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1.  Novel cGMP efflux inhibitors – Identified by virtual ligand screening (VLS) and confirmed by experimental studies 
Journal of medicinal chemistry  2012;55(7):3049-3057.
Elevated intracellular levels of cyclic guanosine monophosphate (cGMP) may induce apoptosis, and at least some cancer cells seem to escape this effect by increased efflux of cGMP, as clinical studies have shown that extracellular cGMP levels are elevated in various types of cancer. The human ATP binding cassette (ABC) transporter ABCC5 transports cGMP out of cells, and inhibition of ABCC5 may have cytotoxic effects. Sildenafil inhibits cGMP efflux by binding to ABCC5, and in order to search for potential novel ABCC5 inhibitors, we have identified sildenafil derivates using structural and computational guidance and tested them for the cGMP efflux effect. Eleven compounds from virtual ligand screening (VLS) were tested in vitro, using inside-out vesicles (IOV), for inhibition of cGMP efflux. 7 of 11 compounds predicted by VLS to bind to ABCC5 were more potent than sildenafil, and the two most potent showed Ki-values of 50 #x02013;100 nM.
PMCID: PMC4181661  PMID: 22380603
2.  Small Molecule Inhibitors of Bacillus anthracis Protective Antigen Proteolytic Activation and Oligomerization 
Journal of medicinal chemistry  2012;55(18):7998-8006.
Protective antigen (PA), lethal factor, and edema factor, the protein toxins of Bacillus anthracis, are among its most important virulence factors and play a key role in infection. We performed a virtual ligand screen of a library of 10,000 members to identify compounds predicted to bind to PA and prevent its oligomerization. Four of these compounds slowed PA association in a FRET-based oligomerization assay, and two of those protected cells from intoxication at concentrations of 1–10 μM. Exploration of the protective mechanism by Western blot showed decreased SDS-resistant PA oligomer on cells, and surprisingly, decreased amounts of activated PA. In vitro assays showed that one of the inhibitors blocked furin-mediated cleavage of PA, apparently through its binding to the PA substrate. Thus, we have identified inhibitors that can independently block both PA’s cleavage by furin and its subsequent oligomerization. Lead optimization on these two backbones may yield compounds with high activity and specificity for the anthrax toxins.
PMCID: PMC3474531  PMID: 22954387
3.  Modeling of the Aryl Hydrocarbon Receptor (AhR) ligand binding domain and its utility in virtual ligand screening to predict new AhR ligands 
Journal of Medicinal Chemistry  2009;52(18):5635-5641.
The Aryl Hydrocarbon Receptor (AhR) is a ligand-activated transcription factor; the AhR Per-AhR/Arnt-Sim (PAS) domain binds ligands. We developed homology models of the AhR PAS domain to characterize previously observed intra- and inter-species differences in ligand binding using Molecular Docking. In silico structure-based virtual ligand screening using our model resulted in the identification of pinocembrin and 5-hydroxy-7-methoxyflavone, which promoted nuclear translocation and transcriptional activation of AhR and AhR-dependent induction of endogenous target genes.
PMCID: PMC3289977  PMID: 19719119
4.  Vaccinia Virus Virulence Factor N1L is a Novel Promising Target for Antiviral Therapeutic Intervention 
Journal of medicinal chemistry  2010;53(10):3899-3906.
The 14 kDa homodimeric N1L protein is a potent vaccinia and variola (smallpox) virulence factor. It is not essential for viral replication, but it causes a strong attenuation of viral production in culture when deleted. The N1L protein is predicted to contain the BH3-like binding domain characteristic of Bcl-2 family proteins, and it is able to bind the BH3 peptides. Its overexpression has been reported to prevent infected cells from committing apoptosis. Therefore, interfering with the N1L apoptotic blockade may be a legitimate therapeutic strategy affecting the viral growth. By using in silico ligand docking and an array of in vitro assays, we have identified sub-micromolar (600 nM) N1L antagonists, belonging to the family of polyphenols. Their affinity is comparable to that of the BH3 peptides (70 nM ÷ 1000 nM). We have also identified the natural polyphenol resveratrol as a moderate N1L inhibitor. Finally, we show that our ligands efficiently inhibit growth of vaccinia virus.
PMCID: PMC2874095  PMID: 20441222
5.  Structure-Based Discovery of Novel Chemotypes for Adenosine A2A Receptor Antagonists 
Journal of medicinal chemistry  2010;53(4):1799-1809.
