Walter A. Szarek

Queen's University, Kingston, Ontario, Canada

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Publications (267)397.47 Total impact

  • Source
    [show abstract] [hide abstract]
    ABSTRACT: Previously, we reported that menadione activated rat, native heme oxygenase-2 (HO-2) and human recombinant heme oxygenase-2 selectively; it did not activate spleen, microsomal heme oxygenase-1. The purpose of this study was to explore some structure-activity relationships of this activation and the idea that redox properties may be an important aspect of menadione efficacy. Heme oxygenase activity was determined in vitro using rat spleen and brain microsomes as the sources of heme oxygenase-1 and -2, respectively, as well as recombinant, human heme oxygenase-2. Menadione analogs with bulky aliphatic groups at position-3, namely vitamins K1 and K2, were not able to activate HO-2. In contrast, several compounds with similar bulky but less lipophilic moieties at position-2 (and -3) were able to activate HO-2 many fold; these compounds included polar, rigid, furan-containing naphthoquinones, furan-benzoxazine naphthoquinones, 2-(aminophenylphenyl)-3-piperidin-1-yl naphthoquinones. To explore the idea that redox properties might be involved in menadione efficacy, we tested analogs such as 1,4-dimethoxy-2-methylnaphthalene, pentafluoromenadione, monohalogenated naphthoquinones, alpha-tetralone and 1,4-naphthoquinone. All of these compounds were inactive except for 1,4-naphthoquinone. Menadione activated full-length recombinant human heme oxygenase-2 (FL-hHO-2) as effectively as rat brain enzyme, but it did not activate rat spleen heme oxygenase. These observations are consistent with the idea that naphthoquinones such as menadione bind to a receptor in HO-2 and activate the enzyme through a mechanism that may involve redox properties.
    Medical gas research. 02/2014; 4(1):4.
  • Gheorghe Roman, Ian E Crandall, Walter A Szarek
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    ABSTRACT: A series of compounds structurally related to astemizole were designed and synthesized with the goal of determining their anti-Plasmodium activity. Several modifications of the astemizole structure, namely the removal of the 4-fluorobenzyl and/or 4-methoxyphenethyl moieties, substitution of the benzene ring of the benzimidazole scaffold, replacement of the fluorine atom in the 4-fluorobenzyl group, and variation of the 4-aminopiperidine moiety, were explored. In vitro evaluation of the anti-Plasmodium activity of these compounds using the ItG strain showed that astemizole and some of its structurally similar derivatives have IC50 values in the nanomolar range and exhibit toxicity towards the parasite over Chinese ovarian hamster (CHO) cells with a selectivity as high as 200. The presence of a secondary cyclic amine at position 2 and substitution with chlorine at positions 4 and 5 in the benzimidazole moiety are two modifications that resulted in potent and selective antimalarials based on astemizole.
    ChemMedChem 09/2013; · 2.84 Impact Factor
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    ABSTRACT: Several analogs based on the lead structure of 1-(4-chlorobenzyl)-2-(pyrrolidin-1-ylmethyl)-1H-benzimidazole (clemizole) were synthesized and evaluated as novel inhibitors of heme oxygenase (HO). Many of the compounds were found to be potent and highly selective for the HO-2 isozyme (constitutive), and had substantially less inhibitory activity on the HO-1 isozyme (inducible). The compounds represent the first report of highly potent and selective inhibitors of HO-2 activity, and complement our suite of selective HO-1 inhibitors. The study has revealed many candidates based on the inhibition of heme oxygenases for potentially useful pharmacological and therapeutic applications.
    Bioorganic & medicinal chemistry 08/2013; · 2.82 Impact Factor
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    ABSTRACT: BACKGROUND: Modifications of proteins by O-glycosylation determine many of the properties and functions of proteins. We wish to understand the mechanisms of O-glycosylation and develop inhibitors that could affect glycoprotein functions and alter cellular behavior. METHODS: We expressed recombinant soluble human Gal- and GlcNAc-transferases that synthesize the O-glycan cores 1 to 4 and are critical for the overall structures of O-glycans. We determined the properties and substrate specificities of these enzymes using synthetic acceptor substrate analogs. Compounds that were inactive as substrates were tested as inhibitors. RESULTS: Enzymes significantly differed in their recognition of the sugar moieties and aglycone groups of substrates. Core 1 synthase was active with glycopeptide substrates but GlcNAc-transferases preferred substrates with hydrophobic aglycone groups. Chemical modifications of the acceptors shed light on enzyme - substrate interactions. Core 1 synthase was weakly inhibited by its substrate analog benzyl 2-butanamido-2-deoxy-α-D-galactoside while two of the three GlcNAc-transferases were selectively and potently inhibited by bis-imidazolium salts which are not substrate analogs. CONCLUSIONS: This work delineates the distinct specificities and properties of the enzymes that synthesize the common O-glycan core structures 1 to 4. New inhibitors were found that could selectively inhibit the synthesis of core 1, 2 and 3 but not core 4. General significance These studies help our understanding of the mechanisms of action of enzymes critical for O-glycosylation. The results may be useful for the re-engineering of O-glycosylation to determine the roles of O-glycans and the enzymes critical for O-glycosylation, and for biotechnology with potential therapeutic applications.
