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Dive into the research topics where Paul S. Charifson is active.

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Featured researches published by Paul S. Charifson.


Proteins | 2004

A detailed comparison of current docking and scoring methods on systems of pharmaceutical relevance.

Emanuele Perola; W. Patrick Walters; Paul S. Charifson

A thorough evaluation of some of the most advanced docking and scoring methods currently available is described, and guidelines for the choice of an appropriate protocol for docking and virtual screening are defined. The generation of a large and highly curated test set of pharmaceutically relevant protein–ligand complexes with known binding affinities is described, and three highly regarded docking programs (Glide, GOLD, and ICM) are evaluated on the same set with respect to their ability to reproduce crystallographic binding orientations. Glide correctly identified the crystallographic pose within 2.0 Å in 61% of the cases, versus 48% for GOLD and 45% for ICM. In general Glide appears to perform most consistently with respect to diversity of binding sites and ligand flexibility, while the performance of ICM and GOLD is more binding site–dependent and it is significantly poorer when binding is predominantly driven by hydrophobic interactions. The results also show that energy minimization and reranking of the top N poses can be an effective means to overcome some of the limitations of a given docking function. The same docking programs are evaluated in conjunction with three different scoring functions for their ability to discriminate actives from inactives in virtual screening. The evaluation, performed on three different systems (HIV‐1 protease, IMPDH, and p38 MAP kinase), confirms that the relative performance of different docking and scoring methods is to some extent binding site–dependent. GlideScore appears to be an effective scoring function for database screening, with consistent performance across several types of binding sites, while ChemScore appears to be most useful in sterically demanding sites since it is more forgiving of repulsive interactions. Energy minimization of docked poses can significantly improve the enrichments in systems with sterically demanding binding sites. Overall Glide appears to be a safe general choice for docking, while the choice of the best scoring tool remains to a larger extent system‐dependent and should be evaluated on a case‐by‐case basis. Proteins 2004.


Chemistry & Biology | 2000

The structures of caspases-1, -3, -7 and -8 reveal the basis for substrate and inhibitor selectivity

Yunyi Wei; Ted Fox; Steve P Chambers; JoAnne Sintchak; Joyce T. Coll; Julian Golec; Lora Swenson; Keith P. Wilson; Paul S. Charifson

BACKGROUND Peptide inhibitors of caspases have helped define the role of these cysteine proteases in biology. Structural and biochemical characterization of the caspase enzymes may contribute to the development of new drugs for the treatment of caspase-mediated inflammation and apoptosis. RESULTS The crystal structure of the previously unpublished caspase-7 (Csp7; 2.35 A) bound to the reversible tetrapeptide aldehyde inhibitor acetyl-Asp-Glu-Val-Asp-CHO is compared with crystal structures of caspases-1 (2.3 A), -3 (2.2 A), and -8 (2.65 A) bound to the same inhibitor. Csp7 is a close homolog of caspase-3 (Csp3), and these two caspases possess some quarternary structural characteristics that support their unique role among the caspase family. However, although Csp3 and Csp7 are quite similar overall, they were found to have a significantly different substitution pattern of amino acids in and around the S4-binding site. CONCLUSIONS These structures span all three caspase subgroups, and provide a basis for inferring substrate and inhibitor binding, as well as selectivity for the entire caspase family. This information will influence the design of selective caspase inhibitors to further elucidate the role of caspases in biology and hopefully lead to the design of therapeutic agents to treat caspase-mediated diseases, such as rheumatoid arthritis, certain neurogenerative diseases and stroke.


