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1.
J Comput Aided Mol Des ; 35(3): 297-314, 2021 03.
Article in English | MEDLINE | ID: mdl-33615401

ABSTRACT

Leishmaniasis is an infectious disease caused by parasites of the genus Leishmania and transmitted by the bite of a sand fly. To date, most available drugs for treatment are toxic and beyond the economic means of those affected by the disease. Protein disulfide isomerase (PDI) is a chaperone protein that plays a major role in the folding of newly synthesized proteins, specifically assisting in disulfide bond formation, breakage, or rearrangement in all non-native proteins. In previous work, we demonstrated that Leishmania major PDI (LmPDI) has an essential role in pathogen virulence. Furthermore, inhibition of LmPDI further blocked parasite infection in macrophages. In this study, we utilized a computer-aided approach to design a series of LmPDI inhibitors. Fragment-based virtual screening allowed for the understanding of the inhibitors' modes of action on LmPDI active sites. The generated compounds obtained after multiple rounds of virtual screening were synthesized and significantly inhibited target LmPDI reductase activity and were shown to decrease in vitro parasite growth in human monocyte-derived macrophages. This novel cheminformatics and synthetic approach led to the identification of a new series of compounds that might be optimized into novel drugs, likely more specific and less toxic for the treatment of leishmaniasis.


Subject(s)
Anti-Infective Agents/chemical synthesis , Enzyme Inhibitors/chemistry , Hexachlorophene/chemical synthesis , Leishmania major/enzymology , Leishmaniasis/drug therapy , Protein Disulfide-Isomerases/antagonists & inhibitors , Small Molecule Libraries/chemical synthesis , Anti-Infective Agents/pharmacology , Catalytic Domain , Computer-Aided Design , Drug Design , Enzyme Inhibitors/pharmacology , Hexachlorophene/pharmacology , Humans , Molecular Docking Simulation , Protein Binding , Protein Conformation , Small Molecule Libraries/pharmacology , Structure-Activity Relationship
2.
Int J Mol Sci ; 21(22)2020 Nov 20.
Article in English | MEDLINE | ID: mdl-33233525

ABSTRACT

We investigated the synthesis of N-docosahexaenoylethanolamine (synaptamide) in neuronal cells from unesterified docosahexaenoic acid (DHA) or DHA-lysophosphatidylcholine (DHA-lysoPC), the two major lipid forms that deliver DHA to the brain, in order to understand the formation of this neurotrophic and neuroprotective metabolite of DHA in the brain. Both substrates were taken up in Neuro2A cells and metabolized to N-docosahexaenoylphosphatidylethanolamine (NDoPE) and synaptamide in a time- and concentration-dependent manner, but unesterified DHA was 1.5 to 2.4 times more effective than DHA-lysoPC at equimolar concentrations. The plasmalogen NDoPE (pNDoPE) amounted more than 80% of NDoPE produced from DHA or DHA-lysoPC, with 16-carbon-pNDoPE being the most abundant species. Inhibition of N-acylphosphatidylethanolamine-phospholipase D (NAPE-PLD) by hexachlorophene or bithionol significantly decreased the synaptamide production, indicating that synaptamide synthesis is mediated at least in part via NDoPE hydrolysis. NDoPE formation occurred much more rapidly than synaptamide production, indicating a precursor-product relationship. Although NDoPE is an intermediate for synaptamide biosynthesis, only about 1% of newly synthesized NDoPE was converted to synaptamide, possibly suggesting additional biological function of NDoPE, particularly for pNDoPE, which is the major form of NDoPE produced.


Subject(s)
Arachidonic Acids/biosynthesis , Docosahexaenoic Acids/metabolism , Endocannabinoids/biosynthesis , Ethanolamines/metabolism , Lysophosphatidylcholines/metabolism , Neurons/metabolism , Animals , Arachidonic Acids/antagonists & inhibitors , Arachidonic Acids/isolation & purification , Bithionol/pharmacology , Carbon Isotopes , Cell Line, Tumor , Chromatography, Liquid , Endocannabinoids/antagonists & inhibitors , Endocannabinoids/isolation & purification , Ethanolamines/antagonists & inhibitors , Ethanolamines/isolation & purification , Hexachlorophene/pharmacology , Kinetics , Mice , Neurons/cytology , Neurons/drug effects , Plasmalogens/antagonists & inhibitors , Plasmalogens/biosynthesis , Plasmalogens/isolation & purification , Polyunsaturated Alkamides/antagonists & inhibitors , Polyunsaturated Alkamides/isolation & purification , Tandem Mass Spectrometry
3.
J Biol Chem ; 295(21): 7289-7300, 2020 05 22.
Article in English | MEDLINE | ID: mdl-32284327

