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Code:
PO
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Time Slot/Poster Number:
280
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Session:
Small Molecules and Drug Discovery, Poster
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Structure-based Investigation of Wnt Signaling Inhibitors
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| Jie Zheng
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St. Jude Children's Research Hospital, Memphis, TN
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| Abstract |
Wnt signaling plays crucial roles in embryonic development and in tissue maintenance in adults. Abnormal activation of Wnt signaling is observed in several types of cancers. Dishevelled (Dvl) is a key molecule in the Wnt pathways that, through its PDZ domain, relays Wnt signals from membrane-bound Wnt receptors to downstream components. We have worked to develop small-molecule inhibitors of the Dvl PDZ protein–protein interaction for use in the elucidation of biological processes and as potential cancer-treatment and prevention agents.
Through structure-based virtual ligand screening and NMR spectroscopy, we initially identified the first inhibitor of the Dvl PDZ domain from the NCI small-molecule database. Then, by virtually exploring the existing chemical space and the development of SAR models of PDZ ligands, we obtained addition ~70 small additional small-molecule inhibitors. We also show that some drug-like inhibitors to interact at low to sub micromolar affinity with the PDZ domain. In additional, we showed that one of the such compounds inhibited Wnt signaling and reduced the levels of apoptosis in the hyaloid vessels of eye. Moreover, this compound also suppressed the growth of prostate cancer PC-3 cells. These biological effects suggest that by blocking the PDZ domain of Dvl, the compound identified in our studies effectively inhibits the Wnt signaling and thus provides a useful tool for studies dissecting the Wnt signaling pathways.
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Code:
PO
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Time Slot/Poster Number:
281
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Session:
Small Molecules and Drug Discovery, Poster
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Characterization of a Hexapeptide with Antiviral Activity Against the SARS Corona Virus by Saturation Transfer Difference (STD) NMR Spectroscopy
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| Anna-Winona Struck1; Susanne Pfefferle2; Marco Axmann1; Christian Drosten3; Bernd Meyer1
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1University of Hamburg, Hamburg, Germany; 2Bernhard Nocht-Institute for Tropical Medicine, Hamburg, Germany; 3University Hospital Bonn, Bonn, Germany
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| Abstract |
A defined receptor binding domain (RBD) on the viral spike protein (S) mediates the attachment of SARS Corona Virus to its cellular receptor, Angiotensin Converting Enzyme 2 (ACE2). We have analyzed binding of the SARS S-protein with ACE2 by saturation transfer difference (STD) NMR spectroscopy and surface plasmon resonance (SPR). From a peptide library a hexapeptide from Tyr438 to Leu443, Tyr-Lys-Tyr-Arg-Tyr-Leu (YKYRYL), of S-protein was identified to have binding affinity to ACE2 (KD= 10 µM). This peptide has also strong antiviral activity and can suppress viral proliferation completely at a concentration of 10 mM. STD NMR spectroscopy was used to detect the interaction of YKYRYL and related peptides with the receptor protein ACE2.
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Code:
PO
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Time Slot/Poster Number:
282
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Session:
Small Molecules and Drug Discovery, Poster
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NMR Structure Elucidation of a CCK1 Receptor Agonist Series – Are They Mixtures?
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| Andrew Butler; Kimberly Cameron; Kathleen Farley; Robert Oliver
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Pfizer Global R&D, Groton, CT
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| Abstract |
The structure elucidation of a series of novel triazolobenzodiazepinone CCK1 receptor agonists, one of which has completed Phase IIA clinical trials for the treatment of obesity and management of glycemic control, using Nuclear Magnetic Resonance (NMR) spectroscopy, is described. In solution, most members of this series exhibit marked atropisomerism/rotamerism, complicating NMR spectra. We show convincing evidence that this is a series of single molecular entities most of which exist in two distinct inter-converting conformations in solution, observable by NMR. The solution-state NMR studies are complimented by corroborating crystallographic, chromatographic, and molecular modeling studies.
