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Find video protocols related to scientific articles indexed in Pubmed.
Antagonism of chemical genetic interaction networks resensitize MRSA to ?-lactam antibiotics.
Chem. Biol.
PUBLISHED: 06-17-2011
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Antibiotic drug resistance among hospital and community acquired methicillin resistant Staphylococcus aureus (MRSA) has dramatically eroded the efficacy of current therapeutics. We describe a chemical genetic strategy using antisense interference to broadly identify new drug targets that potentiate the effects of existing antibiotics against both etiological classes of MRSA infection. Further, we describe the resulting chemical genetic interaction networks and highlight the prominent and overlapping target sets that restore MRSA susceptibility to penicillin, cephalosporins, and carbapenems. Pharmacological validation of this approach is the potent synergy between a known inhibitor to a member of this genetic potentiation network (GlmS) and a broad set of ?-lactam antibiotics against methicillin resistant Staphylococci. Developing drug-like leads to these targets may serve as rational and effective combination agents when paired with existing ?-lactam antibiotics to restore their efficacy against MRSA.
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New indole amide derivatives as potent CRTH2 receptor antagonists.
Bioorg. Med. Chem. Lett.
PUBLISHED: 02-21-2011
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A new series of indole amide acting as hCRTH2 receptor ligands had been explored and are described herein. Several amide derivatives displaying low nanomolar activity in hCRTH2 binding and whole blood assays were identified. They were found to behave as a full antagonists, exhibiting good selectivity over related prostaglandin receptors. Also, prototypical compounds in this novel series which displayed acceptable CYP profiles and were orally bioavailable in rats were identified.
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Pathway analysis of Candida albicans survival and virulence determinants in a murine infection model.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 12-06-2010
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One potentially rich source of possible targets for antifungal therapy are those Candida albicans genes deemed essential for growth under the standard culture (i.e., in vitro) conditions; however, these genes are largely unexplored as drug targets because essential genes are not experimentally amenable to conventional gene deletion and virulence studies. Using tetracycline-regulatable promoter-based conditional mutants, we investigated a murine model of candidiasis in which repressing essential genes in the host was achieved. By adding doxycycline to the drinking water starting 3 days prior to (dox - 3D) or 2 days post (dox + 2D) infection, the phenotypic consequences of temporal gene inactivation were assessed by monitoring animal survival and fungal burden in prophylaxis and acute infection settings. Of 177 selected conditional shut-off strains tested, the virulence of 102 was blocked under both repressing conditions, suggesting that the corresponding genes are essential for growth and survival in a murine host across early and established infection periods. Among these genes were those previously identified as antifungal drug targets (i.e., FKS1, ERG1, and ERG11), verifying that this methodology can be used to validate potential new targets. We also identify genes either conditionally essential or dispensable for in vitro growth but required for survival and virulence, including those in late stage ergosterol synthesis, or early steps in fatty acid or riboflavin biosynthesis. This study evaluates the role of essential genes with respect to pathogen virulence in a large-scale, systems biology context, and provides a general method for gene target validation and for uncovering unexpected antimicrobial targets.
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Azaindoles as potent CRTH2 receptor antagonists.
Bioorg. Med. Chem. Lett.
PUBLISHED: 10-28-2010
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A new class of 7-azaindole analogs of MK-7246 as potent and selective CRTH2 antagonists is reported. The SAR leading to the identification of the optimal azaindole regioisomer as well as the pharmacokinetics and off-target activities of the most potent antagonists are disclosed.
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Discovery of MK-7246, a selective CRTH2 antagonist for the treatment of respiratory diseases.
Bioorg. Med. Chem. Lett.
PUBLISHED: 10-14-2010
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In this manuscript we wish to report the discovery of MK-7246 (4), a potent and selective CRTH2 (DP2) antagonist. SAR studies leading to MK-7246 along with two synthetic sequences enabling the preparation of this novel class of CRTH2 antagonist are reported. Finally, the pharmacokinetic and metabolic profile of MK-7246 is disclosed.
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Pharmacological characterization of MK-7246, a potent and selective CRTH2 (chemoattractant receptor-homologous molecule expressed on T-helper type 2 cells) antagonist.
Mol. Pharmacol.
PUBLISHED: 10-13-2010
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The chemoattractant receptor-homologous molecule expressed on T-helper type 2 cells (CRTH2) is a G protein-coupled receptor that has been reported to modulate inflammatory responses in various rodent models of asthma, allergic rhinitis and atopic dermatitis. In this study, we describe the biological and pharmacological properties of {(7R)-7-[[(4-fluorophenyl)sulfonyl](methyl)amino]-6,7,8,9-tetrahydropyrido[1,2-a]indol-10-yl}acetic acid (MK-7246), a novel synthetic CRTH2 antagonist. We show that MK-7246 1) has high affinity for the human, monkey, dog, rat, and mouse CRTH2, 2) interacts with CRTH2 in a reversible manner, 3) exhibits high selectivity over all prostanoid receptors as well as 157 other receptors and enzymes, 4) acts as a full antagonist on recombinant and endogenously expressed CRTH2, 5) demonstrates good oral bioavailability and metabolic stability in various animal species, 6) yields ex vivo blockade of CRTH2 on eosinophils in monkeys and sheep, and 7) significantly blocks antigen-induced late-phase bronchoconstriction and airway hyper-responsiveness in sheep. MK-7246 represents a potent and selective tool to further investigate the in vivo function of CRTH2.