ABSTRACT

N-Acyl-phosphatidylethanolamine phospholipase D (NAPE-PLD) (EC 3.1.4.4) catalyzes the final step in the biosynthesis of N-acyl-ethanolamides. Reduced NAPE-PLD expression and activity may contribute to obesity and inflammation, but a lack of effective NAPE-PLD inhibitors has been a major obstacle to elucidating the role of NAPE-PLD and N-acyl-ethanolamide biosynthesis in these processes. The endogenous bile acid lithocholic acid (LCA) inhibits NAPE-PLD activity (with an IC50 of 68 µm), but LCA is also a highly potent ligand for TGR5 (EC50 0.52 µm). Recently, the first selective small-molecule inhibitor of NAPE-PLD, ARN19874, has been reported (having an IC50 of 34 µm). To identify more potent inhibitors of NAPE-PLD, here we used a quenched fluorescent NAPE analog, PED-A1, as a substrate for recombinant mouse Nape-pld to screen a panel of bile acids and a library of experimental compounds (the Spectrum Collection). Muricholic acids and several other bile acids inhibited Nape-pld with potency similar to that of LCA. We identified 14 potent Nape-pld inhibitors in the Spectrum Collection, with the two most potent (IC50 = ∼2 µm) being symmetrically substituted dichlorophenes, i.e. hexachlorophene and bithionol. Structure-activity relationship assays using additional substituted dichlorophenes identified key moieties needed for Nape-pld inhibition. Both hexachlorophene and bithionol exhibited significant selectivity for Nape-pld compared with nontarget lipase activities such as Streptomyces chromofuscus PLD or serum lipase. Both also effectively inhibited NAPE-PLD activity in cultured HEK293 cells. We conclude that symmetrically substituted dichlorophenes potently inhibit NAPE-PLD in cultured cells and have significant selectivity for NAPE-PLD versus other tissue-associated lipases.


Subject(s)
Dichlorophen , Enzyme Inhibitors , Phospholipase D , Animals , Bacterial Proteins/antagonists & inhibitors , Bacterial Proteins/chemistry , Bacterial Proteins/metabolism , Bithionol/chemistry , Bithionol/pharmacology , Dichlorophen/chemistry , Dichlorophen/pharmacology , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , HEK293 Cells , Hexachlorophene/chemistry , Hexachlorophene/pharmacology , Humans , Mice , Phospholipase D/antagonists & inhibitors , Phospholipase D/chemistry , Phospholipase D/metabolism , Quinazolines/chemistry , Quinazolines/pharmacology , Streptomyces/enzymology , Sulfonamides/chemistry , Sulfonamides/pharmacology
4.
J Vet Med Sci ; 81(9): 1368-1372, 2019 Oct 10.
Article in English | MEDLINE | ID: mdl-31447458

ABSTRACT

In a previous study, we showed that 2', 3'-Cyclic Nucleotide 3'-Phosphodiesterase (CNPase) expression is induced in different temporal patterns in the cerebrum, cerebellum and medulla oblongata of hexachlorophene (HCP) and cuprizone (CPZ) treated rats. Here, we additionally examined the histopathological changes and CNPase expression in the spinal cord to clarify the reproducibility of different temporal patterns of CNPase expression in the spinal cord showing low degree or lack of spongy changes. Spongy changes were observed in HCP-treated rats, but not in CPZ-treated rats. Immunohistochemistry showed that intense expression of CNPase was not induced following HCP or CPZ treatment. Our data reveal that expression intensity of CNPase may be dependent on the degree of HCP- and CPZ-induced damage of the myelin sheath.


Subject(s)
2',3'-Cyclic-Nucleotide Phosphodiesterases/metabolism , Cuprizone/pharmacology , Hexachlorophene/pharmacology , Spinal Cord/drug effects , Animals , Female , Male , Myelin Sheath/drug effects , Myelin Sheath/pathology , Rats , Spinal Cord/growth & development , Spinal Cord/pathology , Time Factors
5.
Sci Rep ; 9(1): 6841, 2019 05 02.
Article in English | MEDLINE | ID: mdl-31048740

ABSTRACT

Chronic hepatitis viral infection, alcoholic intoxication, and obesity cause liver fibrosis, which progresses to decompensated liver cirrhosis, a disease for which medical demands cannot be met. Since there are currently no approved anti-fibrotic therapies for established liver fibrosis, the development of novel modalities is required to improve patient prognosis. In this study, we clarified the anti-fibrotic effects of cell sheets produced from human bone marrow-derived mesenchymal stem cells (MSCs) incubated on a temperature-sensitive culture dish with the chemical compound IC-2. Orthotopic transplantation of IC-2-engineered MSC sheets (IC-2 sheets) remarkably reduced liver fibrosis induced by chronic CCl4 administration. Further, the marked production of fibrolytic enzymes such as matrix metalloproteinase (MMP)-1 and MMP-14, as well as thioredoxin, which suppresses hepatic stellate cell activation, was observed in IC-2 sheets. Moreover, the anti-fibrotic effect of IC-2 sheets was much better than that of MSC sheets. Finally, knockdown experiments revealed that MMP-14 was primarily responsible for the reduction of liver fibrosis. Here, we show that IC-2 sheets could be a promising therapeutic option for established liver fibrosis.


