Prostaglandin D2 11-ketoreductase Research Articles

Unveiling a Novel Solvatomorphism of Anti-inflammatory Flufenamic Acid: X-ray Structure, Quantum Chemical, and In Silico Studies.

This paper delves into the polymorphism of 2-[3-(trifluoromethyl)anilino]benzoic acid, commonly referred to as flufenamic acid (FA), a pharmaceutical agent employed in treating inflammatory conditions. The central focus of the study is on a newly unearthed solvatomorphic structure of FA in methanol (FAM), and a thorough comparison is conducted with the commercially available standard structure. Employing a comprehensive approach, including X-ray crystallography, Hirshfeld surface analysis, density functional theory (DFT), molecular docking, and molecular dynamics (MD) simulations, the research aims to unravel the structural and functional implications of solvatomorphism. The X-ray crystal structure analysis brings to light notable differences between the standard FA and solvatomorphic FAM, showcasing variations in intermolecular interactions and crystal packing. Key features such as hydrogen bonding, π···π stacking, and C-H···π interactions are identified as influential factors shaping the stability and conformation of the compounds. Hirshfeld surface analysis further quantifies the nature and contribution of intermolecular interactions, providing a comprehensive perspective on molecular stability. Density functional theory offers valuable electronic structure insights, highlighting disparities in frontier molecular orbitals between FA and FAM. Molecular docking studies against prostaglandin D2 11-ketoreductase explore potential drug interactions, unveiling distinct binding modes and hydrogen bonding patterns that shed light on how the solvatomorphic structure may impact drug-target interactions. In-depth molecular dynamics simulations over 100 ns investigate the stability of the protein-ligand complex, with root mean square deviation and root mean square fluctuation analyses revealing minimal deviations and affirming the stability of FAM within the active site of the target protein.

Molecular Docking and Dynamics Simulations of Ammi visnaga L. Constituents as Antimelanogenic, Anti-Inflammatory and Anticoagulant Agents.

In this study, anti-melanogenic, anti-inflammatory and anti-coagulant potentials of eighteen selected constituents of Ammi visnaga L. were investigated by Induced Fit Docking (IFD) and molecular dynamic simulation with Schrödinger software. The binding free energies of the selected natural compounds were computed by means of ΔG MM-GBSA studies. Anti-melanogetic activity of the constituent against agaricus bisporus tyrosinase, Priestia megaterium tyrosinase and Homo sapiens tyrosinase were evaluated. The result showed that apiumetin had more negative binding free energy against three tyrosinase enzymes than cognate ligands, tropolone and kojic acid. Docking analysis was also performed to predict the constituents with anti-inflammatory activity against human Tumor necrosis factor, Cyclooxygenase-2, Prostaglandin D2 11-ketoreductase AKR1C3 and Prostaglandin reductase PTGR2. The results showed that pyranocoumarins (visnadin, dihydrosamidin, samidin) have more negative binding free energy against Cyclooxygenase-2 and Prostaglandin D2 11-ketoreductase receptors than cognate drugs, rofecoxib and indomethacin. In addition, docking analysis shows that pyranocoumarins, apiumetin and cimifugin have more negative binding free energy against Vitamin K epoxide reductase than S-warfarin drug, predicting that they have anticoagulant activity. Furthermore, the constituents and their cognate drugs were subjected to 100 ns MD Simulation to predict their stability at the active sites of the enzymes.

Mouse Akr1cl gene product is a prostaglandin D2 11-ketoreductase with strict substrate specificity

A mouse gene, Akr1cl, encodes a member of the aldo-keto reductase 1C subfamily (AKR1CL), whose function, however, remains unknown. Here, we show that the recombinant AKR1CL is an NADPH-dependent reductase of prostaglandin (PG) D2 (Km 3.2 μM, kcat 5.6 min−1) and oxidizes 9α,11β-PGF2 (Km 30 μM, kcat 3.3 min−1) in the reverse reaction. In contrast, it did not exhibit oxidoreductase activity towards other PGs (E2, A1, B2 and F2α), steroids and nonsteroidal carbonyls and alcohols, which are substrates of other mammalian AKR1C subfamily enzymes. The enzyme activity was inhibited by estradiol, quercetin, benzbromarone, ethacrynic acid and flufenamic acid, of which estradiol was the most potent competitive inhibitor (Ki 3.2 μM). The mRNA for AKR1CL was expressed abundantly in mouse testis, ovary and adrenal gland, and at low levels in the brain, lung, small intestine and prostate. Thus, AKR1CL is the first PGD2 11-ketoreductase with strict substrate specificity in mammals. The site-directed mutagenesis of P85 in AKR1CL to the corresponding residue, W, in other mammalian AKR1C subfamily enzymes resulted in broad substrate specificity for nonsteroidal carbonyls and alcohols, suggesting that P85 plays a critical role in the strict specificity for PGD2 and 9α,11β-PGF2.

