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Chandra, N., Frängsmyr, L. & Arnberg, N. (2019). Decoy Receptor Interactions as Novel Drug Targets against EKC-Causing Human Adenovirus. Viruses, 11(3), Article ID E242.
Open this publication in new window or tab >>Decoy Receptor Interactions as Novel Drug Targets against EKC-Causing Human Adenovirus
2019 (English)In: Viruses, ISSN 1999-4915, E-ISSN 1999-4915, Vol. 11, no 3, article id E242Article in journal (Refereed) Published
Abstract [en]

Epidemic keratoconjunctivitis (EKC) is a severe ocular disease and can lead to visual impairment. Human adenovirus type-37 (HAdV-D37) is one of the major causative agents of EKC and uses sialic acid (SA)-containing glycans as cellular receptors. Currently, there are no approved antivirals available for the treatment of EKC. Recently, we have reported that sulfated glycosaminoglycans (GAGs) bind to HAdV-D37 via the fiber knob (FK) domain of the viral fiber protein and function as decoy receptors. Based on this finding, we speculated that GAG-mimetics may act as artificial decoy receptors and inhibit HAdV-D37 infection. Repurposing of approved drugs to identify new antivirals has drawn great attention in recent years. Here, we report the antiviral effect of suramin, a WHO-approved drug and a widely known GAG-mimetic, against HAdV-D37. Commercially available suramin analogs also show antiviral effects against HAdV-D37. We demonstrate that suramin exerts its antiviral activity by inhibiting the attachment of HAdV-D37 to cells. We also reveal that the antiviral effect of suramin is HAdV species-specific. Collectively, in this proof of concept study, we demonstrate for the first time that virus binding to a decoy receptor constitutes a novel and an unexplored target for antiviral drug development.

Place, publisher, year, edition, pages
MDPI, 2019
Keywords
GAG-mimetic, adenovirus, cellular receptor, decoy receptor, epidemic keratoconjunctivitis, glycosaminoglycans
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:umu:diva-158516 (URN)10.3390/v11030242 (DOI)000464389700002 ()30870979 (PubMedID)
Funder
Knut and Alice Wallenberg Foundation, KAW 2013.0019
Available from: 2019-04-29 Created: 2019-04-29 Last updated: 2019-08-28Bibliographically approved
Chandra, N., Frängsmyr, L., Imhof, S., Caraballo, R., Elofsson, M. & Arnberg, N. (2019). Sialic Acid-Containing Glycans as Cellular Receptors for Ocular Human Adenoviruses: Implications for Tropism and Treatment. Viruses, 11(5), Article ID 395.
Open this publication in new window or tab >>Sialic Acid-Containing Glycans as Cellular Receptors for Ocular Human Adenoviruses: Implications for Tropism and Treatment
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2019 (English)In: Viruses, ISSN 1999-4915, E-ISSN 1999-4915, Vol. 11, no 5, article id 395Article in journal (Refereed) Published
Abstract [en]

Human adenoviruses (HAdV) are the most common cause of ocular infections. Species B human adenovirus type 3 (HAdV-B3) causes pharyngoconjunctival fever (PCF), whereas HAdV-D8, -D37, and -D64 cause epidemic keratoconjunctivitis (EKC). Recently, HAdV-D53, -D54, and -D56 emerged as new EKC-causing agents. HAdV-E4 is associated with both PCF and EKC. We have previously demonstrated that HAdV-D37 uses sialic acid (SA)-containing glycans as cellular receptors on human corneal epithelial (HCE) cells, and the virus interaction with SA is mediated by the knob domain of the viral fiber protein. Here, by means of cell-based assays and using neuraminidase (a SA-cleaving enzyme), we investigated whether ocular HAdVs other than HAdV-D37 also use SA-containing glycans as receptors on HCE cells. We found that HAdV-E4 and -D56 infect HCE cells independent of SAs, whereas HAdV-D53 and -D64 use SAs as cellular receptors. HAdV-D8 and -D54 fiber knobs also bound to cell-surface SAs. Surprisingly, HCE cells were found resistant to HAdV-B3 infection. We also demonstrated that the SA-based molecule i.e., ME0462, designed to bind to SA-binding sites on the HAdV-D37 fiber knob, efficiently prevents binding and infection of several EKC-causing HAdVs. Surface plasmon resonance analysis confirmed a direct interaction between ME0462 and fiber knobs. Altogether, we demonstrate that SA-containing glycans serve as receptors for multiple EKC-causing HAdVs, and, that SA-based compound function as a broad-spectrum antiviral against known and emerging EKC-causing HAdVs.