Subject(s)
Bone Marrow Cells/cytology , Enzyme Inhibitors/pharmacology , Liver Cirrhosis/therapy , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/drug effects , Animals , Bridged Bicyclo Compounds/pharmacology , Chemical and Drug Induced Liver Injury/metabolism , Chemical and Drug Induced Liver Injury/therapy , Collagen Type I/metabolism , Hexachlorophene/pharmacology , Humans , Immunohistochemistry , Liver Cirrhosis/metabolism , Matrix Metalloproteinase 1/metabolism , Matrix Metalloproteinase 14/metabolism , Mesenchymal Stem Cells/metabolism , Mice , Mice, Inbred BALB C , Temperature
6.
Bioorg Med Chem Lett ; 29(14): 1689-1693, 2019 07 15.
Article in English | MEDLINE | ID: mdl-31129054

ABSTRACT

Glucose-regulated protein 78 (GRP78) is the ER resident 70 kDa heat shock protein 70 (HSP70) and has been hypothesized to be a therapeutic target for various forms of cancer due to its role in mitigating proteotoxic stress in the ER, its elevated expression in some cancers, and the correlation between high levels for GRP78 and a poor prognosis. Herein we report the development and use of a high throughput fluorescence polarization-based peptide binding assay as an initial step toward the discovery and development of GRP78 inhibitors. This assay was used in a pilot screen to discover the anti-infective agent, hexachlorophene, as an inhibitor of GRP78. Through biochemical characterization we show that hexachlorophene is a competitive inhibitor of the GRP78-peptide interaction. Biological investigations showed that this molecule induces the unfolded protein response, induces autophagy, and leads to apoptosis in a colon carcinoma cell model, which is known to be sensitive to GRP78 inhibition.


Subject(s)
HSP70 Heat-Shock Proteins/drug effects , Hexachlorophene/therapeutic use , High-Throughput Screening Assays/methods , Endoplasmic Reticulum Chaperone BiP , Hexachlorophene/pharmacology , Humans
7.
Neurotoxicology ; 56: 225-232, 2016 09.
Article in English | MEDLINE | ID: mdl-27555423

ABSTRACT

Longitudinal MRI employing diffusion tensor imaging and T2 mapping approaches has been applied to investigate the mechanisms of white matter damage caused by acute hexachlorophene neurotoxicity in rats in vivo. Male Sprague-Dawley rats were administered hexachlorophene orally once a day for five consecutive days at a dose of 30mg/kg and were monitored in 7T MRI scanner at days 0 (baseline), 3, 6, 13, and 20 following the first hexachlorophene dose. Quantitative T2 maps as well as a number of diffusion tensor parameters (fractional anisotropy, radial and axial diffusivity, apparent diffusion coefficient, and trace) were calculated from corresponding MR images. T2, as well as all diffusion tensor derived parameters (except fractional anisotropy) showed significant changes during the course of neurotoxicity development. These changes peaked at 6days after the first dose of hexachlorophene (one day after the last dose) and recovered to practically baseline levels at the end of observation (20days from the first dose). While such changes in diffusivity and T2 relaxation clearly demonstrate myelin perturbations consistent with edema, the lack of changes of fractional anisotropy suggests that the structure of the myelin sheath was not disrupted significantly by hexachlorophene in this study. This is also confirmed by the rapid recovery of all observed MRI parameters after cessation of hexachlorophene exposure.


Subject(s)
Anti-Infective Agents, Local/pharmacology , Brain/drug effects , Brain/diagnostic imaging , Diffusion Tensor Imaging , Hexachlorophene/pharmacology , Animals , Anisotropy , Image Processing, Computer-Assisted , Longitudinal Studies , Male , Nerve Fibers, Myelinated/drug effects , Rats , Rats, Sprague-Dawley , Time Factors , White Matter/diagnostic imaging , White Matter/drug effects
8.
Food Microbiol ; 60: 1-12, 2016 Dec.
Article in English | MEDLINE | ID: mdl-27554140

ABSTRACT

Lactic acid bacteria (LAB) isolated throughout the fermentation process of Aloreña table olives were found to be resistant at least to three antibiotics (Casado Muñoz et al., 2014); however, most were sensitive to the biocides tested in this study (with minimum inhibitory concentrations [MIC] below the epidemiological cut-off values). 2-15% of the isolates were found to be biocide resistant: Leuconostoc Pseudomesenteroides, which were resistant to hexachlorophene, and Lactobacillus pentosus to cetrimide and hexadecylpiridinium. We analyzed the effect of different physico-chemical stresses, including antimicrobials, on the phenotypic and genotypic responses of LAB, providing new insights on how they become resistant in a changing environment. Results indicated that similar phenotypic responses were obtained under three stress conditions: antimicrobials, chemicals and UV light. Susceptibility patterns to antibiotics changed: increasing MICs for ampicillin, chloramphenicol, ciprofloxacin, teicoplanin and tetracycline, and decreasing the MICs for clindamycin, erythromycin, streptomycin and trimethoprim in most strains. Statistically, cross resistance between different antibiotics was detected in all stress conditions. However, expression profiles of selected genes involved in stress/resistance response (rpsL, recA, uvrB and srtA) differed depending on the stress parameter, LAB species and strain, and the target gene. We conclude that, despite the uniform phenotypic response to stresses, the repertoire of induced and repressed genes differs. So, a search for a target to improve stress tolerance of LAB, especially those of importance as starter/protective cultures or probiotics, may depend on the individual screening of each strain, even though we could predict the antibiotic phenotypic response to all stresses.


