| 1. |
- Adamiak, Beata, et al.
(författare)
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Human antibodies to herpes simplex virus type 1 glycoprotein C are neutralizing and target the heparan sulfate-binding domain
- 2010
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Ingår i: Virology. - : Elsevier BV. - 0042-6822. ; 400:2, s. 197-206
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Tidskriftsartikel (refereegranskat)abstract
- Human antibodies specific for glycoprotein C (gC1) of herpes simplex virus type 1 (HSV-1) neutralized the virus infectivity and efficiently inhibited attachment of HSV-1 to human HaCaT keratinocytes and to murine mutant L cells expressing either heparan sulfate or chondroitin sulfate at the cell surface. Similar activities were observed with anti-gC1 monoclonal antibody B1C1. In addition to HaCaT and L cells, B1C1 antibody neutralized HSV-1 infectivity in simian GMK AH1 cells mildly pre-treated with heparinase III. Human anti-gC1 antibodies efficiently competed with the binding of gC1 to B1C1 antibody whose epitope overlaps a part of the attachment domain of gC1. Human anti-gC1 and B1C1 antibodies extended survival time of mice experimentally infected with HSV-1. We conclude that in HaCaT cells and in cell systems showing restricted expression of glycosaminoglycans, human and some monoclonal anti-gC1 antibodies can target the cell-binding domain of this protein and neutralize viral infectivity.
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| 2. |
- Ekblad, Maria, 1978, et al.
(författare)
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A highly lipophilic sulfated tetrasaccharide glycoside related to muparfostat (PI-88) exhibits virucidal activity against herpes simplex virus.
- 2010
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Ingår i: Antiviral research. - : Elsevier BV. - 1872-9096 .- 0166-3542. ; 86:2, s. 196-203
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Tidskriftsartikel (refereegranskat)abstract
- Although sulfated polysaccharides potently inhibit the infectivity of herpes simplex virus (HSV) and human immunodeficiency virus in cultured cells, these compounds fail to show protective effects in humans, most likely due to their poor virucidal activity. Herein we report on sulfated oligosaccharide glycosides related to muparfostat (formerly known as PI-88) and their assessment for anti-HSV activity. Chemical modifications based on the introduction of specific hydrophobic groups at the reducing end of a sulfated oligosaccharide chain enhanced the compound's capability to inhibit the infection of cells by HSV-1 and HSV-2 and abrogated the cell-to-cell transmission of HSV-2. Furthermore, modification with a highly lipophilic cholestanyl group provided a compound with virucidal activity against HSV. This glycoside targeted the viral particle and, to a lesser degree, the cell, and exhibited an antiviral mode of action typical for sulfated polysaccharides and virucides, i.e., interference with the virus attachment to cells and irreversible inactivation of virus infectivity, respectively. The virucidal activity was decreased in the presence of human cervical secretions suggesting that higher doses of this glycoside might be needed for in vivo application. Altogether, the sulfated oligosaccharide-cholestanyl glycoside exhibits potent anti-HSV activity and is, therefore, a good candidate for development as a virucide.
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| 3. |
- Lundin, Anna, et al.
(författare)
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Potent anti-respiratory syncytial virus activity of a cholestanol-sulfated tetrasaccharide conjugate.
- 2012
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Ingår i: Antiviral research. - : Elsevier BV. - 1872-9096 .- 0166-3542. ; 93:1, s. 101-9
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Tidskriftsartikel (refereegranskat)abstract
- A number of different viruses including respiratory syncytial virus (RSV) initiate infection of cells by binding to cell surface glycosaminoglycans and sulfated oligo- and polysaccharide mimetics of these receptors exhibit potent antiviral activity in cultured cells. We investigated whether the introduction of different lipophilic groups to the reducing end of sulfated oligosaccharides would modulate their anti-RSV activity. Our results demonstrate that the cholestanol-conjugated tetrasaccharide (PG545) exhibited ∼5- to 16-fold enhanced anti-RSV activity in cultured cells compared with unmodified sulfated oligosaccharides. Furthermore, PG545 displayed virus-inactivating (virucidal) activity, a feature absent in sulfated oligosaccharides. To inhibit RSV infectivity PG545 had to be present during the initial steps of viral infection of cells. The anti-RSV activity of PG545 was due to both partial inhibition of the virus attachment to cells and a more profound interference with some post-attachment steps as PG545 efficiently neutralized infectivity of the cell-adsorbed virus. The anti-RSV activity of PG545 was reduced when tested in the presence of human nasal secretions. Serial passages of RSV in the presence of increasing concentrations of PG545 selected for weakly resistant viral variants that comprised the F168S and the P180S amino acid substitutions in the viral G protein. Altogether we identified a novel and potent inhibitor of RSV, which unlike sulfated oligo- and polysaccharide compounds, could irreversibly inactivate RSV infectivity.
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| 4. |
- Lundin, Anna, et al.
(författare)
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Screening and evaluation of anti-respiratory syncytial virus compounds in cultured cells.