    Biochimica et Biophysica Acta 04/2013; · 4.66 Impact Factor
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    ABSTRACT: Galactosyltransferases (GalTs) extend the glycan chains of mammalian glycoproteins by adding Gal to terminal GlcNAc residues, and thus build the scaffolds for biologically important glycan structures. We have shown that positively charged bivalent imidazolium salts in which the two imidazolium groups are linked by an aliphatic chain of 20 or 22 carbons form potent inhibitors of purified human β3-GalT5, using GlcNAcβ-benzyl as acceptor substrate. The inhibitors are not substrate analogs and also inhibited a selected number of other glycosyltransferases. These bis-imidazolium compounds represent a new class of glycosyltransferase inhibitors with potential as anti-cancer and anti-inflammatory drugs.
    Bioorganic & medicinal chemistry 01/2013; · 2.82 Impact Factor
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    ABSTRACT: The interaction between DNA and members of series of bivalent imidazole compounds, monovalent and bivalent imidazolium compounds, and monovalent and bivalent tetrazolium compounds, which had been synthesized and evaluated for their anti-Plasmodium activity, has been examined using the displacement of SYBR Green I as a measure of competitive binding. The degree of interaction with DNA appears to be dependent on both hydrophobic and charge-pairing interactions.
    Bioorganic & medicinal chemistry letters 12/2012; · 2.65 Impact Factor
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    ABSTRACT: The development of heme oxygenase (HO) inhibitors, especially those that are isozyme-selective, promises powerful pharmacological tools to elucidate the regulatory characteristics of the HO system. It is already known that HO has cytoprotective properties and may play a role in several disease states, making it an enticing therapeutic target. Traditionally, the metalloporphyrins have been used as competitive HO inhibitors owing to their structural similarity with the substrate, heme. However, given heme's important role in several other proteins (e.g. cytochromes P450, nitric oxide synthase), non-selectivity is an unfortunate side-effect. Reports that azalanstat and other non-porphyrin molecules inhibited HO led to a multi-faceted effort to develop novel compounds as potent, selective inhibitors of HO. This resulted in the creation of non-competitive inhibitors with selectivity for HO, including a subset with isozyme selectivity for HO-1. Using X-ray crystallography, the structures of several complexes of HO-1 with novel inhibitors have been elucidated, which provided insightful information regarding the salient features required for inhibitor binding. This included the structural basis for non-competitive inhibition, flexibility and adaptability of the inhibitor binding pocket, and multiple, potential interaction subsites, all of which can be exploited in future drug-design strategies.
    Journal of The Royal Society Interface 10/2012; · 4.91 Impact Factor
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    ABSTRACT: The enterohemorrhagic O157 strain of Escherichia coli, which is one of the most well-known bacterial pathogens, has an O-antigen repeating unit structure with the sequence [-2-d-Rha4NAcα1-3-l-Fucα1-4-d-Glcβ1-3-d-GalNAcα1-]. The O-antigen gene cluster of E. coli O157 contains the genes responsible for the assembly of this repeating unit and includes wbdN. In spite of cloning many O-antigen genes, biochemical characterization has been done on very few enzymes involved in O-antigen synthesis. In this work, we expressed the wbdN gene in E. coli BL21, and the His-tagged protein was purified. WbdN activity was characterized using the donor substrate UDP-[(14)C]Glc and the synthetic acceptor substrate GalNAcα-O-PO(3)-PO(3)-(CH(2))(11)-O-Ph. The enzyme product was isolated by high pressure liquid chromatography, and mass spectrometry showed that one Glc residue was transferred to the acceptor by WbdN. Nuclear magnetic resonance analysis of the product structure indicated that Glc was β1-3 linked to GalNAc. WbdN contains a conserved DxD motif and requires divalent metal ions for full activity. WbdN activity has an optimal pH between 7 and 8 and is highly specific for UDP-Glc as the donor substrate. GalNAcα derivatives lacking the diphosphate group were inactive as substrates, and the enzyme did not transfer Glc to GlcNAcα-O-PO(3)-PO(3)-(CH(2))(11)-O-Ph. Our results illustrate that WbdN is a specific UDP-Glc:GalNAcα-diphosphate-lipid β1,3-Glc-transferase. The enzyme is a target for the development of inhibitors to block O157-antigen synthesis.
