- Molecular dynamics simulations of viral neuraminidase inhibitors with the human neuraminidase enzymes: Insights into isoenzyme selectivity
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Inhibitors of viral neuraminidase enzymes have been previously developed as therapeutics. Humans can express multiple forms of neuraminidase enzymes (NEU1, NEU2, NEU3, NEU4) that share a similar active site and enzymatic mechanism with their viral counterparts. Using a panel of purified human neuraminidase enzymes, we tested the inhibitory activity of 2-deoxy-2,3-dehydro-N-acetylneuraminic acid (DANA), zanamivir, oseltamivir, and peramivir against each of the human isoenzymes. We find that, with the exceptions of DANA and zanamivir, these compounds show generally poor activity against the human neuraminidase enzymes. To provide insight into the interactions of viral inhibitors with human neuraminidases, we conducted molecular dynamics simulations using homology models based on coordinates reported for NEU2. Simulations revealed that an organized water is displaced by zanamivir in binding to NEU2 and NEU3 and confirmed the critical importance of engaging the binding pocket of the C7–C9 glycerol sidechain. Our results suggest that compounds designed to target the human neuraminidases should provide more selective tools for interrogating these enzymes. Furthermore, they emphasize a need for additional structural data to enable structure-based drug design in these systems.
- Richards, Michele R.,Guo, Tianlin,Hunter, Carmanah D.,Cairo, Christopher W.
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- Design, in silico studies, synthesis and in?vitro evaluation of oseltamivir derivatives as inhibitors of neuraminidase from influenza A virus H1N1
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Since the neuraminidase (NA) enzyme of the influenza A virus plays a key role in the process of release of new viral particles from a host cell, it is often a target for new drug design. The emergence of NA mutations, such as H275Y, has led to great resis
- Neri-Bazán, Rocío M.,García-Machorro, Jazmín,Méndez-Luna, David,Tolentino-López, Luis E.,Martínez-Ramos, Federico,Padilla-Martínez, Itzia I.,Aguilar-Faisal, Leopoldo,Soriano-Ursúa, Marvin A.,Trujillo-Ferrara, José G.,Fragoso-Vázquez, M. Jonathan,Barrón, Blanca L.,Correa-Basurto, José
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Read Online
- In vitro inhibition of carboxylesterase 1 by major cannabinoids and selected metabolites
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The escalating use of medical cannabis and significant recreational use of cannabis in recent years has led to a higher potential for metabolic interactions between cannabis or one or more of its components and concurrently used medications. Although there have been a significant number of in vitro and in vivo assessments of the effects of cannabis on cytochrome P450 and UDP-glucuronosyltransferase enzyme systems, there is limited information regarding the effects of cannabis on the major hepatic esterase, carboxylesterase 1 (CES1). In this study, we investigated the in vitro inhibitory effects of the individual major cannabinoids and metabolites Δ9-tetrahydrocannabinol (THC), cannabidiol (CBD), cannabinol (CBN), 11-nor-THC-carboxylic acid, and 11-hydroxy-THC on CES1 activity. S9 fractions from human embryonic kidney 293 cells stably expressing CES1 were used in the assessment of cannabinoid inhibitory effects. THC, CBD, and CBN each exhibited substantial inhibitory potency, and were further studied to determine their mechanism of inhibition and kinetic parameters. The inhibition of CES1 by THC, CBD, and CBN was reversible and appears to proceed through a mixed competitive-noncompetitive mechanism. The inhibition constant (Ki) values for THC, CBD, and CBN inhibition were 0.541, 0.974, and 0.263 mM (0.170, 0.306, and 0.0817 mg/ml), respectively. Inhibition potency was increased when THC, CBD, and CBN were combined. Compared with the potential unbound plasma concentrations attainable clinically, the Ki values suggest a potential for clinically significant inhibition of CES1 by THC and CBD. CBN, however, is expected to have a limited impact on CES1. Carefully designed clinical studies are warranted to establish the clinical significance of these in vitro findings.
- Qian, Yuli,Wang, Xinwen,Markowitz, John S.