Subject(s)
Anti-Infective Agents, Local/pharmacology , Disinfectants/pharmacology , Fermentation , Lactobacillaceae/drug effects , Lactobacillaceae/genetics , Olea/microbiology , Aminoacyltransferases/genetics , Bacterial Proteins/genetics , Cetrimonium , Cetrimonium Compounds/pharmacology , Cetylpyridinium/pharmacology , Cysteine Endopeptidases/genetics , Food Microbiology , Hexachlorophene/pharmacology , Lactobacillaceae/isolation & purification , Lactobacillaceae/radiation effects , Lactobacillus/drug effects , Lactobacillus/genetics , Lactobacillus/isolation & purification , Leuconostoc/drug effects , Leuconostoc/isolation & purification , Microbial Sensitivity Tests , Phenotype , Probiotics/isolation & purification , Probiotics/metabolism , Real-Time Polymerase Chain Reaction , Rec A Recombinases/genetics , Stress, Physiological/genetics
9.
Br J Pharmacol ; 173(8): 1339-49, 2016 Apr.
Article in English | MEDLINE | ID: mdl-26774021

ABSTRACT

BACKGROUND AND PURPOSE: High-throughput screening of compound libraries using genetically encoded fluorescent biosensors has identified several second-generation. low MW inhibitors of the calcium-activated chloride channel anoctamin 1 (CaCC/Ano1). Here we have (i) examined the effects of these Ano1 inhibitors on gastric and intestinal pacemaker activity; (ii) compared the effects of these inhibitors with those of the more classical CaCC inhibitor, 5-nitro-2-(3-phenylpropylalanine) benzoate (NPPB); (ii) examined the mode of action of these compounds on the waveform of pacemaker activity; and (iii) compared differences in the sensitivity between gastric and intestinal pacemaker activity to the Ano1 inhibitors. EXPERIMENTAL APPROACH: Using intracellular microelectrode recordings of gastric and intestinal muscle preparations from C57BL/6 mice, the dose-dependent effects of Ano1 inhibitors were examined on spontaneous electrical slow waves. KEY RESULTS: The efficacy of second-generation Ano1 inhibitors on gastric and intestinal pacemaker activity differed significantly. Antral slow waves were more sensitive to these inhibitors than intestinal slow waves. CaCCinh -A01 and benzbromarone were the most potent at inhibiting slow waves in both muscle preparations and more potent than NPPB. Dichlorophene and hexachlorophene were equally potent at inhibiting slow waves. Surprisingly, slow waves were relatively insensitive to T16Ainh -A01 in both preparations. CONCLUSIONS AND IMPLICATIONS: We have identified several second-generation Ano1 inhibitors, blocking gastric and intestinal pacemaker activity. Different sensitivities to Ano1 inhibitors between stomach and intestine suggest the possibility of different splice variants in these two organs or the involvement of other conductances in the generation and propagation of pacemaker activity in these tissues.


Subject(s)
Benzbromarone/pharmacology , Chloride Channels/antagonists & inhibitors , Dichlorophen/pharmacology , Gastrointestinal Tract/drug effects , Hexachlorophene/pharmacology , Thiophenes/pharmacology , Animals , Anoctamin-1 , Benzbromarone/chemistry , Chloride Channels/metabolism , Dichlorophen/chemistry , Dose-Response Relationship, Drug , Gastrointestinal Tract/metabolism , Hexachlorophene/chemistry , High-Throughput Screening Assays , Mice , Mice, Inbred C57BL , Structure-Activity Relationship , Thiophenes/chemistry
10.
Sci Rep ; 5: 16169, 2015 Nov 10.
Article in English | MEDLINE | ID: mdl-26553591

ABSTRACT

Mesenchymal stem cells (MSCs) are an attractive cell source for cell therapy. Based on our hypothesis that suppression of Wnt/ß-catenin signal enhances hepatic differentiation of human MSCs, we developed human mesenchymal stem cell-engineered hepatic cell sheets by a small molecule compound. Screening of 10 small molecule compounds was performed by WST assay, TCF reporter assay, and albumin mRNA expression. Consequently, hexachlorophene suppressed TCF reporter activity in time- and concentration-dependent manner. Hexachlorophene rapidly induced hepatic differentiation of human MSCs judging from expression of liver-specific genes and proteins, PAS staining, and urea production. The effect of orthotopic transplantation of human mesenchymal stem cell-engineered hepatic cell sheets against acute liver injury was examined in one-layered to three-layered cell sheets system. Transplantation of human mesenchymal stem cell-engineered hepatic cell sheets enhanced liver regeneration and suppressed liver injury. The survival rates of the mice were significantly improved. High expression of complement C3 and its downstream signals including C5a, NF-κB, and IL-6/STAT-3 pathway was observed in hepatic cell sheets-grafted tissues. Expression of phosphorylated EGFR and thioredoxin is enhanced, resulting in reduction of oxidative stress. These findings suggest that orthotopic transplantation of hepatic cell sheets manufactured from MSCs accelerates liver regeneration through complement C3, EGFR and thioredoxin.