Episode-Specific Differential Gene Expression of Peripheral Blood Mononuclear Cells in Rapid Cycling Supports Novel Treatment Approaches

Molecular mechanisms underlying bipolar affective disorders are unknown. Difficulties arise from genetic and phenotypic heterogeneity of patients and the lack of animal models. Thus, we focused on only one patient (n = 1) with an extreme form of rapid cycling. Ribonucleic acid (RNA) from peripheral blood mononuclear cells (PBMC) was analyzed in a three-tiered approach under widely standardized conditions. Firstly, RNA was extracted from PBMC of eight blood samples, obtained on two consecutive days within one particular episode, including two different consecutive depressive and two different consecutive manic episodes, and submitted to (1) screening by microarray hybridizations, followed by (2) detailed bioinformatic analysis, and (3) confirmation of episode-specific regulation of genes by quantitative real-time polymerase chain reaction (qRT-PCR).Secondly, results were validated in additional blood samples obtained one to two years later. Among gene transcripts elevated in depressed episodes were prostaglandin D synthetase (PTGDS) and prostaglandin D2 11-ketoreductase (AKR1C3), both involved in hibernation. We hypothesized them to account for some of the rapid cycling symptoms. A subsequent treatment approach over 5 months applying the cyclooxygenase inhibitor celecoxib (2 x 200 mg daily) resulted in reduced severity rating of both depressed and manic episodes. This case suggests that rapid cycling is a systemic disease, resembling hibernation, with prostaglandins playing a mediator role.

Crystal structures of prostaglandin D(2) 11-ketoreductase (AKR1C3) in complex with the nonsteroidal anti-inflammatory drugs flufenamic acid and indomethacin.

It is becoming increasingly well established that nonsteroidal anti-inflammatory drugs (NSAID) protect against tumors of the gastrointestinal tract and that they may also protect against a variety of other tumors. These activities have been widely attributed to the inhibition of cylooxygenases (COX) and, in particular, COX-2. However, several observations have indicated that other targets may be involved. Besides targeting COX, certain NSAID also inhibit enzymes belonging to the aldo-keto reductase (AKR) family, including AKR1C3. We have demonstrated previously that overexpression of AKR1C3 acts to suppress cell differentiation and promote proliferation in myeloid cells. However, this enzyme has a broad tissue distribution and therefore represents a novel candidate for the target of the COX-independent antineoplastic actions of NSAID. Here we report on the X-ray crystal structures of AKR1C3 complexed with the NSAID indomethacin (1.8 A resolution) or flufenamic acid (1.7 A resolution). One molecule of indomethacin is bound in the active site, whereas flufenamic acid binds to both the active site and the beta-hairpin loop, at the opposite end of the central beta-barrel. Two other crystal structures (1.20 and 2.1 A resolution) show acetate bound in the active site occupying the proposed oxyanion hole. The data underline AKR1C3 as a COX-independent target for NSAID and will provide a structural basis for the future development of new cancer therapies with reduced COX-dependent side effects.

Identification of a principal mRNA species for human 3alpha-hydroxysteroid dehydrogenase isoform (AKR1C3) that exhibits high prostaglandin D2 11-ketoreductase activity.