- 2013
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Ingår i: Antiviral Methods and Protocols. Ed. Edwin Yunhao Gong. (Methods in molecular biology, 1030). - Totowa, NJ : Humana Press. - 1940-6029. - 9781627034838 ; 1030, s. 345-63
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Bokkapitel (refereegranskat)abstract
- Respiratory syncytial virus (RSV) is a highly contagious pathogen that infects mainly ciliated cells of respiratory epithelium and type 1 pneumocytes in the alveoli frequently causing serious respiratory disease in infants, elderly, and immunocompromised patients. At present, prevention/treatment of RSV infection is limited to the use of specific anti-RSV antibody or an aerosol formulation of ribavirin, a drug of suboptimal efficacy and low safety profile. There is an urgent need for development of novel anti-RSV drugs and virucides. Here we describe the cell culture-based methods used in our laboratory in identification of novel inhibitors of RSV including the P13 fusion inhibitor, and the PG545 virucide. Protocols for antiviral screening, evaluation of anti-RSV potency, and elucidation of mode of antiviral activity of test compounds are described.
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| 5. |
- Lundin, Anna, et al.
(författare)
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Targeting Membrane-Bound Viral RNA Synthesis Reveals Potent Inhibition of Diverse Coronaviruses Including the Middle East Respiratory Syndrome Virus
- 2014
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Ingår i: Plos Pathogens. - : Public Library of Science (PLoS). - 1553-7366 .- 1553-7374. ; 10:5
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Tidskriftsartikel (refereegranskat)abstract
- Coronaviruses raise serious concerns as emerging zoonotic viruses without specific antiviral drugs available. Here we screened a collection of 16671 diverse compounds for anti-human coronavirus 229E activity and identified an inhibitor, designated K22, that specifically targets membrane-bound coronaviral RNA synthesis. K22 exerts most potent antiviral activity after virus entry during an early step of the viral life cycle. Specifically, the formation of double membrane vesicles (DMVs), a hallmark of coronavirus replication, was greatly impaired upon K22 treatment accompanied by near-complete inhibition of viral RNA synthesis. K22-resistant viruses contained substitutions in non-structural protein 6 (nsp6), a membrane-spanning integral component of the viral replication complex implicated in DMV formation, corroborating that K22 targets membrane bound viral RNA synthesis. Besides K22 resistance, the nsp6 mutants induced a reduced number of DMVs, displayed decreased specific infectivity, while RNA synthesis was not affected. Importantly, K22 inhibits a broad range of coronaviruses, including Middle East respiratory syndrome coronavirus (MERS-CoV), and efficient inhibition was achieved in primary human epithelia cultures representing the entry port of human coronavirus infection. Collectively, this study proposes an evolutionary conserved step in the life cycle of positive-stranded RNA viruses, the recruitment of cellular membranes for viral replication, as vulnerable and, most importantly, druggable target for antiviral intervention. We expect this mode of action to serve as a paradigm for the development of potent antiviral drugs to combat many animal and human virus infections.
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| 6. |
- Lundin, Anna, et al.
(författare)
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Two novel fusion inhibitors of human respiratory syncytial virus
- 2010
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Ingår i: Antiviral Research. - : Elsevier BV. - 0166-3542. ; 88:3, s. 317-324
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Tidskriftsartikel (refereegranskat)abstract
- To search for novel drugs against human respiratory syncytial virus (RSV), we have screened a diversity collection of 16,671 compounds for anti-RSV activity in cultures of HEp-2 cells. Two of the hit compounds, i.e., the N-(2-hydroxyethyl)-4-methoxy-N-methyl-3-(6-methyl[1,2,4]triazolo[3,4-a]phthalazin-3-yl)benzenesulfonamide (designated as P13) and the 1,4-bis(3-methyl-4-pyridinyl)-1,4-diazepane (designated as C15), reduced the virus infectivity with IC₅₀ values of 0.11 and 0.13μM respectively. The concentration of P13 and C15 that reduced the viability of HEp-2 cells by 50% was 310 and 75μM respectively. Both P13 and C15 exhibited no direct virucidal activity or inhibitory effects on the virus attachment to cells. However, to inhibit formation of RSV-induced syncytial plaques P13 and C15 had to be present during the virus entry into the cells and the cell-to-cell transmission of the virus. The RSV multiplication in HEp-2 cells in the presence of P13 or C15 resulted in rapid selection of viral variants that were ∼1000 times less sensitive to these drugs than original virus. Sequencing of resistant viruses revealed presence of amino acid substitutions in the F protein of RSV, i.e., the D489G for C15-selected, and the T400I and N197T (some clones) for the P13-selected virus variants. In conclusion, we have identified two novel fusion inhibitors of RSV, and the detailed understanding of their mode of antiviral activity including selection for the drug resistant viral variants may help to develop selective and efficient anti-RSV drugs.
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| 7. |
- Nordén, Rickard, 1977, et al.
(författare)
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Involvement of viral glycoprotein gC-1 in expression of the selectin ligand sialyl-Lewis X induced after infection with herpes simplex virus type 1.