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Code:
PO
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Time Slot/Poster Number:
283
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Session:
Small Molecules and Drug Discovery, Poster
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Progress in the Isolation, Identification and Characterization of the Pristionchus pacificus Mating Pheromone
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| Arthur S Edison1; Hans Alborn2; Ramadan Ajredini1; Andrea Choe3; Tatsuji Chuman1; Chaevien Clendinen1; Aaron T Dossey1; Adrien Gauna1; Fatma Kaplan2; Hasan Rasheed1; Frank Schroeder4; Paul W Sternberg3
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1University of Florida, Gainesville, FL; 2USDA Laboratory, Gainesville, FL; 3HHMI and Caltech, Pasadena, CA; 4Boyce Thompson Institute, Cornell University, Ithica, NY
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| Abstract |
Pristionchus pacificus is related to the nematode, Caenorhabditis elegans. C. elegans produces several related small molecules called ascarosides that often act synergistically and produce multiple biological responses including male-specific attraction and dauer formation. Exudates of P. pacificus hermaphrodites are attractive to P. pacificus males but not to C. elegans males. We have collected liquid bacterial co-cultures as well as biologically active worm water samples of P. pacificus. We have used NMR and LC-MS to search for known C. elegans ascarosides in the P. pacificus co-cultures and worm water. We have detected 3 known ascarosides in bacterial co-cultures, but we have found little or no evidence for any of the known ascarosides in the worm water, suggesting an environmental dependence.
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Code:
PO
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Time Slot/Poster Number:
284
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Session:
Small Molecules and Drug Discovery, Poster
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Characterizing the Binding of Disopyramide Enantiomers to
Alpha-1 Acid Glycoprotein with Ligand-Detected NMR Experiments
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| Jennifer Cruz; Cynthia K. Larive
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UC Riverside, Riverside, CA
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| Abstract |
Specific ligand binding studies are integral to drug design and development. This information can be derived by using ligand detected NMR epitope mapping experiments such as nuclear Overhauser effect spectroscopy (NOESY) and saturation transfer difference (STD). Each of these experiments gives different information about ligand-protein interactions through measurements of dipolar cross-relaxation of ligand and protein protons. This presentation compares the results of the NOESY and STD experiments and analyzes their ability to differentiate the binding epitopes of disopyramide enantiomers with the human serum protein α1-acid glycoprotein (AGP).
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Code:
PO
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Time Slot/Poster Number:
285
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Session:
Small Molecules and Drug Discovery, Poster
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Structure Elucidation of Conopeptides by NMR
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| Marloes Schurink; Henry Hocking; Rolf Boelens
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Utrecht University, Utrecht, NETHERLANDS
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| Abstract |
Venomous animals represent a major source of highly selective and efficient bioactive compounds that have already led to the development of several new drugs. The CONCO project focuses on therapeutically relevant molecules from marine venomous cone snails. Each individual venom typically contains hundreds of different peptides and small proteins. Such conotoxin or conopeptide families are usually disulfide-constrained peptides, sharing a characteristic well-conserved cysteine framework. The non-cysteinyl residues are highly variable, leading to the extreme diversity of conopeptides. Structure elucidation using NMR combined with bioactivity assays will lead to a better understanding of their structure-function relationship. Eventually, optimization of conopeptides may lead to alternative anesthetics and new treatments against a wide range of diseases affecting the nervous system.
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Code:
PO
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Time Slot/Poster Number:
286
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Session:
Small Molecules and Drug Discovery, Poster
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Multidisciplinary NMR, Biophysical, X-Ray and Computational Techniques in Support of FBDD to Generate Novel Lead Series of BACE Inhibitors
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| Thomas O'connell; Kris Borzilleri; Michael Brodney; Tommy Chen; Ivan Efremov; Zachary Hendsch; Alexander McColl; Felix Vajdos; Hong Wang; Jane Withka
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Pfizer Global R&D, Groton, CT
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| Abstract |
Primary screening of a proprietary fragment library using NMR methods led to the discovery of a novel lead for BACE inhibitors in support of an Alzheimer's Disease drug discovery program. Fragment hits were confirmed and the active site binding mode was determined by backbone chemical shift perturbations obtained from TROSY based 1H-15N correlation methods. In an effort to guide chemistry optimization and drive SAR of fragments, a robust NMR based functional activity assay was developed to assess biochemical activity of weak lead compounds. In addition, Surface Plasmon Resonance, X-Ray crystal structures and computational modeling techniques along with knowledge from previous aspartyl protease drug discovery programs were used to efficiently expand this fragment to generate a novel series of BACE inhibitors.