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- Development and validation of a stability-indicating spectrofluorimetric method for the determination of H1N1 antiviral drug (oseltamivir phosphate) in human plasma through the Hantzsch reaction
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A simple and sensitive spectrofluorimetric method has been described and validated for the determination of oseltamivir phosphate (OSP) in its pure form and pharmaceutical dosage forms. The method is based on the reaction of acetylacetone and formaldehyde with the primary amino group of OSP through the Hantzsch reaction, forming a yellow fluorescent dihydropyridine derivative measured spectrofluorimetrically at 475 nm (excitation at 408 nm). At the optimum reaction conditions, the fluorescence intensity concentration plot is rectilinear over the range of 0.4-2.8 μg mL-1. The lower limits of detection and quantitation were 0.08 and 0.24 μg mL-1, respectively. The developed method was successfully applied for determination of OSP in its commercial capsules and suspension with an average percentage recovery of 99.86 ± 1.20 and 98.36 ± 2.42, respectively (n = 5) without interference from common excipients. The proposed method was utilized for in vitro determination of the cited drug in spiked human plasma, with a percent mean recovery (n = 3) of 98.22 ± 1.27%. Furthermore, the proposed method was extended to study the stability of OSP under different stress conditions; such as hydrolysis (acidic and alkaline), oxidation (30% H2O2 v/v), photolysis, (as per ICH guideline Q1B option 1) and a thermal degradation study. Also the developed method was used to investigate the kinetics of the alkaline and acidic degradations of the cited drug.
- Omar, Mahmoud A.,Derayea, Sayed M.,Mostafa, Islam M.
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- Stereoisomers of oseltamivir-synthesis, in silico prediction and biological evaluation
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Oseltamivir is an important antiviral drug, which possess three chirality centers in its structure. From eight possible stereoisomers, only two have been synthesized and evaluated so far. We describe herein the stereoselective synthesis, computational activity prediction and biological testing of another three diastereoisomers of oseltamivir. These isomers have been synthesized using stereoselective organocatalytic Michael addition, cyclization and reduction. Their binding to viral neuraminidase N1 of influenza A virus was evaluated by quantum-chemical calculations and their anti-influenza activities were tested by an in vitro virus-inhibition assay. All three isomers displayed antiviral activity lower than that of oseltamivir, however, one of the stereoisomers, (3S,4R,5S)-isomer, of oseltamivir showed in vitro potency towards the Tamiflu-sensitive influenza viral strain A/Perth/265/2009(H5N1) comparable to Tamiflu.
- Hajzer, Viktória,Fi?era, Roman,Latika, Attila,Durmis, Július,Kollár, Jakub,Frecer, Vladimír,Tu?eková, Zuzana,Miertu?, Stanislav,Kostolansky, Franti?ek,Vare?ková, Eva,?ebesta, Radovan
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p. 1828 - 1841
(2017/03/09)
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- Novel compound, its synthetic method and therapeutic use (by machine translation)
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Novel compounds are described. The compounds generally comprise an acidic group, a basic group, a substituted amino or N-acyl and a group having an optionally hydroxylated alkane moiety. Pharmaceutical compositions comprising the inhibitors of the invention are also described. Methods of inhibiting neuraminidase in samples suspected of containing neuraminidase are also described. Antigenic materials, polymers, antibodies, conjugates of the compounds of the invention with labels, and assay methods for detecting neuraminidase activity are also described.
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Paragraph 2230-2241
(2016/10/07)
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- Kinetic, thermodynamic and structural analysis of tamiphosphor binding to neuraminidase of H1N1 (2009) pandemic influenza
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Influenza virus causes severe respiratory infections that are responsible for up to half a million deaths worldwide each year. Two inhibitors targeting viral neuraminidase have been approved to date (oseltamivir, zanamivir). However, the rapid development of antiviral drug resistance and the efficient transmission of resistant viruses among humans represent serious threats to public health. The approved influenza neuraminidase inhibitors have (oxa)cyclohexene scaffolds designed to mimic the oxonium transition state during enzymatic cleavage of sialic acid. Their active forms contain a carboxylate that interacts with three arginine residues in the enzyme active site. Recently, the phosphonate group was successfully used as an isostere of the carboxylate in oseltamivir, and the resulting compound, tamiphosphor, was identified as a highly active neuraminidase inhibitor. However, the structure of the complex of this promising inhibitor with neuraminidase has not yet been reported. Here, we analyzed the interaction of a set of oseltamivir and tamiphosphor derivatives with neuraminidase from the A/California/07/2009 (H1N1) influenza virus. We thermodynamically characterized the binding of oseltamivir carboxylate or tamiphosphor to the neuraminidase catalytic domain by protein microcalorimetry, and we determined crystal structure of the catalytic domain in complex with tamiphosphor at 1.8 ? resolution. This structural information should aid rational design of the next generation of neuraminidase inhibitors.