Subject(s)
Hepatocytes/transplantation , Liver Regeneration/physiology , Mesenchymal Stem Cells/cytology , Animals , Cell Differentiation/drug effects , Cells, Cultured , Chemical and Drug Induced Liver Injury/mortality , Chemical and Drug Induced Liver Injury/therapy , Complement C3/metabolism , Complement C5a/metabolism , Cytokines/metabolism , ErbB Receptors/metabolism , Hepatocytes/cytology , Hepatocytes/metabolism , Hexachlorophene/pharmacology , Humans , Interleukin-6/metabolism , Mesenchymal Stem Cells/drug effects , Mice , Mice, Inbred NOD , Mice, SCID , NF-kappa B/metabolism , Oxidative Stress/drug effects , Phosphorylation/drug effects , STAT3 Transcription Factor/metabolism , Survival Rate , Thioredoxins/metabolism , Wnt Signaling Pathway/drug effects
11.
PLoS One ; 10(7): e0132816, 2015.
Article in English | MEDLINE | ID: mdl-26201074

ABSTRACT

Biocides are used without restriction for several purposes. As a consequence, large amounts of biocides are released without any control in the environment, a situation that can challenge the microbial population dynamics, including selection of antibiotic resistant bacteria. Previous work has shown that triclosan selects Stenotrophomonas maltophilia antibiotic resistant mutants overexpressing the efflux pump SmeDEF and induces expression of this pump triggering transient low-level resistance. In the present work we analyze if two other common biocides, benzalkonium chloride and hexachlorophene, trigger antibiotic resistance in S. maltophilia. Bioinformatic and biochemical methods showed that benzalkonium chloride and hexachlorophene bind the repressor of smeDEF, SmeT. Only benzalkonium chloride triggers expression of smeD and its effect in transient antibiotic resistance is minor. None of the hexachlorophene-selected mutants was antibiotic resistant. Two benzalkonium chloride resistant mutants presented reduced susceptibility to antibiotics and were impaired in growth. Metabolic profiling showed they were more proficient than their parental strain in the use of some dipeptides. We can then conclude that although bioinformatic predictions and biochemical studies suggest that both hexachlorophene and benzalkonium chloride should induce smeDEF expression leading to transient S. maltophilia resistance to antibiotics, phenotypic assays showed this not to be true. The facts that hexachlorophene resistant mutants are not antibiotic resistant and that the benzalkonium chloride resistant mutants presenting altered susceptibility to antibiotics were impaired in growth suggests that the risk for the selection (and fixation) of S. maltophilia antibiotic resistant mutants by these biocides is likely low, at least in the absence of constant selection pressure.


Subject(s)
Bacterial Proteins/metabolism , Disinfectants/pharmacology , Drug Resistance, Bacterial , Stenotrophomonas maltophilia/growth & development , Bacterial Proteins/chemistry , Bacterial Proteins/genetics , Benzalkonium Compounds/pharmacology , Computational Biology/methods , Drug Resistance, Bacterial/drug effects , Hexachlorophene/pharmacology , Models, Molecular , Molecular Docking Simulation , Stenotrophomonas maltophilia/drug effects , Stenotrophomonas maltophilia/enzymology
12.
J Biotechnol ; 207: 34-8, 2015 Aug 10.
Article in English | MEDLINE | ID: mdl-25987361

ABSTRACT

TAR DNA binding protein (TDP43) is a DNA- and RNA-binding protein that is implicated in several neurodegenerative disorders termed as "TDP43 proteinopathies" including Alzheimer's disease (AD), amyotrophic lateral sclerosis (ALS) and fronto-temporal lobe dementia (FTLD). We have developed an InCell Western (ICW) technique for screening TDP targeting drugs in 96 well plates. We tested 281 compounds and identified a novel compound hexachlorophene (referred to as B10) that showed potent reduction in TDP43 levels. The effect of B10 on TDP protein level was validated in two different cellular models: endogenous TDP43 expressing N9 microglial cells and TDP43-over-expressing HEK293 and HeLa cells. We also analyzed effect of B10 on various pathological forms of TDP such as the C25 cleaved fragment that localizes to the cytosol, insoluble high molecular weight species, and ALS-linked mutants. Our data suggest that B10 effectively reduces all forms of TDP. Overall, our data suggest that B10 could serve as a potential drug molecule for the treatment of AD, ALS and other TDP43 proteinopathies.


Subject(s)
DNA-Binding Proteins/antagonists & inhibitors , Drug Discovery/methods , Hexachlorophene/pharmacology , Animals , Cell Line , DNA-Binding Proteins/metabolism , HEK293 Cells , HeLa Cells , Hexachlorophene/chemistry , Humans , Mice , Molecular Targeted Therapy , TDP-43 Proteinopathies/drug therapy
13.
Antiviral Res ; 114: 1-10, 2015 Feb.
Article in English | MEDLINE | ID: mdl-25451075

ABSTRACT

With the recent emergence of Middle East Respiratory Syndrome coronavirus in humans and the outbreak of devastating porcine epidemic diarrhea coronavirus in swine, therapeutic intervention is urgently needed. However, anti-coronavirus drugs currently are not available. In an effort to assist rapid development of anti-coronavirus drugs, here we screened the NIH Clinical Collection in cell culture using a luciferase reporter-expressing recombinant murine coronavirus. Of the 727 compounds screened, 84 were found to have a significant anti-coronavirus effect. Further experiments revealed that 51 compounds blocked virus entry while 19 others inhibited viral replication. Additional validation studies with the top 3 inhibitors (hexachlorophene, nitazoxanide and homoharringtonine) demonstrated robust anti-coronavirus activities (a reduction of 6 to 8log10 in virus titer) with an IC50 ranging from 11nM to 1.2µM. Furthermore, homoharringtonine and hexachlorophene exhibited broad antiviral activity against diverse species of human and animal coronaviruses. Since the NIH Clinical Collection consists of compounds that have already been through clinical trials, these small molecule inhibitors have a great potential for rapid development as anti-coronavirus drugs.