Human 3alpha-hydroxysteroid dehydrogenase exists in four isoforms, which belong to the aldo-keto reductase (AKR) superfamily and are named AKR1C1-AKR1C4. The properties of AKR1C3 have not been fully characterized compared to the other isoforms. In addition, a cDNA that shows more than 99% homology with AKR1C3 cDNA has been cloned from human myeloblasts. We have here expressed and purified a recombinant enzyme (designated as DBDH) from this cDNA. DBDH oxidized xenobiotic alicyclic alcohols and 3alpha- or 17beta-hydroxy-5beta-androstanes, and catalyzed the reversible conversion between prostaglandin D2 and 9alpha,11beta-prostaglandin F2 more efficiently than that of 3alpha- or 17beta-hydroxysteroids:the respective Km values were 0.6 and 6.8 microM, and kcat/Km values were about 1,000 min-1.mM-1. Anti-inflammatory drugs highly inhibited the enzyme. The recombinant AKR1C3 prepared by site-directed mutagenesis of DBDH also showed the same properties as the wild-type DBDH. Analyses of expression of mRNAs for DBDH and AKR1C3 by reverse transcription-PCR indicated that only one mRNA species for DBDH is expressed in 33 human specimens of liver, kidney, lung, brain, heart, spleen, adrenal gland, small intestine, placenta, prostate, and testis. These results suggest that AKR1C3 acts as prostaglandin D2 11-ketoreductase, and that its principal gene in the human has a coding region represented by DBDH cDNA.

REDUCTASES FOR XENOBIOTIC CARBONYL COMPOUNDS. REDUCTASES FOR XENOBIOTIC CARBONYL COMPOUNDS. Structures and Specificities of the Human Liver Enzymes

Dihydrodiol dehydrogenase (DD) is involved in both the metabolic inactivation and intoxication of aromatic hydrocarbons. Human liver contains four DDs, DD1-DD4, of which DD3 was identified with aldehyde reductase. In order to further evaluate the nature of the other enzymes, we performed the molecular cloning of the two enzymes, DD2 and DD4, and examined the reactivity towards endogenous and xenobiotic compounds. cDNA cloning of the enzymes demonstrated significant sequence homology to members of the aldo-keto reductase family. The sequences of DD2 and DD4, having 82% identity, were virtually identical with those of human liver bile-acid binder and chlordecone reductase, respectively. The results of substrate specificity for endogenous compounds indicated that DD1, DD2 and DD4 act as 3(20)α or 3α-hydroxysteroid dehydrogenases, and suggested that DD1 and DD2 are also the major species of prostaglandin D2 11-ketoreductase in human liver cytosol. On the other hand, the DDs catalyzed the reduction of several drug ketones more efficiently than did aldehyde reductase and carbonyl reductase of human liver.

9 alpha,11 beta-prostaglandin F2 formation in various bovine tissues. Different isozymes of prostaglandin D2 11-ketoreductase, contribution of prostaglandin F synthetase and its cellular localization.

9 alpha,11 beta-prostaglandin F2 was formed from prostaglandin D2 by its 11-ketoreductases in 100,000 x g supernatants of various bovine tissues in the presence of an NADPH-generating system. The reductase activities were high in liver (51.09 nmol/h/mg of protein), lung (24.99), and spleen (14.20); moderate in heart and pancreas (3.09-3.61); weak in stomach, intestine, colon, kidney, uterus, adrenal gland, and thymus (0.11-2.63); and undetectable in brain, retina, carotid artery, and blood (less than 0.10). No formation of prostaglandin F2 alpha from prostaglandin D2 was detected in all tissues. In immunotitration analyses with a polyclonal antibody specific for prostaglandin F synthetase, the reductase activities in lung and spleen showed identical titration curves to that of the purified synthetase and decreased to less than 15% of the initial activity under the condition of antibody excess. Prostaglandin F synthetase-immunoreactive protein in these two tissues showed peptide fingerprints identical to that of the purified enzyme after partial digestion with Staphylococcus aureus V8 protease. The antibody was partially cross-reactive to the reductase in liver (about 20% of that to the synthetase) but not to the reductase(s) in other tissues. The Km value for prostaglandin D2 of the reductase activity was the same in lung and spleen as that of the purified prostaglandin F synthetase (120 microM) but differed in liver (6 microM), heart, and pancreas (15 microM). The predominant distribution of prostaglandin F synthetase in lung and spleen was confirmed by radioimmunoassay (2.8 and 1.0 micrograms/mg protein, respectively) and Northern blot analyses. In immunoperoxidase staining, this enzyme was localized in alveolar interstitial cells and nonciliated epithelial cells in lung, histiocytes and/or dendritic cells in spleen, and a few interstitial cells in kidney and adrenal cortex.