- Albina?a, Carlos Berenguer,MacHara, Ale?,Rezacov, Pavlína,Pachl, Petr,Konvalinka, Jan,Kozísek, Milan
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p. 100 - 109
(2016/06/09)
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- Oseltamivir analogues bearing N-substituted guanidines as potent neuraminidase inhibitors
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A series of oseltamivir analogues bearing an N-substituted guanidine unit were prepared and evaluated as inhibitors of neuraminidases from four strains of influenza the two most potent analogues identified contain relatively small N-guanidine substituents (N-methyl and N-hydroxyl) and display enhanced inhibition with IC50 values in the low nanomolar range against neuraminidases from wild-type and oseltamivir-resistant strains. Potential advantages of including the N-hydroxyguanidine moiety in neuraminidase inhibitors are also discussed.
- Mooney, Caitlin A.,Johnson, Stuart A.,'T Hart, Peter,Quarles Van Ufford, Linda,De Haan, Cornelis A. M.,Moret, Ed E.,Martin, Nathaniel I.
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p. 3154 - 3160
(2014/05/06)
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- Synthesis, isolation and characterization of process-related impurities in oseltamivir phosphate
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Three known impurities in oseltamivir phosphate bulk drug at level 0.1% (ranging from 0.05-0.1%) were detected by gradient reverse phase high performance liquid chromatography. These impurities were preliminarily identified by the mass number of the impur
- Sharma, Yogesh Kumar,Agarwal, Dau Dayal,Bhure, Sudesh,Rathore, Sanjay Singh,Rawat, Chakravir,Mukharjee, Rajendranath
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experimental part
p. 113 - 120
(2012/06/18)
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- Formal synthesis of tamiflu: Conversion of tamiflu into tamiphosphor
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A short, enantiomeric, synthesis of Shibasakis 3rd generation intermediate to form (-)-oseltamivir phosphate (Tamiflu) has been achieved in eight steps with the use of inexpensive starting materials. A formal synthetic route to convert tamiflu into tamiphosphor via Fangs tamiphosphor intermediate has been accomplished. Georg Thieme Verlag Stuttgart · New York.
- Gunasekera, Dinara S.
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experimental part
p. 573 - 576
(2012/04/17)
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- Preparation of human drug metabolites using fungal peroxygenases
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The synthesis of hydroxylated and O- or N-dealkylated human drug metabolites (HDMs) via selective monooxygenation remains a challenging task for synthetic organic chemists. Here we report that aromatic peroxygenases (APOs; EC 1.11.2.1) secreted by the agaric fungi Agrocybe aegerita and Coprinellus radians catalyzed the H2O2-dependent selective monooxygenation of diverse drugs, including acetanilide, dextrorphan, ibuprofen, naproxen, phenacetin, sildenafil and tolbutamide. Reactions included the hydroxylation of aromatic rings and aliphatic side chains, as well as O- and N-dealkylations and exhibited different regioselectivities depending on the particular APO used. At best, desired HDMs were obtained in yields greater than 80% and with isomeric purities up to 99%. Oxidations of tolbutamide, acetanilide and carbamazepine in the presence of H218O2 resulted in almost complete incorporation of 18O into the corresponding products, thus establishing that these reactions are peroxygenations. The deethylation of phenacetin-d1 showed an observed intramolecular deuterium isotope effect [(kH/kD) obs] of 3.1 ± 0.2, which is consistent with the existence of a cytochrome P450-like intermediate in the reaction cycle of APOs. Our results indicate that fungal peroxygenases may be useful biocatalytic tools to prepare pharmacologically relevant drug metabolites.
