Structural and functional characterization of HQL-79, an orally selective inhibitor of human hematopoietic prostaglandin D synthase.
We determined the crystal structure of human hematopoietic prostaglandin (PG) D synthase (H-PGDS) as the quaternary complex with glutathione (GSH), Mg2+, and an inhibitor, HQL-79, having anti-inflammatory activities in vivo, at a 1.45-A resolution. In the quaternary complex, HQL-79 was found to reside within the catalytic cleft between Trp104 and GSH. HQL-79 was stabilized by interaction of a phenyl ring of its diphenyl group with Trp104 and by its piperidine group with GSH and Arg14 through water molecules, which form a network with hydrogen bonding and salt bridges linked to Mg2+. HQL-79 inhibited human H-PGDS competitively against the substrate PGH2 and non-competitively against GSH with Ki of 5 and 3 microm, respectively. Surface plasmon resonance analysis revealed that HQL-79 bound to H-PGDS with an affinity that was 12-fold higher in the presence of GSH and Mg2+ (Kd, 0.8 microm) than in their absence. Mutational studies revealed that Arg14 was important for the Mg2+-mediated increase in the binding affinity of H-PGDS for HQL-79, and that Trp104, Lys112, and Lys198 were important for maintaining the HQL-binding pocket. HQL-79 selectively inhibited PGD2 production by H-PGDS-expressing human megakaryocytes and rat mastocytoma cells with an IC50 value of about 100 microm but only marginally affected the production of other prostanoids, suggesting the tight functional engagement between H-PGDS and cyclooxygenase. Orally administered HQL-79 (30 mg/kg body weight) inhibited antigen-induced production of PGD2, without affecting the production of PGE2 and PGF2alpha, and ameliorated airway inflammation in wild-type and human H-PGDS-overexpressing mice. Knowledge about this structure of quaternary complex is useful for understanding the inhibitory mechanism of HQL-79 and should accelerate the structure-based development of novel anti-inflammatory drugs that inhibit PGD2 production specifically.
Department of Molecular Behavioral Biology, Osaka Bioscience Institute, 6-2-4, Furuedai, Suita, Osaka 565-0874, Japan.
Kosuke
K
Aritake
Yuji
Y
Kado
Tsuyoshi
T
Inoue
Masashi
M
Miyano
Yoshihiro
Y
Urade
D000284:Administration, Oral; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002460:Cell Line; D018360:Crystallography, X-Ray; D004791:Enzyme Inhibitors / Q000008:administration & dosage / Q000737:chemistry* / Q000494:pharmacology*; D006413:Hematopoietic System / Q000187:drug effects* / Q000201:enzymology*; D006801:Humans; D006860:Hydrogen Bonding; D066298:In Vitro Techniques; D019746:Intramolecular Oxidoreductases / Q000037:antagonists & inhibitors* / Q000737:chemistry*; D054834:Lipocalins; D051379:Mice; D008958:Models, Molecular; D015394:Molecular Structure; D010880:Piperidines / Q000008:administration & dosage / Q000737:chemistry* / Q000494:pharmacology*; D011487:Protein Conformation; D051381:Rats
C116322:4-benzhydryloxy-1-(3-(1H-tetrazol-5-yl-)-propyl)piperidine; D004791:Enzyme Inhibitors; D054834:Lipocalins; D010880:Piperidines; D019746:Intramolecular Oxidoreductases; C022466:prostaglandin R2 D-isomerase