Place, publisher, year, edition, pages
MDPI, 2019
Keywords
adenovirus, cellular receptor, epidemic keratoconjunctivitis, pharyngoconjunctival fever, sialic acid, tropism
National Category
Microbiology in the medical area
Identifiers
urn:nbn:se:umu:diva-159268 (URN)10.3390/v11050395 (DOI)000472676600006 ()31035532 (PubMedID)2-s2.0-85065483937 (Scopus ID)
Funder
Knut and Alice Wallenberg Foundation, 2013.0019
Available from: 2019-05-23 Created: 2019-05-23 Last updated: 2019-08-28Bibliographically approved
Chandra, N., Liu, Y., Liu, J.-X., Frängsmyr, L., Wu, N., Silva, L. M., . . . Arnberg, N. (2019). Sulfated Glycosaminoglycans as Viral Decoy Receptors for Human Adenovirus Type 37. Viruses, 11(3), Article ID E247.
Open this publication in new window or tab >>Sulfated Glycosaminoglycans as Viral Decoy Receptors for Human Adenovirus Type 37
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2019 (English)In: Viruses, ISSN 1999-4915, E-ISSN 1999-4915, Vol. 11, no 3, article id E247Article in journal (Refereed) Published
Abstract [en]

Glycans on plasma membranes and in secretions play important roles in infection by many viruses. Species D human adenovirus type 37 (HAdV-D37) is a major cause of epidemic keratoconjunctivitis (EKC) and infects target cells by interacting with sialic acid (SA)-containing glycans via the fiber knob domain of the viral fiber protein. HAdV-D37 also interacts with sulfated glycosaminoglycans (GAGs), but the outcome of this interaction remains unknown. Here, we investigated the molecular requirements of HAdV-D37 fiber knob:GAG interactions using a GAG microarray and demonstrated that fiber knob interacts with a broad range of sulfated GAGs. These interactions were corroborated in cell-based assays and by surface plasmon resonance analysis. Removal of heparan sulfate (HS) and sulfate groups from human corneal epithelial (HCE) cells by heparinase III and sodium chlorate treatments, respectively, reduced HAdV-D37 binding to cells. Remarkably, removal of HS by heparinase III enhanced the virus infection. Our results suggest that interaction of HAdV-D37 with sulfated GAGs in secretions and on plasma membranes prevents/delays the virus binding to SA-containing receptors and inhibits subsequent infection. We also found abundant HS in the basement membrane of the human corneal epithelium, which may act as a barrier to sub-epithelial infection. Collectively, our findings provide novel insights into the role of GAGs as viral decoy receptors and highlight the therapeutic potential of GAGs and/or GAG-mimetics in HAdV-D37 infection.