    Glycobiology 05/2012; 22(8):1092-102. · 3.54 Impact Factor
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    ABSTRACT: Several α-(1H-imidazol-1-yl)-ω-phenylalkanes were synthesized and evaluated as novel inhibitors of heme oxygenase (HO). These compounds were found to be potent and selective for the stress-induced isozyme HO-1, showing mostly weak activity toward the constitutive isozyme HO-2. The introduction of an oxygen atom in the alkyl linker produced analogues with decreased potency toward HO-1, whereas the presence of a sulfur atom in the linker gave rise to analogues with greater potency toward HO-1 than the carbon-containing analogues. The most potent compounds studied contained a five-atom linker between the imidazolyl and phenyl moieties, whereas the most HO-1-selective compounds contained a four-atom linker between these groups. The compounds with a five-atom linker containing a heteroatom (O or S) were found to be the most potent inhibitors of HO-2; 1-(N-benzylamino)-3-(1H-imidazol-1-yl)propane dihydrochloride, with a nitrogen atom in the linker, was found to be inactive.
    ChemMedChem 03/2012; 7(5):897-902. · 2.84 Impact Factor
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    ABSTRACT: The development of heme oxygenase (HO) inhibitors is critical in dissecting and understanding the HO system and for potential therapeutic applications. We have established a program to design and optimize HO inhibitors using structure-activity relationships in conjunction with X-ray crystallographic analyses. One of our previous complex crystal structures revealed a putative secondary hydrophobic binding pocket which could be exploited for a new design strategy by introducing a functional group that would fit into this potential site. To test this hypothesis and gain further insights into the structural basis of inhibitor binding, we have synthesized and characterized 1-(1H-imidazol-1-yl)-4,4-diphenyl-2-butanone (QC-308). Using a carbon monoxide (CO) formation assay on rat spleen microsomes, the compound was found to be ∼15 times more potent (IC(50) = 0.27±0.07 µM) than its monophenyl analogue, which is already a potent compound in its own right (QC-65; IC(50) = 4.0±1.8 µM). The crystal structure of hHO-1 with QC-308 revealed that the second phenyl group in the western region of the compound is indeed accommodated by a definitive secondary proximal hydrophobic pocket. Thus, the two phenyl moieties are each stabilized by distinct hydrophobic pockets. This "double-clamp" binding offers additional inhibitor stabilization and provides a new route for improvement of human heme oxygenase inhibitors.
    PLoS ONE 01/2012; 7(1):e29514. · 3.73 Impact Factor
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    ABSTRACT: While substantial progress has been made in elucidating the roles of heme oxygenases-1 (HO-1) and -2 (HO-2) in mammals, our understanding of the functions of these enzymes in health and disease is still incomplete. A significant amount of our knowledge has been garnered through the use of nonselective inhibitors of HOs, and our laboratory has recently described more selective inhibitors for HO-1. In addition, our appreciation of HO-1 has benefitted from the availability of tools for increasing its activity through enzyme induction. By comparison, there is a paucity of information about HO-2 activation, with only a few reports appearing in the literature. This communication describes our observations of the up to 30-fold increase in the in-vitro activation of HO-2 by menadione. This activation was due to an increase in V(max) and was selective, in that menadione did not increase HO-1 activity.