Antimicrobial Agents and Chemotherapy | 2004

Crystal Structures of Escherichia coli Topoisomerase IV ParE Subunit (24 and 43 Kilodaltons): a Single Residue Dictates Differences in Novobiocin Potency against Topoisomerase IV and DNA Gyrase

Steven Bellon; Jonathan D. Parsons; Yunyi Wei; Koto Hayakawa; Lora Swenson; Paul S. Charifson; Judith A. Lippke; Robert Aldape; Christian H. Gross

ABSTRACT Topoisomerase IV and DNA gyrase are related bacterial type II topoisomerases that utilize the free energy from ATP hydrolysis to catalyze topological changes in the bacterial genome. The essential function of DNA gyrase is the introduction of negative DNA supercoils into the genome, whereas the essential function of topoisomerase IV is to decatenate daughter chromosomes following replication. Here, we report the crystal structures of a 43-kDa N-terminal fragment of Escherichia coli topoisomerase IV ParE subunit complexed with adenylyl-imidodiphosphate at 2.0-Å resolution and a 24-kDa N-terminal fragment of the ParE subunit complexed with novobiocin at 2.1-Å resolution. The solved ParE structures are strikingly similar to the known gyrase B (GyrB) subunit structures. We also identified single-position equivalent amino acid residues in ParE (M74) and in GyrB (I78) that, when exchanged, increased the potency of novobiocin against topoisomerase IV by nearly 20-fold (to 12 nM). The corresponding exchange in gyrase (I78 M) yielded a 20-fold decrease in the potency of novobiocin (to 1.0 μM). These data offer an explanation for the observation that novobiocin is significantly less potent against topoisomerase IV than against DNA gyrase. Additionally, the enzyme kinetic parameters were affected. In gyrase, the ATP Km increased ≈5-fold and the Vmax decreased ≈30%. In contrast, the topoisomerase IV ATP Km decreased by a factor of 6, and the Vmax increased ≈2-fold from the wild-type values. These data demonstrate that the ParE M74 and GyrB I78 side chains impart opposite effects on the enzymes substrate affinity and catalytic efficiency.


Journal of Pharmacology and Experimental Therapeutics | 2007

(S)-1-((S)-2-{[1-(4-Amino-3-chloro-phenyl)-methanoyl]-amino}-3,3-dimethyl-butanoyl)-pyrrolidine-2-carboxylic acid ((2R,3S)-2-ethoxy-5-oxo-tetrahydro-furan-3-yl)-amide (VX-765), an Orally Available Selective Interleukin (IL)-Converting Enzyme/Caspase-1 Inhibitor, Exhibits Potent Anti-Inflammatory Activities by Inhibiting the Release of IL-1β and IL-18

Woods Wannamaker; Robert J. Davies; Mark Namchuk; John Pollard; Pamella J. Ford; George Ku; Caroline Decker; Paul S. Charifson; Peter Weber; Ursula A. Germann; Keisuke Kuida; John C. R. Randle

(S)-1-((S)-2-{[1-(4-Amino-3-chloro-phenyl)-methanoyl]-amino}-3,3-dimethyl-butanoyl)-pyrrolidine-2-carboxylic acid ((2R,3S)-2-ethoxy-5-oxo-tetrahydro-furan-3-yl)-amide (VX-765) is an orally absorbed prodrug of (S)-3-({1-[(S)-1-((S)-2-{[1-(4-amino-3-chlorophenyl)-methanoyl]-amino}-3,3-dimethyl-butanoyl)-pyrrolidin-2yl]-methanoyl}-amino)-4-oxo-butyric acid (VRT-043198), a potent and selective inhibitor of interleukin-converting enzyme/caspase-1 subfamily caspases. VRT-043198 exhibits 100- to 10,000-fold selectivity against other caspase-3 and -6 to -9. The therapeutic potential of VX-765 was assessed by determining the effects of VRT-043198 on cytokine release by monocytes in vitro and of orally administered VX-765 in several animal models in vivo. In cultures of peripheral blood mononuclear cells and whole blood from healthy subjects stimulated with bacterial products, VRT-043198 inhibited the release of interleukin (IL)-1β and IL-18, but it had little effect on the release of several other cytokines, including IL-1α, tumor necrosis factor-α, IL-6 and IL-8. In contrast, VRT-043198 had little or no demonstrable activity in cellular models of apoptosis, and it did not affect the proliferation of activated primary T cells or T-cell lines. VX-765 was efficiently converted to VRT-043198 when administered orally to mice, and it inhibited lipopolysaccharide-induced cytokine secretion. In addition, VX-765 reduced disease severity and the expression of inflammatory mediators in models of rheumatoid arthritis and skin inflammation. These data suggest that VX-765 is a novel cytokine inhibitor useful for treatment of inflammatory diseases.


