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[{"key": "dc.contributor.advisor", "value": "Ja\u00e9n, Karim", "language": "", "element": "contributor", "qualifier": "advisor", "schema": "dc"}, {"key": "dc.contributor.advisor", "value": "Ju\u00e1rez-Osorio, Mariana", "language": "", "element": "contributor", "qualifier": "advisor", "schema": "dc"}, {"key": "dc.contributor.advisor", "value": "Ihalainen, Janne", "language": "", "element": "contributor", "qualifier": "advisor", "schema": "dc"}, {"key": "dc.contributor.author", "value": "Zhurina, Anastasiia", "language": "", "element": "contributor", "qualifier": "author", "schema": "dc"}, {"key": "dc.date.accessioned", "value": "2024-06-26T07:39:28Z", "language": null, "element": "date", "qualifier": "accessioned", "schema": "dc"}, {"key": "dc.date.available", "value": "2024-06-26T07:39:28Z", "language": null, "element": "date", "qualifier": "available", "schema": "dc"}, {"key": "dc.date.issued", "value": "2024", "language": "", "element": "date", "qualifier": "issued", "schema": "dc"}, {"key": "dc.identifier.uri", "value": "https://jyx.jyu.fi/handle/123456789/96162", "language": null, "element": "identifier", "qualifier": "uri", "schema": "dc"}, {"key": "dc.description.abstract", "value": "Sugar modifications, in particular, N-glycans, largely determine protein quality, functionality, and, in case of therapeutic biomolecules, safety. Therapeutic monoclonal antibodies (mAbs) generate immune response through effector functions, for which they require complex N-glycans with high levels of galactose, sialic acids, and lack of core fucose, often unachievable in commercial production yielding suboptimal and heterogenous glycosylation pattern. Current glycoengineering strategies successfully tailor N-glycan composition on mAbs, yet fail to improve glycoprofile homogeneity. In this work, a recently developed cell-free in vitro glycoengineering approach was evaluated for generation of homogenous fully galactosylated N-glycoforms on therapeutic effector antibody Rituximab, characterized by low levels of galactose and glycan homogeneity in the originally marketed product. With an aim of targeting commercial mAbs, an industry-oriented protein production and glycoengineering strategy was designed on the basis of Lepidopteran insect cells. Rituximab was expressed in baculovirus-infected HighFive\u2122 cells to obtain highly uniform starting \nN-glycoprofile, which was sequentially modified by recombinant human \u03b2-1,2 N-acetylglucosamintransferases I and II (MGAT1\u0394TM and MGAT2\u0394TM), and \u03b2-1,4-galactosyltransferase (GalT\u0394TM). The enzymatic cascade remodelled simple insect cell oligosaccharides on protein A-immobilized mAb species into complex human-like structures with a near 100% conversion rate. Recombinant enzyme production was tested in HighFive\u2122 host, for which a functional expression protocol was developed, and a promising purification approach incorporating solubilizers was proposed. Production of Rituximab in HighFive\u2122 cells was challenged by poor mAb assembly despite optimization of cell culture conditions. The results indicated a need for significant effort in improving insect cell-based protein production strategy to achieve higher enzyme and antibody yields. With suggested directions for further research, assessed in vitro glycoengineering platform can become an effective toolbox for generating high-efficacy therapeutic mAbs in industrial settings.", "language": "en", "element": "description", "qualifier": "abstract", "schema": "dc"}, {"key": "dc.description.provenance", "value": "Submitted by Miia Hakanen (mihakane@jyu.fi) on 2024-06-26T07:39:28Z\nNo. of bitstreams: 0", "language": "en", "element": "description", "qualifier": "provenance", "schema": "dc"}, {"key": "dc.description.provenance", "value": "Made available in DSpace on 2024-06-26T07:39:28Z (GMT). No. of bitstreams: 0\n Previous issue date: 2024", "language": "en", "element": "description", "qualifier": "provenance", "schema": "dc"}, {"key": "dc.format.extent", "value": "56", "language": "", "element": "format", "qualifier": "extent", "schema": "dc"}, {"key": "dc.language.iso", "value": "eng", "language": null, "element": "language", "qualifier": "iso", "schema": "dc"}, {"key": "dc.rights", "value": "In