    Canadian Journal of Physiology and Pharmacology 11/2011; · 1.56 Impact Factor
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    ABSTRACT: Cell surface mucins configure the cell surface by presenting extended protein backbones that are heavily O-glycosylated. The glycopeptide structures establish physicochemical properties at the cell surface that enable and block the formation of biologically important molecular complexes. Some mucins, such as MUC1, associate with receptor tyrosine kinases and other cell surface receptors, and engage in signal transduction in order to communicate information regarding conditions at the cell surface to the nucleus. In that context, the MUC1 cytoplasmic tail (MUC1CT) receives phosphorylation signals from receptor tyrosine kinases and serine/threonine kinases, which enables its association with different signaling complexes that conduct these signals to the nucleus and perhaps other subcellular organelles. We have detected the MUC1CT at promoters of over 500 genes, in association with several different transcription factors, and have shown that promoter occupancy can vary under different growth factor conditions. However, the full biochemical nature of the nuclear forms of MUC1 and its function at these promoter regions remain undefined. I will present evidence that nuclear forms of the MUC1CT include extracellular and cytoplasmic tail domains. In addition, I will discuss evidence for a hypothesis that the MUC1CT possesses a novel catalytic function that enables remodeling of the transcription factor occupancy of promoters, and thereby engages in regulation of gene expression.
    Glycobiology 11/2011; 21(11):1454-531. · 3.54 Impact Factor
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    ABSTRACT: A series of compounds containing bivalent imidazolium rings and one triazolium analog were synthesized and evaluated for their ability to inhibit the replication of Plasmodium falciparum cultures. The activity and selectivity of the compounds for P. falciparum cultures were found to depend on the presence of electron-deficient rings that were spaced an appropriate distance apart. The activity of the compounds was not critically dependent on the nature of the linker between the electron-deficient rings, an observation that suggests that the rings were responsible for the primary interaction with the molecular target of the compounds in the parasite. The bivalent imidazolium and triazolium compounds disrupted the process whereby merozoites gain entry into erythrocytes, however, they did not appear to prevent merozoites from forming. The compounds were also found to be active in a murine Plasmodium berghei infection, a result consistent with the compounds specifically interacting with a parasite component that is required for replication and is conserved between two Plasmodium species.
    Bioorganic & medicinal chemistry 06/2011; 19(21):6525-42. · 2.82 Impact Factor
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    ABSTRACT: AK1401 is a mutant of Pseudomonas aeruginosa strain PAO1 (serotype 05) that does not express O-antigen, but does express "A-band" lipopolysaccharide (LPS). The polysaccharide portion of the A-band LPS (A-PS) from AK1401 was found to consist mainly of D-rhamnose, with smaller amounts of 3-O-methylrhamnose, ribose, mannose, glucose, and a 3-O-methylhexose. 1H nuclear magnetic resonance spectra of the intact A-PS indicated that the main structural feature was a repeating trisaccharide of α-D-rhamnose having the following structure:The 1H NMR spectrum of the repeating unit was completely assigned through the use of 2D shift-correlated and relayed coherence transfer NMR spectroscopy. The linkage positions and sequence of residues were found by nuclear Overhauser enhancement difference spectroscopy. Key words: Pseudomonas aeruginosa, lipopolysaccharide, 1H NMR.
    Canadian Journal of Chemistry 02/2011; 69(8):1273-1280. · 0.96 Impact Factor
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    Walter A. Szarek, B. Mario Pinto, Masaharu Iwakawa
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    ABSTRACT: The synthesis of a variety of nucleoside analogs involving modifications in the carbohydrate ring is described. In particular, 6-substituted purin-9-yl derivatives of 1-oxa-4-thiacyclohexane and 1,4-dioxacyclohexane have been synthesized. A number of 6-chloropurin-9-yl derivatives of substituted 1-oxa-4-thiacyclohexane have also been derived from the parent compounds by way of a Pummerer rearrangement. A route to nucleoside analogs of 1-oxa-4-thiacyclohexane from naturally occurring nucleosides is illustrated for the case of inosine. A route to nucleoside analogs in which the carbohydrate moiety is replaced by an acyclic moiety bearing α,β-unsaturated esters is also illustrated for the case of uridine. The results of biological screening of these analogs and others previously synthesized in our program against leukemia L-1210 cells in vitro are presented; some of these compounds showed marginal antitumor activity. The screening results of selected compounds against the human HeLa cell line in vitro are also presented; none of the compounds that were tested showed significant inhibitory activity of cell growth.