Journal of Pharmacology and Experimental Therapeutics | 2007

VX-765, an orally available selective interleukin converting enzyme (ICE)/caspase-1 inhibitor exhibits potent anti-inflammatory activities by inhibiting the release of IL-1b and IL-18

Woods Wannamaker; Robert J. Davies; Mark Namchuk; John Pollard; Pamella J. Ford; George Ku; Caroline Decker; Paul S. Charifson; Peter Weber; Ursula A. Germann; Keisuke Kuida; John C. R. Randle

(S)-1-((S)-2-{[1-(4-Amino-3-chloro-phenyl)-methanoyl]-amino}-3,3-dimethyl-butanoyl)-pyrrolidine-2-carboxylic acid ((2R,3S)-2-ethoxy-5-oxo-tetrahydro-furan-3-yl)-amide (VX-765) is an orally absorbed prodrug of (S)-3-({1-[(S)-1-((S)-2-{[1-(4-amino-3-chlorophenyl)-methanoyl]-amino}-3,3-dimethyl-butanoyl)-pyrrolidin-2yl]-methanoyl}-amino)-4-oxo-butyric acid (VRT-043198), a potent and selective inhibitor of interleukin-converting enzyme/caspase-1 subfamily caspases. VRT-043198 exhibits 100- to 10,000-fold selectivity against other caspase-3 and -6 to -9. The therapeutic potential of VX-765 was assessed by determining the effects of VRT-043198 on cytokine release by monocytes in vitro and of orally administered VX-765 in several animal models in vivo. In cultures of peripheral blood mononuclear cells and whole blood from healthy subjects stimulated with bacterial products, VRT-043198 inhibited the release of interleukin (IL)-1β and IL-18, but it had little effect on the release of several other cytokines, including IL-1α, tumor necrosis factor-α, IL-6 and IL-8. In contrast, VRT-043198 had little or no demonstrable activity in cellular models of apoptosis, and it did not affect the proliferation of activated primary T cells or T-cell lines. VX-765 was efficiently converted to VRT-043198 when administered orally to mice, and it inhibited lipopolysaccharide-induced cytokine secretion. In addition, VX-765 reduced disease severity and the expression of inflammatory mediators in models of rheumatoid arthritis and skin inflammation. These data suggest that VX-765 is a novel cytokine inhibitor useful for treatment of inflammatory diseases.


Antimicrobial Agents and Chemotherapy | 2007

Dual Targeting of GyrB and ParE by a Novel Aminobenzimidazole Class of Antibacterial Compounds

Trudy H. Grossman; Douglas J. Bartels; Steve Mullin; Christian H. Gross; Jonathan D. Parsons; Yusheng Liao; Anne-Laure Grillot; Dean Stamos; Eric R. Olson; Paul S. Charifson; Nagraj Mani