Subject(s)
Antiviral Agents/pharmacology , Coronavirus/drug effects , Drug Discovery , Murine hepatitis virus/drug effects , Small Molecule Libraries , Animals , Cell Line , Coronavirus/physiology , Harringtonines/pharmacology , Hexachlorophene/pharmacology , Homoharringtonine , Humans , Luciferases/genetics , Mice , Nitro Compounds , Thiazoles/pharmacology , Viral Load/drug effects , Virus Internalization/drug effects , Virus Replication/drug effects
14.
Neurobiol Dis ; 74: 144-57, 2015 Feb.
Article in English | MEDLINE | ID: mdl-25173807

ABSTRACT

Aß accumulation plays a central role in the pathogenesis of Alzheimer's disease (AD). Recent studies suggest that the process of Aß nucleated polymerization is essential for Aß fibril formation, pathology spreading and toxicity. Therefore, targeting this process represents an effective therapeutic strategy to slow or block disease progression. To discover compounds that might interfere with the Aß seeding capacity, toxicity and pathology spreading, we screened a focused library of FDA-approved drugs in vitro using a seeding polymerization assay and identified small molecule inhibitors that specifically interfered with Aß seeding-mediated fibril growth and toxicity. Mitoxantrone, bithionol and hexachlorophene were found to be the strongest inhibitors of fibril growth and protected primary cortical neuronal cultures against Aß-induced toxicity. Next, we assessed the effects of these three inhibitors in vivo in the mThy1-APPtg mouse model of AD (8-month-old mice). We found that mitoxantrone and bithionol, but not hexachlorophene, stabilized diffuse amyloid plaques, reduced the levels of Aß42 oligomers and ameliorated synapse loss, neuronal damage and astrogliosis. Together, our findings suggest that targeting fibril growth and Aß seeding capacity constitutes a viable and effective strategy for protecting against neurodegeneration and disease progression in AD.


Subject(s)
Alzheimer Disease/drug therapy , Alzheimer Disease/physiopathology , Amyloid beta-Peptides/drug effects , Nerve Degeneration/drug therapy , Nerve Degeneration/physiopathology , Neuroprotective Agents/pharmacology , Peptide Fragments/drug effects , Alzheimer Disease/pathology , Amyloid beta-Peptides/toxicity , Amyloid beta-Protein Precursor/genetics , Amyloid beta-Protein Precursor/metabolism , Animals , Bithionol/pharmacokinetics , Bithionol/pharmacology , Cells, Cultured , Cerebral Cortex/drug effects , Cerebral Cortex/pathology , Cerebral Cortex/physiopathology , Disease Models, Animal , Drug Evaluation, Preclinical , Female , Gliosis/drug therapy , Gliosis/pathology , Gliosis/physiopathology , Hexachlorophene/pharmacokinetics , Hexachlorophene/pharmacology , Humans , Mice, Inbred C57BL , Mice, Transgenic , Mitoxantrone/pharmacokinetics , Mitoxantrone/pharmacology , Nerve Degeneration/pathology , Neurons/drug effects , Neurons/pathology , Neurons/physiology , Neuroprotective Agents/pharmacokinetics , Peptide Fragments/toxicity , Plaque, Amyloid/drug therapy , Plaque, Amyloid/pathology , Plaque, Amyloid/physiopathology , Rats
15.
Antimicrob Agents Chemother ; 59(1): 421-6, 2015 Jan.
Article in English | MEDLINE | ID: mdl-25367916

ABSTRACT

Mucosal biofilm-related fungal infections are very common, and the incidence of recurrent oral and vulvovaginal candidiasis is significant. As resistance to azoles (the preferred treatment) is occurring, we aimed at identifying compounds that increase the activity of miconazole against Candida albicans biofilms. We screened 1,600 compounds of a drug-repositioning library in combination with a subinhibitory concentration of miconazole. Synergy between the best identified potentiators and miconazole was characterized by checkerboard analyses and fractional inhibitory concentration indices. Hexachlorophene, pyrvinium pamoate, and artesunate act synergistically with miconazole in affecting C. albicans biofilms. Synergy was most pronounced for artesunate and structural homologues thereof. No synergistic effect could be observed between artesunate and fluconazole, caspofungin, or amphotericin B. Our data reveal enhancement of the antibiofilm activity of miconazole by artesunate, pointing to potential combination therapy consisting of miconazole and artesunate to treat C. albicans biofilm-related infections.