Place, publisher, year, edition, pages
MDPI, 2019
Keywords
adenovirus, antiviral drugs, cellular receptor, decoy receptor, epidemic keratoconjunctivitis, glycosaminoglycan, tropism
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:umu:diva-158515 (URN)10.3390/v11030247 (DOI)000464389700003 ()30871026 (PubMedID)
Funder
Knut and Alice Wallenberg Foundation, KAW 2013.0019Swedish Research Council, 2018-02401Västerbotten County CouncilWellcome trust, 099197MA
Available from: 2019-04-29 Created: 2019-04-29 Last updated: 2019-08-28Bibliographically approved
Chandra, N. (2019). The glycobiology of human adenovirus infections: implications for tropism and treatment. (Doctoral dissertation). Umeå: Umeå University
Open this publication in new window or tab >>The glycobiology of human adenovirus infections: implications for tropism and treatment
2019 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Human adenoviruses (HAdVs) are common human pathogens, causing gastrointestinal, ocular, and respiratory infections on a regular basis. Epidemic keratoconjunctivitis (EKC) is a severe ocular infection for which no approved antivirals are available. HAdV-D37 is one of the causative agents of EKC and uses sialic acid (SA)-containing glycans as cellular receptors. HAdV-D37 interacts with SA via the knob domain of the trimeric virus fiber protein, containing three SA-binding sites. HAdV-D37 also bind to glycosaminoglycans (GAGs), but the outcome of this interaction remains unknown. Here, using biochemical and cell-based assays, the impact of GAGs on HAdV-D37 infection (paper I) was investigated. We found that HAdV-D37 interacts with both soluble and cell-surface sulfated GAGs via the knob domain of the viral fiber protein. Remarkably, removal of heparan sulfate (HS; a type of GAG) from human corneal epithelial (HCE) cells by heparinase III enhanced HAdV-D37 infection. We propose that sulfated GAGs in bodily secretions and on plasma membranes function as decoy receptors that prevent the virus from binding to SA-containing receptors and inhibit subsequent virus infection. We also found abundant HS in the basement membrane of the human corneal epithelium. We suggest that this layer of HS functions as a barrier to sub-epithelial infection of HAdV-D37. Based on this finding, we hypothesized that GAG-mimetics may act as artificial decoy receptors and inhibit HAdV-D37 infection. Here, the antiviral effect of suramin (a known GAG-mimetic) and its analogs against HAdV-D37 (paper II) was evaluated. Interestingly, all compounds displayed antiviral effects by inhibiting the binding of HAdV-D37 to HCE cells. The antiviral effect of suramin was HAdV species-specific. We report for the first time that virus binding to cell-surface decoy receptor constitutes a potential target for antiviral drug development.

HAdVs are the major cause of infectious conjunctivitis, constituting up to 75% of all conjunctivitis cases worldwide. Species B HAdV type 3 (HAdV-B3) causes pharyngoconjunctival fever (PCF), whereas HAdV-D8, -D37, and -D64 cause EKC. Recently, HAdV-D53, -D54, and -D56 have emerged as new EKC-causing agents. HAdV-E4 causes both PCF and EKC. SA-containing glycans have been established as cellular receptors for HAdV-D37. By means of cell-based assays, we investigated if ocular HAdVs other than HAdV-D37 also use SA-containing glycans as receptors on HCE cells (paper III). It was found that SA-containing glycans function as cellular receptors for five (HAdV-D8, -D37, -D53, -D54, and -D64) out of six EKC-causing species D HAdVs. We showed that these viruses interact with SAs via the knob domain of the viral fiber protein. HAdV-E4 and -D56 infection of cells was independent of SAs. Surprisingly, HCE cells were completely refractory to HAdV-B3 infection. A trivalent sialic acid (TSA) derivative ME0462 (compound 17a in paper II), designed to bind to SA-binding sites on HAdV-D37 fiber knob, also showed potent antiviral activity against several EKC-causing HAdVs. This suggests that ME0462 can be used as a broad-spectrum antiviral against known and emerging EKC-causing HAdVs. Surface plasmon resonance (SPR) analysis confirmed a direct interaction between ME0462 and fiber knobs of EKC-causing HAdVs.