Copyright", "language": "en", "element": "rights", "qualifier": null, "schema": "dc"}, {"key": "dc.subject.other", "value": "effector functions", "language": "", "element": "subject", "qualifier": "other", "schema": "dc"}, {"key": "dc.subject.other", "value": "glycoengineering", "language": "", "element": "subject", "qualifier": "other", "schema": "dc"}, {"key": "dc.subject.other", "value": "homogenous glycoprofile", "language": "", "element": "subject", "qualifier": "other", "schema": "dc"}, {"key": "dc.subject.other", "value": "Lepidopteran insect cells", "language": "", "element": "subject", "qualifier": "other", "schema": "dc"}, {"key": "dc.subject.other", "value": "monoclonal antibody", "language": "", "element": "subject", "qualifier": "other", "schema": "dc"}, {"key": "dc.title", "value": "In vitro glycoengineering of a monoclonal antibody produced in insect cells", "language": "", "element": "title", "qualifier": null, "schema": "dc"}, {"key": "dc.type", "value": "master thesis", "language": null, "element": "type", "qualifier": null, "schema": "dc"}, {"key": "dc.identifier.urn", "value": "URN:NBN:fi:jyu-202406265006", "language": null, "element": "identifier", "qualifier": "urn", "schema": "dc"}, {"key": "dc.type.ontasot", "value": "Master\u2019s thesis", "language": "en", "element": "type", "qualifier": "ontasot", "schema": "dc"}, {"key": "dc.type.ontasot", "value": "Pro gradu -tutkielma", "language": "fi", "element": "type", "qualifier": "ontasot", "schema": "dc"}, {"key": "dc.contributor.faculty", "value": "Matemaattis-luonnontieteellinen tiedekunta", "language": "fi", "element": "contributor", "qualifier": "faculty", "schema": "dc"}, {"key": "dc.contributor.faculty", "value": "Faculty of Sciences", "language": "en", "element": "contributor", "qualifier": "faculty", "schema": "dc"}, {"key": "dc.contributor.department", "value": "Bio- ja ymp\u00e4rist\u00f6tieteiden laitos", "language": "fi", "element": "contributor", "qualifier": "department", "schema": "dc"}, {"key": "dc.contributor.department", "value": "Department of Biological and Environmental Science", "language": "en", "element": "contributor", "qualifier": "department", "schema": "dc"}, {"key": "dc.contributor.organization", "value": "Jyv\u00e4skyl\u00e4n yliopisto", "language": "fi", "element": "contributor", "qualifier": "organization", "schema": "dc"}, {"key": "dc.contributor.organization", "value": "University of Jyv\u00e4skyl\u00e4", "language": "en", "element": "contributor", "qualifier": "organization", "schema": "dc"}, {"key": "dc.subject.discipline", "value": "Solu- ja molekyylibiologia", "language": "fi", "element": "subject", "qualifier": "discipline", "schema": "dc"}, {"key": "dc.subject.discipline", "value": "Cell and molecular biology", "language": "en", "element": "subject", "qualifier": "discipline", "schema": "dc"}, {"key": "yvv.contractresearch.collaborator", "value": "community", "language": "", "element": "contractresearch", "qualifier": "collaborator", "schema": "yvv"}, {"key": "yvv.contractresearch.funding", "value": "0", "language": "", "element": "contractresearch", "qualifier": "funding", "schema": "yvv"}, {"key": "yvv.contractresearch.initiative", "value": "student", "language": "", "element": "contractresearch", "qualifier": "initiative", "schema": "yvv"}, {"key": "dc.type.coar", "value": "http://purl.org/coar/resource_type/c_bdcc", "language": null, "element": "type", "qualifier": "coar", "schema": "dc"}, {"key": "dc.rights.accesslevel", "value": "openAccess", "language": null, "element": "rights", "qualifier": "accesslevel", "schema": "dc"}, {"key": "dc.type.publication", "value": "masterThesis", "language": null, "element": "type", "qualifier": "publication", "schema": "dc"}, {"key": "dc.subject.oppiainekoodi", "value": "4013", "language": "", "element": "subject", "qualifier": "oppiainekoodi", "schema": "dc"}, {"key": "dc.rights.url", "value": "https://rightsstatements.org/page/InC/1.0/", "language": null, "element": "rights", "qualifier": "url", "schema": "dc"}]
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