Subject(s)
Antifungal Agents/pharmacology , Artemisinins/pharmacology , Biofilms/drug effects , Candida albicans/drug effects , Miconazole/pharmacology , Amphotericin B/pharmacology , Artesunate , Candidiasis/drug therapy , Candidiasis/microbiology , Caspofungin , Drug Synergism , Echinocandins/pharmacology , Fluconazole/pharmacology , Hexachlorophene/pharmacology , Lipopeptides , Miconazole/therapeutic use , Microbial Sensitivity Tests , Pyrvinium Compounds/pharmacology , Reactive Oxygen Species/metabolism
16.
Chem Biol Interact ; 205(1): 53-62, 2013 Sep 05.
Article in English | MEDLINE | ID: mdl-23769903

ABSTRACT

OBJECTIVE: Glutathione transferase P1-1 (GST P1-1) is often overexpressed in tumor cells and is regarded as a contributor to their drug resistance. Inhibitors of GST P1-1 are expected to counteract drug resistance and may therefore serve as adjuvants in the chemotherapy of cancer by increasing the efficacy of cytostatic drugs. Finding useful inhibitors among compounds used for other indications would be a shortcut to clinical applications and a search for GST P1-1 inhibitors among approved drugs and other compounds was therefore conducted. METHODS: We tested 1040 FDA-approved compounds as inhibitors of the catalytic activity of purified human GST P1-1 in vitro. RESULTS: We identified chlorophyllide, merbromine, hexachlorophene, and ethacrynic acid as the most effective GST P1-1 inhibitors with IC50 values in the low micromolar range. For comparison, these compounds were even more potent in the inhibition of human GST A3-3, an enzyme implicated in steroid hormone biosynthesis. In distinction from the other inhibitors, which showed conventional inhibition patterns, the competitive inhibitor ethacrynic acid elicited strong kinetic cooperativity in the glutathione saturation of GST P1-1. Apparently, ethacrynic acid serves as an allosteric inhibitor of the enzyme. CONCLUSION AND PRACTICAL IMPLICATIONS: In their own right, the compounds investigated are less potent than desired for adjuvants in cancer chemotherapy, but the structures of the most potent inhibitors could serve as leads for the synthesis of more efficient adjuvants.


Subject(s)
Enzyme Inhibitors/pharmacology , Glutathione S-Transferase pi/antagonists & inhibitors , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Chlorophyllides/pharmacology , Drug Approval , Drug Evaluation, Preclinical , Drug Resistance, Neoplasm , Enzyme Inhibitors/chemistry , Ethacrynic Acid/pharmacology , Glutathione Transferase/antagonists & inhibitors , Hexachlorophene/pharmacology , Humans , Kinetics , Merbromin/pharmacology , Neoplasms/drug therapy , Neoplasms/enzymology , Recombinant Proteins/antagonists & inhibitors , United States , United States Food and Drug Administration
17.
PLoS One ; 7(12): e51820, 2012.
Article in English | MEDLINE | ID: mdl-23251633

ABSTRACT

The voltage-gated KCNQ1 potassium channel is expressed in cardiac tissues, and coassembly of KCNQ1 with an auxiliary KCNE1 subunit mediates a slowly activating current that accelerates the repolarization of action potential in cardiomyocytes. Mutations of KCNQ1 genes that result in reduction or loss of channel activity cause prolongation of repolarization during action potential, thereby causing long QT syndrome (LQTs). Small molecule activators of KCNQ1/KCNE1 are useful both for understanding the mechanism of the complex activity and for developing therapeutics for LQTs. In this study we report that hexachlorophene (HCP), the active component of the topical anti-infective prescription drug pHisoHex, is a KCNQ1/KCNE1 activator. HCP potently increases the current amplitude of KCNQ1/KCNE1 expressed by stabilizing the channel in an open state with an EC(50) of 4.61 ± 1.29 µM. Further studies in cardiomyocytes showed that HCP significantly shortens the action potential duration at 1 µM. In addition, HCP is capable of rescuing the loss of function of the LQTs mutants caused by either impaired activation gating or phosphatidylinositol-4,5-bisphosphate (PIP2) binding affinity. Our results indicate HCP is a novel KCNQ1/KCNE1 activator and may be a useful tool compound for the development of LQTs therapeutics.


Subject(s)
Hexachlorophene/pharmacology , KCNQ1 Potassium Channel/metabolism , Long QT Syndrome/drug therapy , Long QT Syndrome/metabolism , Action Potentials/drug effects , Animals , CHO Cells , Cell Line , Cricetinae , Heart/drug effects , Heart/physiopathology , KCNQ1 Potassium Channel/genetics , Long QT Syndrome/genetics , Mutation/drug effects , Myocytes, Cardiac/drug effects , Myocytes, Cardiac/metabolism , Myocytes, Cardiac/physiology , Phosphatidylinositol 4,5-Diphosphate/pharmacology
18.
J Vet Med Sci ; 74(7): 837-43, 2012 Jul.
Article in English | MEDLINE | ID: mdl-22313968

ABSTRACT

The goal of this research was to identify mechanisms responsible for the spongy change induced in rats after repeated hexachlorophene (HCP) or cuprizone (CPZ) dosing. Rats were dosed with 35 mg/kg HCP for 5 days followed by drug withdrawal for 7 days suffered spongy changes to the white matter of the cerebrum, cerebellum, medulla oblongata, and spinal cord that were accompanied by degeneration of oligodendroglia. The severity of both lesions increased prominently on day 5; however, the spongy change decreased and degeneration of oligodendroglia reversed on day 12 (7 days after dosing ceased). On day 12, cerebral cortex oligodendroglia were stained strongly by anti-CNPase. Other rats were fed for 8 days with powdered chow containing 1% (w/w) CPZ, which was then withdrawn for 16 days. The rats exhibited the spongy change in the white matter of the cerebrum and cerebellum as well as oligodendroglial cell death from day 3. The severity of both lesions increased prominently on day 8. Cerebral cortex oligodendroglia were stained strongly by anti-CNPase on days 3 to 8 and decreased to the control levels by day 24 (16 days after dosing ceased). Electron microscopy revealed that oligodendroglia frequently displayed apoptotic morphology. These findings suggest that CNPase expression was induced in the course of restoration following HCP-induced insults to oligodendroglia and the myelin sheath, and in the course of demyelination by CPZ-induced damage to oligodendroglia. However, the role of CNPase on both courses is unclear.