ABSTRACT A structure-guided drug design approach was used to optimize a novel series of aminobenzimidazoles that inhibit the essential ATPase activities of bacterial DNA gyrase and topoisomerase IV and that show potent activities against a variety of bacterial pathogens. Two such compounds, VRT-125853 and VRT-752586, were characterized for their target specificities and preferences in bacteria. In metabolite incorporation assays, VRT-125853 inhibited both DNA and RNA synthesis but had little effect on protein synthesis. Both compounds inhibited the maintenance of negative supercoils in plasmid DNA in Escherichia coli at the MIC. Sequencing of DNA corresponding to the GyrB and ParE ATP-binding regions in VRT-125853- and VRT-752586-resistant mutants revealed that their primary target in Staphylococcus aureus and Haemophilus influenzae was GyrB, whereas in Streptococcus pneumoniae it was ParE. In Enterococcus faecalis, the primary target of VRT-125853 was ParE, whereas for VRT-752586 it was GyrB. DNA transformation experiments with H. influenzae and S. aureus proved that the mutations observed in gyrB resulted in decreased susceptibilities to both compounds. Novobiocin resistance-conferring mutations in S. aureus, H. influenzae, and S. pneumoniae were found in gyrB, and these mutants showed little or no cross-resistance to VRT-125853 or VRT-752586 and vice versa. Furthermore, gyrB and parE double mutations increased the MICs of VRT-125853 and VRT-752586 significantly, providing evidence of dual targeting. Spontaneous frequencies of resistance to VRT-752586 were below detectable levels (<5.2 × 10−10) for wild-type E. faecalis but were significantly elevated for strains containing single and double target-based mutations, demonstrating that dual targeting confers low levels of resistance emergence and the maintenance of susceptibility in vitro.


Antimicrobial Agents and Chemotherapy | 2015

Preclinical Activity of VX-787, a First-in-Class, Orally Bioavailable Inhibitor of the Influenza Virus Polymerase PB2 Subunit

Randal Byrn; Steven M. Jones; Hamilton Bennett; Chris M Bral; Michael P. Clark; Marc Jacobs; Ann D. Kwong; Mark Ledeboer; Joshua R. Leeman; Colleen F. McNeil; Mark A. Murcko; Azin Nezami; Emanuele Perola; Rene Rijnbrand; Kumkum Saxena; Alice W. Tsai; Yi Zhou; Paul S. Charifson

ABSTRACT VX-787 is a novel inhibitor of influenza virus replication that blocks the PB2 cap-snatching activity of the influenza viral polymerase complex. Viral genetics and X-ray crystallography studies provide support for the idea that VX-787 occupies the 7-methyl GTP (m7GTP) cap-binding site of PB2. VX-787 binds the cap-binding domain of the PB2 subunit with a KD (dissociation constant) of 24 nM as determined by isothermal titration calorimetry (ITC). The cell-based EC50 (the concentration of compound that ensures 50% cell viability of an uninfected control) for VX-787 is 1.6 nM in a cytopathic effect (CPE) assay, with a similar EC50 in a viral RNA replication assay. VX-787 is active against a diverse panel of influenza A virus strains, including H1N1pdm09 and H5N1 strains, as well as strains with reduced susceptibility to neuraminidase inhibitors (NAIs). VX-787 was highly efficacious in both prophylaxis and treatment models of mouse influenza and was superior to the neuraminidase inhibitor, oseltamivir, including in delayed-start-to-treat experiments, with 100% survival at up to 96 h postinfection and partial survival in groups where the initiation of therapy was delayed up to 120 h postinfection. At different doses, VX-787 showed a 1-log to >5-log reduction in viral load (relative to vehicle controls) in mouse lungs. Overall, these favorable findings validate the PB2 subunit of the viral polymerase as a drug target for influenza therapy and support the continued development of VX-787 as a novel antiviral agent for the treatment of influenza infection.


Antimicrobial Agents and Chemotherapy | 2006

In vitro characterization of the antibacterial spectrum of novel bacterial type II topoisomerase inhibitors of the aminobenzimidazole class.

Nagraj Mani; Christian H. Gross; Jonathan D. Parsons; Brian Hanzelka; Ute Müh; Steve Mullin; Yusheng Liao; Anne-Laure Grillot; Dean Stamos; Paul S. Charifson; Trudy H. Grossman