Recently, a TSA derivative (ME0322; designed to bind to SA-binding sites on HAdV-D37 fiber knob) was shown potent antiviral against HAdV-D37 in vitro. To improve the antiviral potency of this compound, six new TSA derivatives were synthesized and their inhibitory effects were evaluated against HAdV-D37 (paper IV). Interestingly, the best compound 17a was found approximately three orders of magnitude more potent (IC50 (binding) = 1.4 nM, IC50 (infection) = 2.9 nM) than ME0322 (IC50 in µM range). SPR data showed that HAdV-D37 fiber knob binds to TSA compounds with high affinities. Structural data revealed the trivalent binding mode of all newly synthesized TSA compounds to HAdV-D37 fiber knob. Ophthalmic toxicity of compound 17a (best compound) was also investigated in rabbits without any sign of toxicity.

HAdV-D36 is a member of species D HAdV and has the ability to infect a broad range of animals, which is unusual for HAdVs. Another remarkable feature of HAdV-D36 is that this virus induces obesity in experimental animals. Several epidemiological studies highlighted a link between HAdV-D36 and human obesity. There is no information about the cellular receptor usage by HAdV-D36. Using structural biology and cell-based approaches, we investigated the cellular receptor(s) for HAdV-D36 (paper V).  We show that HAdV-D36 attaches to host cells (via the fiber knob) using the coxsackie and adenovirus receptor (CAR), SA-containing glycans, and one or more unknown proteins or glycoproteins. Using glycan microarray, we found that HAdV-D36 displays binding preference to a rare SA-variant: 4-O,5-N-diacetylneuraminic acid (Neu4,5Ac2), over the more common SA (in humans) i.e. 5-N-acetylneuraminic acid (Neu5Ac). Structural analysis of HAdV-D36 fiber knob:Neu4,5Ac2 complex explained this preference. To date, Neu4,5Achas not been detected in humans, although it is synthesized by many domestic and livestock animals. Our results indicate that HAdV-D36 has evolved to utilize a specialized set of cellular receptors that coincide with a unique host range and pathogenicity profile.

These studies provide insights into multiple roles of glycans in HAdV infection cycle and highlight the therapeutic potential of glycans/glycan-mimetics in HAdV-D37 infection.

Place, publisher, year, edition, pages
Umeå: Umeå University, 2019. p. 111
Series
Umeå University medical dissertations, ISSN 0346-6612 ; 2046
Keywords
denovirus, cellular receptor, glycosaminoglycan, sialic acid, decoy receptors, tropism, epidemic keratoconjunctivitis, antiviral drugs, GAG-mimetic, obesity
National Category
Microbiology in the medical area Infectious Medicine
Research subject
Medical Virology
Identifiers
urn:nbn:se:umu:diva-162709 (URN)978-91-7855-104-0 (ISBN)
Public defence
2019-09-20, Sal E04, byggnad 6A, Umeå, 09:00 (English)
Opponent
Supervisors
Note

I publikationen felaktigt ISBN: 978-91-7601-940-5

Available from: 2019-08-30 Created: 2019-08-26 Last updated: 2019-09-04Bibliographically approved
Duffy, M. R., Alonso-Padilla, J., John, L., Chandra, N., Khan, S., Ballmann, M. Z., . . . Lemckert, A. (2018). Generation and characterization of a novel candidate gene therapy and vaccination vector based on human species D adenovirus type 56. Journal of General Virology, 99, 135-147
Open this publication in new window or tab >>Generation and characterization of a novel candidate gene therapy and vaccination vector based on human species D adenovirus type 56
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2018 (English)In: Journal of General Virology, ISSN 0022-1317, E-ISSN 1465-2099, Vol. 99, p. 135-147Article in journal (Refereed) Published
Abstract [en]