The recent progress in crystallography of G-protein coupled receptors opens an unprecedented venue for structure-based GPCR drug discovery. To test efficiency of the structure-based approach, we performed molecular docking and virtual ligand screening (VLS) of more than 4 million commercially available “drug-like” and ‘‘lead-like’’ compounds against the A2AAR 2.6 Å resolution crystal structure. Out of 56 high ranking compounds tested in A2AAR binding assays, 23 showed affinities under 10 µM, eleven of those had sub-µM affinities, and two compounds had affinities under 60 nM. The identified hits represent at least 9 different chemical scaffolds and are characterized by very high ligand efficiency (0.3–0.5 kcal/mol per heavy atom). Significant A2AAR antagonist activities were confirmed for 10 out of 13 ligands tested in functional assays. High success rate, novelty and diversity of the chemical scaffolds and strong ligand efficiency of the A2AAR antagonists identified in this study suggest practical applicability of receptor-based VLS in GPCR drug discovery.
PMCID: PMC2826142  PMID: 20095623
6.  Type-II Kinase Inhibitor Docking, Screening, and Profiling Using Modified Structures of Active Kinase States 
Journal of medicinal chemistry  2008;51(24):7921-7932.
Type-II kinase inhibitors represent a class of chemicals that trap their target kinases in an inactive, so-called DFG-out, state, occupying a hydrophobic pocket adjacent to the ATP binding site. These compounds are often more specific than those targeting active, DFG-in, kinase conformations. Unfortunately, the discovery of novel type-II scaffolds presents a considerable challenge, partly because the lack of compatible kinase structures makes structure-based methods inapplicable. We present a computational protocol for converting multiple available DFG-in structures of various kinases (∼70% of mammalian structural kinome) into accurate and specific models of their type-II-bound state. The models, described as Deletion-Of-Loop asp-PHe-gly-IN (DOLPHIN) kinase models, demonstrate exceptional performance in various inhibitor discovery applications, including compound pose prediction, screening, and in silico activity profiling. Given the abundance of the DFG-in structures, the presented approach opens possibilities for kinome-wide discovery of specific molecules targeting inactive kinase states.
PMCID: PMC2669721  PMID: 19053777
kinase; DFG-in; DFG-out; type-II inhibitor; imatinib; structure-based inhibitor discovery; compound screening; compound profiling; phosphorylation; cancer therapeutics
7.  Small molecules block the polymerisation of Z α1-antitrypsin and increase the clearance of intracellular aggregates 
Journal of medicinal chemistry  2007;50(22):5357-5363.
The Z mutant of α1-antitrypsin (Glu342Lys) causes a domain-swap and the formation of intrahepatic polymers that aggregate as inclusions and predispose the homozygote to cirrhosis. We have identified an allosteric cavity that is distinct from the interface involved in polymerisation for rational structure-based drug design to block polymer formation. Virtual ligand screening was performed on 1.2 million small molecules and 6 compounds were identified that reduced polymer formation in vitro. Modelling the effects of ligand binding on the cavity and re-screening the library identified an additional 10 compounds that completely blocked polymerisation. The best antagonists were effective at ratios of compound to Z α1-antitrypsin of 2.5:1 and reduced the intracellular accumulation of Z α1-antitrypsin by 70% in a cell model of disease. Identifying small molecules provides a novel therapy for the treatment of liver disease associated with the Z allele of α1-antitrypsin.
PMCID: PMC2631427  PMID: 17918823
hepatic inclusions; cirrhosis; serpinopathies; drug design; serpins
8.  De novo Discovery of Serotonin N-acetyltransferase Inhibitors 
Journal of medicinal chemistry  2007;50(22):5330-5338.
Serotonin N-acetyltransferase (arylalkylamine N-acetyltransferase, AANAT) is a member of the GCN5 N-acetyltransferase (GNAT) superfamily and catalyzes the penultimate step in the biosynthesis of melatonin; a large daily rhythm in AANAT activity drives the daily rhythm in circulating melatonin. We have used a structure-based computational approach to identify the first drug-like and selective inhibitors of AANAT. Approximately 1.2 million compounds were virtually screened by 3D high-throughput docking into the active site of X-ray structures for AANAT and in total 241 compounds were tested as inhibitors. One compound class, containing a rhodanine scaffold, exhibited low micromolar competitive inhibition against acetyl-CoA (AcCoA), and proved effective in blocking melatonin production in pineal cells. Compounds from this class are predicted to bind as bisubstrate inhibitors through interactions with the AcCoA and serotonin binding sites. Overall, this study demonstrates the feasibility of using virtual screening (VS) to identify small molecules which are selective inhibitors of AANAT.
PMCID: PMC2531295  PMID: 17924613

Results 1-8 (8)