ABSTRACT Antibiotics with novel mechanisms of action are becoming increasingly important in the battle against bacterial resistance to all currently used classes of antibiotics. Bacterial DNA gyrase and topoisomerase IV (topoIV) are the familiar targets of fluoroquinolone and coumarin antibiotics. Here we present the characterization of two members of a new class of synthetic bacterial topoII ATPase inhibitors: VRT-125853 and VRT-752586. These aminobenzimidazole compounds were potent inhibitors of both DNA gyrase and topoIV and had excellent antibacterial activities against a wide spectrum of problematic pathogens responsible for both nosocomial and community-acquired infections, including staphylococci, streptococci, enterococci, and mycobacteria. Consistent with the novelty of their structures and mechanisms of action, antibacterial potency was unaffected by commonly encountered resistance phenotypes, including fluoroquinolone resistance. In time-kill assays, VRT-125853 and VRT-752586 were bactericidal against Staphylococcus aureus, Streptococcus pneumoniae, Enterococcus faecalis, and Haemophilus influenzae, causing 3-log reductions in viable cells within 24 h. Finally, similar to the fluoroquinolones, relatively low frequencies of spontaneous resistance to VRT-125853 and VRT-752586 were found, a property consistent with their in vitro dual-targeting activities.


Antimicrobial Agents and Chemotherapy | 2003

Active-Site Residues of Escherichia coli DNA Gyrase Required in Coupling ATP Hydrolysis to DNA Supercoiling and Amino Acid Substitutions Leading to Novobiocin Resistance

Christian H. Gross; Jonathan D. Parsons; Trudy H. Grossman; Paul S. Charifson; Steven Bellon; James Jernee; Maureen Dwyer; Stephen P. Chambers; William Markland; Martyn Botfield; Scott A. Raybuck

ABSTRACT DNA gyrase is a bacterial type II topoisomerase which couples the free energy of ATP hydrolysis to the introduction of negative supercoils into DNA. Amino acids in proximity to bound nonhydrolyzable ATP analog (AMP · PNP) or novobiocin in the gyrase B (GyrB) subunit crystal structures were examined for their roles in enzyme function and novobiocin resistance by site-directed mutagenesis. Purified Escherichia coli GyrB mutant proteins were complexed with the gyrase A subunit to form the functional A2B2 gyrase enzyme. Mutant proteins with alanine substitutions at residues E42, N46, E50, D73, R76, G77, and I78 had reduced or no detectable ATPase activity, indicating a role for these residues in ATP hydrolysis. Interestingly, GyrB proteins with P79A and K103A substitutions retained significant levels of ATPase activity yet demonstrated no DNA supercoiling activity, even with 40-fold more enzyme than the wild-type enzyme, suggesting that these amino acid side chains have a role in the coupling of the two activities. All enzymes relaxed supercoiled DNA to the same extent as the wild-type enzyme did, implying that only ATP-dependent reactions were affected. Mutant genes were examined in vivo for their abilities to complement a temperature-sensitive E. coli gyrB mutant, and the activities correlated well with the in vitro activities. We show that the known R136 novobiocin resistance mutations bestow a significant loss of inhibitor potency in the ATPase assay. Four new residues (D73, G77, I78, and T165) that, when changed to the appropriate amino acid, result in both significant levels of novobiocin resistance and maintain in vivo function were identified in E. coli.


Bioorganic & Medicinal Chemistry Letters | 2010

Discovery of pyrazolthiazoles as novel and potent inhibitors of bacterial gyrase.

Steven Ronkin; Michael C. Badia; Steve Bellon; Anne-Laure Grillot; Christian H. Gross; Trudy H. Grossman; Nagraj Mani; Jonathan D. Parsons; Dean Stamos; Martin Trudeau; Yunyi Wei; Paul S. Charifson

Bacterial DNA gyrase is an attractive target for the investigation of new antibacterial agents. Inhibitors of the GyrB subunit, which contains the ATP-binding site, are described in this communication. Novel, substituted 5-(1H-pyrazol-3-yl)thiazole compounds were identified as inhibitors of bacterial gyrase. Structure-guided optimization led to greater enzymatic potency and moderate antibacterial potency. Data are presented for the demonstration of selective enzyme inhibition of Escherichia coli GyrB over Staphylococcus aureus GyrB.

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Dean Stamos

Vertex Pharmaceuticals

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Wenxin Gu

Vertex Pharmaceuticals

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