The vectorization of rare human adenovirus (HAdV) types will widen our knowledge of this family and their interaction with cells, tissues and organs. In this study we focus on HAdV-56, a member of human Ad species D, and create ease-of-use cloning systems to generate recombinant HAdV-56 vectors carrying foreign genes. We present in vitro transduction profiles for HAdV-56 in direct comparison to the most commonly used HAdV-5-based vector. In vivo characterizations demonstrate that when it is delivered intravenously (i.v.) HAdV-56 mainly targets the spleen and, to a lesser extent, the lungs, whilst largely bypassing liver transduction in mice. HAdV-56 triggered robust inflammatory and cellular immune responses, with higher induction of IFNγ, TNFα, IL5, IL6, IP10, MCP1 and MIG1 compared to HAdV-5 following i.v. administration. We also investigated its potential as a vaccine vector candidate by performing prime immunizations in mice with HAdV-56 encoding luciferase (HAdV-56-Luc). Direct comparisons were made to HAdV-26, a highly potent human vaccine vector currently in phase II clinical trials. HAdV-56-Luc induced luciferase 'antigen'-specific IFNγ-producing cells and anti-HAdV-56 neutralizing antibodies in Balb/c mice, demonstrating a near identical profile to that of HAdV-26. Taken together, the data presented provides further insight into human Ad receptor/co-receptor usage, and the first report on HAdV-56 vectors and their potential for gene therapy and vaccine applications.

Keywords
adenovirus, HAdV-56, gene therapy, vaccine vector
National Category
Microbiology in the medical area Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:umu:diva-143059 (URN)10.1099/jgv.0.000978 (DOI)000431016500015 ()29154744 (PubMedID)
Available from: 2017-12-14 Created: 2017-12-14 Last updated: 2018-06-09Bibliographically approved
Lasswitz, L., Chandra, N., Arnberg, N. & Gerold, G. (2018). Glycomics and Proteomics Approaches to Investigate Early Adenovirus-Host Cell Interactions. Journal of Molecular Biology, 430(13), 1863-1882
Open this publication in new window or tab >>Glycomics and Proteomics Approaches to Investigate Early Adenovirus-Host Cell Interactions
2018 (English)In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 430, no 13, p. 1863-1882Article in journal (Refereed) Published
Abstract [en]

Adenoviruses as most viruses rely on glycan and protein interactions to attach to and enter susceptible host cells. The Adenoviridae family comprises more than 80 human types and they differ in their attachment factor and receptor usage, which likely contributes to the diverse tropism of the different types. In the past years, methods to systematically identify glycan and protein interactions have advanced. In particular sensitivity, speed and coverage of mass spectrometric analyses allow for high-throughput identification of glycans and peptides separated by liquid chromatography. Also, developments in glycan microarray technologies have led to targeted, high-throughput screening and identification of glycan-based receptors. The mapping of cell surface interactions of the diverse adenovirus types has implications for cell, tissue, and species tropism as well as drug development. Here we review known adenovirus interactions with glycan- and protein-based receptors, as well as glycomics and proteomics strategies to identify yet elusive virus receptors and attachment factors. We finally discuss challenges, bottlenecks, and future research directions in the field of non-enveloped virus entry into host cells.

Keywords
adenovirus, glycomis, host cell interactions, proteomics, virus entry
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy) Microbiology in the medical area
Identifiers
urn:nbn:se:umu:diva-148152 (URN)10.1016/j.jmb.2018.04.039 (DOI)000436224800004 ()29746851 (PubMedID)2-s2.0-85047296623 (Scopus ID)
Available from: 2018-05-29 Created: 2018-05-29 Last updated: 2018-09-28Bibliographically approved
Caraballo, R., Saleeb, M., Bauer, J., Liaci, A.-M., Chandra, N., Storm, R. J., . . . Elofsson, M. (2015). Triazole linker-based trivalent sialic acid inhibitors of adenovirus type 37 infection of human corneal epithelial cells. Organic and biomolecular chemistry, 13(35), 9194-9205
Open this publication in new window or tab >>Triazole linker-based trivalent sialic acid inhibitors of adenovirus type 37 infection of human corneal epithelial cells
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2015 (English)In: Organic and biomolecular chemistry, ISSN 1477-0520, E-ISSN 1477-0539, Vol. 13, no 35, p. 9194-9205Article in journal (Refereed) Published
Abstract [en]