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Code:
PO
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Time Slot/Poster Number:
287
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Session:
Small Molecules and Drug Discovery, Poster
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Use of Solid State NMR to Aid the Salt Selection Process for Pharmaceutical Products
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| Richard Lewis; David Martin; Anke Fiebig; Talbir Austin; Austen Pimm
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AstraZeneca R&D Charnwood, Loughborough, United Kingdom
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| Abstract |
Selection of a suitable drug compound for development involves not only selecting the right compound, but also the correct salt form with the most desirable properties such as solubility or stability. This work describes how we have used solid state and solution NMR to understand three issues occurring at the salt selection stage of a project. We show causes of variable stoichiometry, and a study on the protonation state of salts which showed a change in site of protonation between aqueous solution and the solid state. We will also describe a correlation of solid state mobility with the stability for different salts of a compound and how this might be used as a quick assessment of stability for that compound.
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Code:
PO
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Time Slot/Poster Number:
289
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Session:
Small Molecules and Drug Discovery, Poster
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Orientation of Steroids in the Active Site of Drug-metabolizing Mutants of the Cytochrome P450BM3: Comparison between Holo-enzyme and Heme Domain
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| Ard Kolkman
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Radboud University Nijmegen, Nijmegen, Netherlands
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| Abstract |
Steroids can be metabolized by Cytochrome P450BM3 mutants and generate potential new lead compounds. Based on these recently published results, we present here new methodologies for the biophysical characterization, by NMR and UV, of the binding of low-soluble steroids to mutant forms of the P450BM3 holo-enzyme and its heme domain alone. This includes, deriving the orientation of steroids in the active site of the enzyme by paramagnetic proton spin relaxation. The NMR-derived orientations of the steroids in the active site well explains their observed metabolization pattern and may provide ways for the rational design of new lead compounds.
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Code:
PO
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Time Slot/Poster Number:
290
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Session:
Small Molecules and Drug Discovery, Poster
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Automatic Identification and Retrieval of the Best Candidates for NMR Prediction Training
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| Mikhail Kvasha1; Aleksandr Sakharov1; Sergey Golotvin1; Ryan Sasaki2; Peter Maas3
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1Advanced Chemistry Development, Moscow, Russia; 2Advanced, Chemistry Development, Inc., Toronto, Canada; 3Specs, Kluyverweg 6, Netherlands
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| Abstract |
Presented in this work, is a tool that helps to intelligently build up prediction training databases to increase the accuracy of HOSE-code based predictions. The tool ranks all structures in a database according to their utility score. This utility score measures the prediction relevance a particular structure will have on 1H and 13C prediction. The top score is given to a structure that is able to train the largest number of atoms found in other structures to achieve high impact prediction training. A validation of this system, using an automated 1D 1H verification routine will be presented against several sets of structures from the Specs collection with varied degrees of similarity.
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Code:
PO
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Time Slot/Poster Number:
291
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Session:
Small Molecules and Drug Discovery, Poster
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Evaluation of the Benefit of Including COSY and HSQC 2D Data in Automated Structure Verification
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| Ryan R. Sasaki1; Sergey Golotvin2; Randy Rutkowske3; Timothy Spitzer3; Duncan Farrant4
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1ACD/Labs, Toronto, Canada; 2Advanced Chemistry Development, Moscow, Russia; 3GlaxoSmithKline, RTP, RTP, NC; 4GlaxoSmithKline UK, Stevenage, UK
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| Abstract |
In previous publications, the results of an automated structure verification process for 1D 1H NMR data as well as a combined approach using 1D 1H and 2D HSQC data have been presented. The inclusion of 2D HSQC data in the process has proven to provide a substantial improvement on overall pass-fail performance as well as a reduction in both false negative and false positive results. Here we evaluate the impact of 1H-1H homonuclear connectivities on the reliability of the structure verification and chemical shift assignments.