- 2013
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Ingår i: APMIS : acta pathologica, microbiologica, et immunologica Scandinavica. - : Wiley. - 1600-0463. ; 121:4, s. 280-289
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Tidskriftsartikel (refereegranskat)abstract
- Several herpesviruses induce expression of the selectin receptor sialyl-Lewis X (sLe(x) ) by activating transcription of one or more of silent host FUT genes, each one encoding a fucosyltransferase that catalyses the rate-limiting step of sLe(x) synthesis. The aim here was to identify the identity of the glycoconjugate associated with sLe(x) glycoepitope in herpes simplex virus type 1 (HSV-1) infected human diploid fibroblasts, using immunofluorescence confocal microscopy. Cells infected with all tested HSV-1 strains analysed demonstrated bright sLe(x) fluorescence, except for two mutant viruses that were unable to induce proper expression of viral glycoprotein gC-1: One gC-1 null mutant and another mutant expressing gC-1 devoid of its major O-glycan-containing region (aa 33-116). The sLe(x) reactivity of HSV-1 infected cells was abolished by mild alkali treatment. Altogether the results indicated that the detectable sLe(x) was associated with O-linked glycans, situated in the mucin region of gC-1. No evidence for sLe(x) (i) in other HSV-1 glycoproteins with mucin domains such as gI-1 or (ii) in host cell glycoproteins/glycolipids was found. Thus, the mucin domain of HSV-1 gC-1 may support expression of selectin ligands such as sLe(x) and other larger O-linked glycans in cell types lacking endogenous mucin domain-containing glycoproteins, optimized for O-glycan expression, provided that the adequate host glycosyltransferase genes are activated.
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| 8. |
- Said, Joanna, et al.
(författare)
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HIV-1 variants with reduced sensitivity to sulfated oligosaccharide muparfostat contain mutations in the envelope glycoproteins gp120 and gp41
- 2013
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Ingår i: Journal of Antivirals and Antiretrovirals. - : OMICS Publishing Group. - 1948-5964. ; 5:3, s. 50-56
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Tidskriftsartikel (refereegranskat)abstract
- Attachment of human immunodefciency virus type 1 (HIV-1) to host cells is primarily mediated by cell surface molecules CD4 and either of the chemokine co-receptors CCR5 or CXCR4, and is facilitated by cellular heparansulfate chains of syndecans. Although mimetics of heparansulfate exhibit potent anti-HIV-1 activity in cultured cells, these compounds failed to prevent infection in humans when used in clinical trials as microbicides. We have previously shown that the low molecular weight and extensively sulfated oligosaccharide muparfostat coupled to cholestanol exhibited virucidal activity while the non-conjugated muparfostat (formerly known as PI-88) inhibited HIV-1 infection of cultured cells in a reversible manner only. To initiate clarifcation of distinct anti-HIV-1 potencies of muparfostat and muparfostat-cholestanol conjugate, in this work we sought to select for viral resistance using the less potent muparfostat. The laboratory strain HIV-1IIIB was successively propagated in H9 cells in the presence of the compound. The virus selected for after 21-24 passages appeared to be approximately 3-4 times less sensitive to muparfostat than the original HIV-1IIIB strain or control virus passaged in parallel in the absence of muparfostat. Comparative analysis of nucleotide sequences of these viruses revealed presence of the I152V substitution in V2, the K276R change in V3, the deletion of fve amino acid repeat 366FNSTW370 in V4 of gp120, and the L33S and A101T alterations in transmembrane gp41 component of the muparfostat passaged virus. Selection for viral variants with mutations in gp41 was an unexpected observation as this protein of HIV-1 is seldom targeted by sulfated polysaccharides. © 2013 Said J, et al.
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| 9. |
- Said, Joanna, et al.
(författare)
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Lipophile-conjugated sulfated oligosaccharides as novel microbicides against HIV-1.
- 2010
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Ingår i: Antiviral research. - : Elsevier BV. - 1872-9096 .- 0166-3542. ; 86:3, s. 286-295
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Tidskriftsartikel (refereegranskat)abstract
- With the aim of providing compounds suitable for further development as microbicides active against human immunodeficiency virus 1 (HIV-1) a library containing 37 lipophile-conjugated sulfated oligosaccharides was screened for antiviral and virucidal activity against this virus. Four highly active compounds had low drug inhibition concentrations (IC(50)) for HIV-1 and inactivated viral particles, suggestive of virucidal properties. Two of these compounds comprising a sulfated tetrasaccharide linked to a cholestanol group by a glycosidic bond, showed low toxicity and high selectivity indices. The two compounds were active both against CCR5 and dual-tropic CCR5/CXCR4 clinical HIV-1 isolates. Since herpes simplex virus type 2 (HSV-2) may be a cofactor for HIV-1 infection, the virucidal effect of the compounds was demonstrated against both viruses when mixed and incubated together on permissive cells. Incubation of compounds with serum, and to a lesser degree, cervical secretions, reduced the HIV-1 inactivating capacity, which suggests the need for molecular modification to reduce host protein binding. Considering the virucidal effect and low toxicity, these sulfated oligosaccharides with lipophilic tails may offer new possibilities of microbicide development.
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