Subject(s)
2',3'-Cyclic-Nucleotide Phosphodiesterases/metabolism , Brain/drug effects , Cuprizone/pharmacology , Hexachlorophene/pharmacology , Myelin Sheath/drug effects , Animals , Brain/growth & development , Brain/pathology , Microscopy, Electron , Myelin Sheath/pathology , Rats , Time Factors
19.
Antimicrob Agents Chemother ; 56(2): 1084-6, 2012 Feb.
Article in English | MEDLINE | ID: mdl-22083485

ABSTRACT

Recurrent community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) skin infections are an increasingly common problem. However, there are no data on the efficacy of decolonization regimens. We prospectively evaluated 31 patients with recurrent CA-MRSA skin infections who received nasal mupirocin, topical hexachlorophene body wash, and an oral anti-MRSA antibiotic. The mean number of MRSA infections after the intervention decreased significantly from baseline (0.03 versus 0.84 infections/month, P = <0.0001). This regimen appears promising at preventing recurrent CA-MRSA infections.


Subject(s)
Anti-Bacterial Agents/therapeutic use , Community-Acquired Infections/prevention & control , Hexachlorophene/therapeutic use , Methicillin-Resistant Staphylococcus aureus/drug effects , Mupirocin/therapeutic use , Staphylococcal Skin Infections/prevention & control , Administration, Intranasal , Administration, Oral , Administration, Topical , Adolescent , Adult , Aged , Aged, 80 and over , Anti-Bacterial Agents/administration & dosage , Anti-Bacterial Agents/pharmacology , Anti-Infective Agents, Local , Community-Acquired Infections/drug therapy , Community-Acquired Infections/microbiology , Female , Hexachlorophene/administration & dosage , Hexachlorophene/pharmacology , Humans , Male , Middle Aged , Mupirocin/administration & dosage , Mupirocin/pharmacology , Recurrence , Staphylococcal Skin Infections/drug therapy , Staphylococcal Skin Infections/microbiology , Treatment Outcome , Young Adult
20.
J Biol Chem ; 284(34): 22988-3000, 2009 Aug 21.
Article in English | MEDLINE | ID: mdl-19531491

ABSTRACT

Mammalian glutamate dehydrogenase (GDH) is a homohexameric enzyme that catalyzes the reversible oxidative deamination of l-glutamate to 2-oxoglutarate using NAD(P)(+) as coenzyme. Unlike its counterparts from other animal kingdoms, mammalian GDH is regulated by a host of ligands. The recently discovered hyperinsulinism/hyperammonemia disorder showed that the loss of allosteric inhibition of GDH by GTP causes excessive secretion of insulin. Subsequent studies demonstrated that wild-type and hyperinsulinemia/hyperammonemia forms of GDH are inhibited by the green tea polyphenols, epigallocatechin gallate and epicatechin gallate. This was followed by high throughput studies that identified more stable inhibitors, including hexachlorophene, GW5074, and bithionol. Shown here are the structures of GDH complexed with these three compounds. Hexachlorophene forms a ring around the internal cavity in GDH through aromatic stacking interactions between the drug and GDH as well as between the drug molecules themselves. In contrast, GW5074 and bithionol both bind as pairs of stacked compounds at hexameric 2-fold axes between the dimers of subunits. The internal core of GDH contracts when the catalytic cleft closes during enzymatic turnover. None of the drugs cause conformational changes in the contact residues, but all bind to key interfaces involved in this contraction process. Therefore, it seems likely that the drugs inhibit enzymatic turnover by inhibiting this transition. Indeed, this expansion/contraction process may play a major role in the inter-subunit communication and allosteric regulation observed in GDH.


Subject(s)
Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Glutamate Dehydrogenase/antagonists & inhibitors , Glutamate Dehydrogenase/chemistry , Indoles/chemistry , Phenols/chemistry , Allosteric Regulation/drug effects , Animals , Bithionol/chemistry , Bithionol/pharmacology , Cattle , Escherichia coli Proteins/antagonists & inhibitors , Escherichia coli Proteins/chemistry , Escherichia coli Proteins/metabolism , Glutamate Dehydrogenase/metabolism , Guanosine Triphosphate/chemistry , Guanosine Triphosphate/pharmacology , Hexachlorophene/chemistry , Hexachlorophene/pharmacology , Indoles/pharmacology , Kinetics , NADP/chemistry , NADP/pharmacology , Phenols/pharmacology , Protein Conformation/drug effects , Protein Structure, Quaternary , Protein Structure, Secondary , Protein Structure, Tertiary , Tetrahymena/enzymology
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