Adenovirus type 37 (Ad37) is one of the principal agents responsible for epidemic keratoconjunctivitis (EKC), a severe ocular infection that remains without any available treatment. Recently, a trivalent sialic acid derivative (ME0322, Angew. Chem. Int. Ed., 2011, 50, 6519) was shown to function as a highly potent inhibitor of Ad37, efficiently preventing the attachment of the virion to the host cells and subsequent infection. Here, new trivalent sialic acid derivatives were designed, synthesized and their inhibitory properties against Ad37 infection of the human corneal epithelial cells were investigated. In comparison to ME0322, the best compound (17a) was found to be over three orders of magnitude more potent in a cell-attachment assay (IC50 = 1.4 nM) and about 140 times more potent in a cell-infection assay (IC50 = 2.9nM). X-ray crystallographic analysis demonstrated a trivalent binding mode of all compounds to the Ad37 fiber knob. For the most potent compound ophthalmic toxicity in rabbits was investigated and it was concluded that repeated eye administration did not cause any adverse effects.

National Category
Microbiology in the medical area
Identifiers
urn:nbn:se:umu:diva-100014 (URN)10.1039/C5OB01025J (DOI)000360115100007 ()
Available from: 2015-02-18 Created: 2015-02-18 Last updated: 2019-08-28Bibliographically approved
Majhen, D., Calderon, H., Chandra, N., Alberto Fajardo, C., Rajan, A., Alemany, R. & Custers, J. (2014). Adenovirus-based vaccines for fighting infectious diseases and cancer: progress in the field. Human Gene Therapy, 25(4), 301-317
Open this publication in new window or tab >>Adenovirus-based vaccines for fighting infectious diseases and cancer: progress in the field
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2014 (English)In: Human Gene Therapy, ISSN 1043-0342, E-ISSN 1557-7422, Vol. 25, no 4, p. 301-317Article, review/survey (Refereed) Published
Abstract [en]

The field of adenovirology is undergoing rapid change in response to increasing appreciation of the potential advantages of adenoviruses as the basis for new vaccines and as vectors for gene and cancer therapy. Substantial knowledge and understanding of adenoviruses at a molecular level has made their manipulation for use as vaccines and therapeutics relatively straightforward in comparison with other viral vectors. In this review we summarize the structure and life cycle of the adenovirus and focus on the use of adenovirus-based vectors in vaccines against infectious diseases and cancers. Strategies to overcome the problem of preexisting antiadenovirus immunity, which can hamper the immunogenicity of adenovirus-based vaccines, are discussed. When armed with tumor-associated antigens, replication-deficient and oncolytic adenoviruses can efficiently activate an antitumor immune response. We present concepts on how to use adenoviruses as therapeutic cancer vaccines and consider some of the strategies used to further improve antitumor immune responses. Studies that explore the prospect of adenoviruses as vaccines against infectious diseases and cancer are underway, and here we give an overview of the latest developments.

Place, publisher, year, edition, pages
Mary Ann Liebert, 2014
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy) Medical Genetics
Identifiers
urn:nbn:se:umu:diva-89232 (URN)10.1089/hum.2013.235 (DOI)000334915500006 ()
Available from: 2014-05-26 Created: 2014-05-26 Last updated: 2018-06-07Bibliographically approved
Liaci, A., Chandra, N., Munender, S., Liu, Y., Pfenning, V., Bachmann, P., . . . Stehle, T.Primary attachment receptors of human adenovirus type 36.
Open this publication in new window or tab >>Primary attachment receptors of human adenovirus type 36
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(English)Manuscript (preprint) (Other academic)
National Category
Microbiology in the medical area
Identifiers
urn:nbn:se:umu:diva-162795 (URN)
Available from: 2019-08-28 Created: 2019-08-28 Last updated: 2019-08-28
Organisations
Identifiers
ORCID iD: ORCID iD iconorcid.org/0000-0002-8551-3256

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