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Staphylococcus aureus TargetArray: comprehensive differential essential gene expression as a mechanistic tool to profile antibacterials.
Antimicrob. Agents Chemother.
PUBLISHED: 06-14-2010
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The widespread emergence of antibiotic-resistant bacteria and a lack of new pharmaceutical development have catalyzed a need for new and innovative approaches for antibiotic drug discovery. One bottleneck in antibiotic discovery is the lack of a rapid and comprehensive method to identify compound mode of action (MOA). Since a hallmark of antibiotic action is as an inhibitor of essential cellular targets and processes, we identify a set of 308 essential genes in the clinically important pathogen Staphylococcus aureus. A total of 446 strains differentially expressing these genes were constructed in a comprehensive platform of sensitized and resistant strains. A subset of strains allows either target underexpression or target overexpression by heterologous promoter replacements with a suite of tetracycline-regulatable promoters. A further subset of 236 antisense RNA-expressing clones allows knockdown expression of cognate targets. Knockdown expression confers selective antibiotic hypersensitivity, while target overexpression confers resistance. The antisense strains were configured into a TargetArray in which pools of sensitized strains were challenged in fitness tests. A rapid detection method measures strain responses toward antibiotics. The TargetArray antibiotic fitness test results show mechanistically informative biological fingerprints that allow MOA elucidation.
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Chemical genetic profiling and characterization of small-molecule compounds that affect the biosynthesis of unsaturated fatty acids in Candida albicans.
J. Biol. Chem.
PUBLISHED: 06-01-2009
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The balance between saturated and unsaturated fatty acids plays a crucial role in determining the membrane fluidity. In the diploid fungal pathogen Candida albicans, the gene for fatty acid Delta9 desaturase, OLE1, is essential for viability. Using a reverse genetic approach, termed the fitness test, we identified a group of structurally related synthetic compounds that induce specific hypersensitivity of the OLE1(+/-) strain. Genetic repression of OLE1 and chemical inhibition by two selected compounds, ECC145 and ECC188, resulted in a marked decrease in the total unsaturated fatty acids and impaired hyphal development. The resulting auxotroph of both was suppressed by the exogenous monounsaturated fatty acids (16:1Delta9 and 18:1Delta9). These correlations suggest that both compounds affect the level of unsaturated fatty acids, likely by impairing Ole1p directly or indirectly. However, the residual levels of monounsaturated fatty acids (MUFAs) resulted from chemical inhibition were significantly higher than OLE1 repression, indicating even partial inhibition of MUFAs is sufficient to stop cellular proliferation. Although the essentiality of OLE1 was suppressed by MUFAs in vitro, we demonstrated that it was required for virulence in a murine model of systemic candidiasis even when the animals were supplemented with a high fat diet. Thus, the fungal fatty acid desaturase is an attractive antifungal drug target. Taking advantage of the inhibitors and the relevant conditional shut-off strains, we validated several chemical genetic interactions observed in the fitness test profiles that reveal novel genetic interactions between OLE1/unsaturated fatty acids and other cellular processes.
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Restoring methicillin-resistant Staphylococcus aureus susceptibility to ?-lactam antibiotics.
Sci Transl Med
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Despite the need for new antibiotics to treat drug-resistant bacteria, current clinical combinations are largely restricted to ?-lactam antibiotics paired with ?-lactamase inhibitors. We have adapted a Staphylococcus aureus antisense knockdown strategy to genetically identify the cell division Z ring components-FtsA, FtsZ, and FtsW-as ?-lactam susceptibility determinants of methicillin-resistant S. aureus (MRSA). We demonstrate that the FtsZ-specific inhibitor PC190723 acts synergistically with ?-lactam antibiotics in vitro and in vivo and that this combination is efficacious in a murine model of MRSA infection. Fluorescence microscopy localization studies reveal that synergy between these agents is likely to be elicited by the concomitant delocalization of their cognate drug targets (FtsZ and PBP2) in MRSA treated with PC190723. A 2.0 Å crystal structure of S. aureus FtsZ in complex with PC190723 identifies the compound binding site, which corresponds to the predominant location of mutations conferring resistance to PC190723 (PC190723(R)). Although structural studies suggested that these drug resistance mutations may be difficult to combat through chemical modification of PC190723, combining PC190723 with the ?-lactam antibiotic imipenem markedly reduced the spontaneous frequency of PC190723(R) mutants. Multiple MRSA PC190723(R) FtsZ mutants also displayed attenuated virulence and restored susceptibility to ?-lactam antibiotics in vitro and in a mouse model of imipenem efficacy. Collectively, these data support a target-based approach to rationally develop synergistic combination agents that mitigate drug resistance and effectively treat MRSA infections.
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What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

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We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

Video X seems to be unrelated to Abstract Y...

In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.