    Canadian Journal of Chemistry 02/2011; 63(8):2149-2161. · 0.96 Impact Factor
  • Lucjan J. J. Hronowski, Walter A. Szarek
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    ABSTRACT: Synthesis of new carbocyclic analogs of 1-(2′,3′-dideoxy-glycero-pentofuranosyl)pyrimidine nucleosides having the uracil (34), 2-thiouracil (33), 2-thiothymine (31), cytosine (44), and 5-methylcytosine (43) bases is described. The nucleoside analogs having the uracil, 2-thiouracil, and 2-thiothymine bases were prepared by coupling cis-3-aminocyclopentanemethanol (8) with 3-ethoxypropenoyl isocyanate (26), 3-ethoxypropenoyl isothiocyanate (25), and 3-methoxy-2-methylpropenoyl isothiocyanate (23), respectively, to give the corresponding acyl urea (30) and acyl thioureas (29 and 27). The acyl urea was cyclized in 2 N H2SO4 and the acyl thioureas in 15 N aqueous ammonia to give the corresponding nucleoside analogs. The nucleoside analogs containing the cytosine (44) and 5-methylcytosine (43) bases were prepared from the uracil and thymine nucleoside analogs, respectively, by way of the 4-chloropyrimidinone intermediates (42 and 41). The synthesis of cis-3-aminocyclopentanemethanol (8) from norbornene by way of cis-1,3-cyclopentanedicarboxylic acid anhydride (3) is also described. In addition, the ease of nucleophilic opening of compound 3 is compared to the opening of camphoric anhydride (9), which contains a cis-vicinal substituent at position 2. The relative ease of opening of compound 3 is discussed with respect to the effect, observed in an earlier study, that a cis-vicinal acetoxy group has on the course of the nucleophilic opening of such anhydrides. The 1H magnetic resonance spectra at 200 MHz of all of the synthetic intermediates and of the nucleoside analogs have been determined and discussed. The nucleoside analogs were screened for cell-growth inhibition using K-562 cells. Nucleoside analogs having the 2-thiouracil (33), 2-thiothymine (31), cytosine (44), and 5-methylcytosine (43) bases showed some growth inhibition with activity 150 to 300 times lower than that shown by 5-fluorouracil in this test system.
    Canadian Journal of Chemistry 02/2011; 66(1):61-70. · 0.96 Impact Factor
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    ABSTRACT: L-Glucose has been synthesized from D-glucose by a convenient method involving methyl 2,3-O-isopropylidene-β-D-gulo-furanosiduronic acid (6) as a key intermediate. Compound 6 was converted into L-glucono-1,5-lactone (8), which, by a selective reduction, afforded L-glucose (9).
    Canadian Journal of Chemistry 02/2011; 62(4):671-674. · 0.96 Impact Factor
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    Philip G. Hultin, Walter A. Szarek
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    ABSTRACT: The 1-(3-thianyl)uracil (9) and 9-(3-thianyl)adenine (14) nucleoside analogs have been prepared from the key intermediate, (±)-(3β,5β)-3-amino-5-(hydroxymethyl)thiane (6). Analog 9 was converted into a mixture of diastereomeric sulfoxides (10) that afforded, by a Pummerer reaction, a mixture of (±)-1-{(2′β,3′β,,5′β)-2′-acetoxy-5′-(acetoxymethyl)thian-3′-yl}-2,4(1H,3H)-pyrimidinedione (11a) and its 6′-β isomer (11b). The EI mass spectra of the nucleoside analogs are discussed. The uracil nucleoside analogs have been evaluated also for their anti-HIV and antitumor activities.
    Canadian Journal of Chemistry 02/2011; 72(1):208-213. · 0.96 Impact Factor
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    ABSTRACT: The utility of the Pictet–Spengler condensation of (R)-(+)-glyceraldehyde with dopamine hydrochloride in the enantioselective synthesis of isoquinoline alkaloids is demonstrated by the preparation of (S)-(−)-carnegine, (R)-(−)-calycotomine, and (S)-(+)-laudanosine.
    Canadian Journal of Chemistry 02/2011; 64(11):2205-2210. · 0.96 Impact Factor
  • Canadian Journal of Chemistry 02/2011; 62(12):2709-2711. · 0.96 Impact Factor

Publication Stats

1k Citations
609 Downloads
397.47 Total Impact Points

Institutions

  • 1968–2013
    • Queen's University
      • • Department of Chemistry
      • • Division of Rheumatology
      • • Department of Pharmacology and Toxicology
      • • Department of Biochemistry
      • • Department of Paediatrics
      Kingston, Ontario, Canada
    • Rutgers, The State University of New Jersey
      • Department of Chemical Biology
      New Brunswick, NJ, United States
  • 2007–2012
    • University of Toronto
      • Division of Infectious Diseases
      Toronto, Ontario, Canada
  • 1996
    • McGill University
      • Department of Anatomy and Cell Biology
      Montréal, Quebec, Canada