- Poraj-Kobielska, Marzena,Kinne, Matthias,Ullrich, Rene,Scheibner, Katrin,Kayser, Gernot,Hammel, Kenneth E.,Hofrichter, Martin
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experimental part
p. 789 - 796
(2012/07/14)
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- Sugar-based synthesis of tamiflu and its inhibitory effects on cell secretion
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Tamiflu is currently the most effective drug for the treatment of influenza, but the insufficient supply and side-effects of this drug demand urgent solutions. We present a practical synthesis of Tamiflu by using novel synthetic routes, cheap reagents, and the abundantly available starting material D-glucal. The strategy features a Claisen rearrangement of hexose to obtain the cyclohexene backbone and introduction of diamino groups through tandem intramolecular aziridination and ring opening. In addition, this synthetic protocol allows late-stage functionalization for the flexible synthesis of Tamiflu analogues. By using the synthesized Tamiflu and its active metabolite (oseltamivir carboxylate), we inves-tigated their influences on neuroendocrine PC12 cells in various aspects. It was discovered that oseltamivir carboxylate significantly inhibits the vesicular exocytosis (regulated secretion) of PC 12 cells, and suggests a mechanism underlying the Tamiflu side-effects, in particular its possible adverse influences on neurotransmitter release in the central nervous system.
- Ma, Jimei,Zhao, Yanying,Ng, Simon,Zhang, Jing,Zeng, Jing,Than, Aung,Chen, Peng,Liu, Xue-Wei
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supporting information; experimental part
p. 4533 - 4540
(2010/08/19)
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- SYNTHESIS OF OSELTAMIVIR CONTAINING PHOSPHONATE CONGENERS WITH ANTI-INFLUENZA ACTIVITY
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Novel phosphonate compounds are described. The compounds have activity as neuraminidase inhibitors against wild-type and H274Y mutant of H1N1 and H5N1 viruses. The present disclosure also provides an enantioselective synthetic route to known neuraminidase inhibitors oseltamivir and the anti-flu drug Tamiflu, as well as novel phosphonate compounds, via D-xylose. Another efficient and flexible synthesis of Tamiflu and the highly potent neuraminidase inhibitor Tamiphosphor was also achieved in 11 steps and > 20% overall yields from the readily available fermentation product (1S-cis)-3-bromo-3,5- cyclohexadiene-1,2-diol. Most of the reaction intermediates were obtained as crystals without tedious purification procedures. The key transformations include an initial regio- and stereoselective bromoamidation of a bromoarene cis- dihydrodiol, as well as the final palladium-catalyzed carbonylation and phosphonylation.
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Page/Page column 57
(2009/04/25)
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- Radiosyntheses of two positron emission tomography probes: [11C]Oseltamivir and its active metabolite [11C]Ro 64-0802
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Oseltamivir phosphate (Tamiflu, 1·H3PO4 is an orally active anti-influenza drug, which is hydrolyzed by esterase to its carboxylate metabolite Ro 64-0802 (2) with potent activity inhibiting neuraminidase. In this study, fo
- Konno, Fujiko,Arai, Takuya,Zhang, Ming-Rong,Hatori, Akiko,Yanamoto, Kazuhiko,Ogawa, Masanao,Ito, Gukuto,Odawara, Chika,Yamasaki, Tomoteru,Kato, Koichi,Suzuki, Kazutoshi
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p. 1260 - 1263
(2008/09/18)
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- Synthesis of tamiflu and its phosphonate congeners possessing potent anti-influenza activity
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Using d-xylose as an appropriate chiral precursor, we have synthesized active neuraminidase inhibitor oseltamivir, antiflu drug Tamiflu, and novel phosphonate congeners that exhibit even stronger antiflu activities by inhibiting the neuraminidases of the wild-type and H274Y mutant of H1N1 and H5N1 viruses. Molecular modeling of the neuraminidase-phosphonate complex indicates a pertinent binding mode of the phosphonate with three arginine residues in the active site. Discovery of such potent neuraminidase inhibitors will offer an opportunity to the development of new anti-influenza drugs. Copyright
