoria Ullah, Grigorios Nikitidis, Pernilla GladerAbstract:While bronchodilators and inhaled corticosteroids are the mainstay of asthma treatment, up to 50% of asthmatics remain uncontrolled. Many studies show that the cysteinyl Leukotriene cascade remains highly activated in some asthmatics, even those on high-dose inhaled or oral corticosteroids. Hence, inhibition of the Leukotriene C4 Synthase (LTC4S) enzyme could provide a new and differentiated core treatment for patients with a highly activated cysteinyl Leukotriene cascade. Starting from a screening hit (3), a program to discover oral inhibitors of LTC4S led to (1S,2S)-2-({5-[(5-chloro-2,4-difluorophenyl)(2-fluoro-2-methylpropyl)amino]-3-methoxypyrazin-2-yl}carbonyl)cyclopropanecarboxylic acid (AZD9898) (36), a picomolar LTC4S inhibitor (IC50 = 0.28 nM) with high lipophilic ligand efficiency (LLE = 8.5), which displays nanomolar potency in cells (peripheral blood mononuclear cell, IC50,free = 6.2 nM) and good in vivo pharmacodynamics in a calcium ionophore-stimulated rat model after oral dosing (in vivo, I...
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discovery of the oral Leukotriene C4 Synthase inhibitor 1s 2s 2 5 5 chloro 2 4 difluorophenyl 2 fluoro 2 methylpropyl amino 3 methoxypyrazin 2 yl carbonyl cyclopropanecarboxylic acid azd9898 as a new treatment for asthma
Journal of Medicinal Chemistry, 2019Co-Authors: Magnus Munck Af Rosenschöld, Petra Johannesson, Antonios Nikitidis, Christian Tyrchan, Hui-fang Chang, Robert Ronn, Dave Chapman, Victoria Ullah, Grigorios Nikitidis, Pernilla GladerAbstract:While bronchodilators and inhaled corticosteroids are the mainstay of asthma treatment, up to 50% of asthmatics remain uncontrolled. Many studies show that the cysteinyl Leukotriene cascade remains highly activated in some asthmatics, even those on high-dose inhaled or oral corticosteroids. Hence, inhibition of the Leukotriene C4 Synthase (LTC4S) enzyme could provide a new and differentiated core treatment for patients with a highly activated cysteinyl Leukotriene cascade. Starting from a screening hit (3), a program to discover oral inhibitors of LTC4S led to (1S,2S)-2-({5-[(5-chloro-2,4-difluorophenyl)(2-fluoro-2-methylpropyl)amino]-3-methoxypyrazin-2-yl}carbonyl)cyclopropanecarboxylic acid (AZD9898) (36), a picomolar LTC4S inhibitor (IC50 = 0.28 nM) with high lipophilic ligand efficiency (LLE = 8.5), which displays nanomolar potency in cells (peripheral blood mononuclear cell, IC50,free = 6.2 nM) and good in vivo pharmacodynamics in a calcium ionophore-stimulated rat model after oral dosing (in vivo, IC50,free = 34 nM). Compound 36 mitigates the GABA binding, hepatic toxicity signal, and in vivo toxicology findings of an early lead compound 7 with a human dose predicted to be 30 mg once daily.