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Code:
PO
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Time Slot/Poster Number:
292
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Session:
Small Molecules and Drug Discovery, Poster
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Efficient Multi-Site Analysis of Optical Purity Using Cryoprobe-Assisted 13C-Detected NMR Spectroscopy
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| SiYi Wang; Istvan Pelczer
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Princeton University, Princeton, NJ
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| Abstract |
We propose using cryoprobe-optimized 13C-NMR to assess presence and relative quantity of diastereomers. A few milligrams sample of small molecules can provide a satisfactory spectrum for quantitative analysis in a short time. The 13C-NMR spectrum only has singlets, the dispersion and resolution of such spectra is very high, and differential relaxation is only a minimal problem. The singlet resonances can be easily integrated, possibly using curve fitting approaches to improve accuracy. In a single spectrum one can usually find many sites available for the same measurement of the relative integrals. The 13C-NMR approach is very tolerant to presence of impurities, to variable sample conditions, and works well also for very complex systems and when there are multiple diastereomers present.
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Code:
PO
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Time Slot/Poster Number:
293
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Session:
Small Molecules and Drug Discovery, Poster
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Evaluation of Polymorphs of Cephalosporins by 13C and 15N Solid State NMR
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| Daniel L.M. Aguiar1; Rosane A.S. San Gil1; Stefano Caldarelli3; Lea Lopes2
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1Instituto de Quimica, Universidade Federal do RJ, Rio de Janeiro, Brazil; 2IMA, Universidade Federal do RJ, Rio de Janeiro, Brazil; 3Spectropolle, Universite Marseille, Marseille, France
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| Abstract |
Cephalexin and cefadroxil are cephalosporins, antibiotics with antibactericidal activity and wide pharmaceutical use. In this communication the combination of C-13 and N-15 solid state NMR and thermal analysis (DSC and TGA) was employed to address the presence of polymorphs in commercial samples of cephalexin and cephadroxil.
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Code:
PO
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Time Slot/Poster Number:
294
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Session:
Small Molecules and Drug Discovery, Poster
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Microcoil NMR Characterization of Heparin Derived Oligosaccharides
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| Szabolcs Beni1, 2; John Limtiaco1; Christopher Jones1; Derek Langeslay1; Cynthia Larive1
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1UC Riverside, Riverside, CA; 2Semmelweis University, Budapest, Hungary
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| Abstract |
Microcoil NMR can be a powerful technique to characterize mass limited heparin oligosaccharides. The presented research used microcoil NMR to identify various size uniform oligosaccharide fractions of digested heparin. Microgram amounts of tetra- and hexasaccharides were isolated using both strong anion exchange and reverse-phase ion pairing chromatography. NMR-pH titrations were carried out to characterize carboxylate basicities as a function of oligosaccharide chain length.
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Code:
PO
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Time Slot/Poster Number:
295
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Session:
Small Molecules and Drug Discovery, Poster
|
NMR Monitoring of H-bond Network Perturbations in the Active Site of hMAGL: A Straightforward Approach for NMR Ligand Screening
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| Sergiy Tyukhtenko; Ioannis Karageorgos; Nikolai Zvonok; Vidyanand Shukla; Alexandros Makriyannis
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Northeastern University, Boston, MA
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| Abstract |
A new protein-based NMR screening method accelerating the discovery of high-affinity inhibitors of therapeutically important human monoacylglycerol lipase (MAGL) has been developed. Perturbations of low field 1D 1H NMR signals associated with a network of short hydrogen bonds in the active site of MAGL strongly correlated with affinity of ligands to enzyme. This method allows the identification of specific interactions with enzyme for compounds with broad range of affinities. The technique does not require protein isotopic labeling and sequential NMR resonance assignment.
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Code:
PO
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Time Slot/Poster Number:
296
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Session:
Small Molecules and Drug Discovery, Poster
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Database-assisted NMR Structure Elucidation of Small Molecules in Drug Discovery Research
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| Yingjie Li; Mark Mulvihill; Lifu Ma
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OSI Pharmaceuticals, Farmingdale, NY
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| Abstract |
NMR is considered a choice technique for structure elucidation of drug-like organic molecules and unknown byproducts such as drug metabolites and impurities. In order to enhance the accuracy and efficiency of structure characterization requested by our drug discovery laboratories, we have developed a comprehensive workflow where key tasks were sub-categorized as structure verification, isomer identification, and unknown structure elucidation. Application of our novel, undisclosed database in computer-aided structure elucidation has proven to significantly facilitate the identification of compounds that are commonly investigated by our chemists. In this poster three case studies will be presented to illustrate the powerful capability of our workflow and the benefit of fragment databases for significantly increasing the efficiency of structure elucidation.
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