- Shie, Jiun-Jie,Fang, Jim-Min,Wang, Shi-Yun,Tsai, Keng-Chang,Cheng, Yih-Shyun E.,Yang, An-Suei,Hsiao, Shih-Chia,Su, Ching-Yao,Wong, Chi-Huey
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p. 11892 - 11893
(2008/03/27)
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- Structure-activity relationship studies of novel carbocyclic influenza neuraminidase inhibitors
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A series of influenza neuraminidase inhibitors with the cyclohexene scaffold containing lipophilic side chains have been synthesized and evaluated for influenza A and B neuraminidase inhibitory activity. The size and geometry of side chains have been modified systematically in order to investigate structure-activity relationships of this class of compounds. The X-ray crystal structures of several analogues complexed with neuraminidase revealed that the lipophilic side chains bound to the hydrophobic pocket consisted of Glu276, Ala246, Arg224, and Ile222 of the enzyme active site. The structure-activity relationship studies of this series have also demonstrated remarkably different inhibitory potency between influenza A and B neuraminidase. This indicated that the lipophilic side chains had quite different hydrophobic interactions with influenza A and B neuraminidase despite their complete homology in the active site. Influenza B neuraminidase appeared to be much more sensitive toward the increased steric bulkiness of inhibitors compared to influenza A neuraminidase. From the extensive structure-activity relationship investigation reported in this article, GS 4071 emerged as one of the most potent influenza neuraminidase inhibitors against both influenza A and B strains.
- Kim, Choung U.,Lew, Willard,Williams, Matthew A.,Wu, Huiwei,Zhang, Lijun,Chen, Xiaowu,Escarpe, Paul A.,Mendel, Dirk B.,Laver, W. Graeme,Stevens, Raymond C.
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p. 2451 - 2460
(2007/10/03)
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- Biexponential decomposition of a neuraminidase inhibitor prodrug (GS- 4104) in aqueous solution
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Purpose. To examine the degradation kinetics and identify the degradation products of a neuraminidase inhibitor prodrug, GS-4104. Methods. Degradation was studied as a function of pH and temperature using a stability-indicating RP-HPLC assay. Degradation products were isolated by RP- HPLC and identified by NMR. Specific rate constants were calculated based on a scheme defined by product(s) analysis. Results. Three distinct degradation products were observed in the pH region studied (pH 2-8): Isomer I, GS-4071, and isomer II. Isomer I resulted from the N, N-migration of the acetyl group. GS-4071 was formed by the hydrolysis of the ethyl ester. Both GS-4071 and isomer I degraded further to isomer [I by N, N-acyl migration and ester hydrolysis, respectively. The N, N-acyl migration reaction was characterized using two dimensional heteronuclear multiple bond correlation (HMBC) NMR. The decomposition kinetics of GS-4104 follow a biexponential decay at pH 2-7. The degradation kinetics of GS-4104 at pH 4.0, 70°C were independent of the initial GS-4104 concentration. Conclusions. The degradation profile indicates that development of solution or solid dosage form of GS-4104 with adequate shelf-life stability at room temperature is feasible.
- Oliyai, Reza,Yuan, Lung-Chi,Dahl, Terrence C.,Swaminathan,Wang, Ke-Yu,Lee, William A.
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p. 1300 - 1304
(2007/10/03)
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- Influenza neuraminidase inhibitors possessing a novel hydrophobic interaction in the enzyme active site: Design, synthesis, and structural analysis of carbocyclic sialic acid analogues with potent anti-influenza activity
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The design, synthesis, and in vitro evaluation of the novel carbocycles as transition-state-based inhibitors of influenza neuraminidase (NA) are described. The double bond position in the carbocyclic analogues plays an important role in NA inhibition as d
- Kim, Choung U.,Lew, Willard,Williams, Matthew A.,Liu, Hongtao,Zhang, Lijun,Swaminathan,Bischofberger, Norbert,Chen, Ming S.,Mendel, Dirk B.,Tai, Chun Y.,Laver, W. Graeme,Stevens, Raymond C.
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p. 681 - 690
(2007/10/03)
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