Glycosylation and antiproliferative activity of hyperglycosylated IFN-α2 potentiate HEK293 cells as biofactories

Autores
Gugliotta, Agustina; Ceaglio, Natalia Analia; Raud, Brenda; Forno, Angela Guillermina; Mauro, Laura Valeria; Kratje, Ricardo Bertoldo; Oggero Eberhardt, Marcos Rafael
Año de publicación
2017
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Both CHO and HEK cells are interesting hosts for the production of biotherapeutics due to their ability to introduce post-translational modifications such as glycosylation. Even though oligosaccharide structures attached to proteins are conserved among eukaryotes, many differences have been found between therapeutic glycoproteins expressed in hamster and human derived cells. In this work, a hyperglycosylated IFN-α2b mutein (IFN4N) was produced in CHO and HEK cell lines and an extensive characterization of their properties was performed. IFN4NCHO exhibited a higher average molecular mass and more acidic isoforms compared to IFN4NHEK. In agreement with these results, a 2-times higher sialic acid content was found for IFN4NCHO in comparison with the HEK-derived protein. This result was in agreement with monosaccharide quantification and glycan's analysis using WAX chromatography and HILIC coupled to mass spectrometry; all methods supported the existence of highly sialylated and also branched structures for IFN4NCHO glycans, in contrast with smaller and truncated structures among IFN4NHEK glycans. Unexpectedly, those remarkable differences in the glycosylation pattern had not a considerable impact on the clearance rate of both molecules in rats. In fact, although IFN4NHEK reached maximum plasma concentration 3-times faster than IFN4NCHO, their elimination profile did not differ significantly. Also, despite the in vitro antiviral specific biological activity of both proteins was the same, IFN4NHEK was more efficient as an antiproliferative agent in different tumor-derived cell lines. Accordingly, IFN4NHEK showed a higher in vivo antitumor activity in animal models. Our results show the importance of an appropriate host selection to set up a bioprocess and potentiate the use of HEK293 cells for the production of a new hyperglycosylated protein-based pharmaceutical.
Fil: Gugliotta, Agustina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Cultivos Celulares; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina
Fil: Ceaglio, Natalia Analia. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Cultivos Celulares; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina
Fil: Raud, Brenda. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Cultivos Celulares; Argentina
Fil: Forno, Angela Guillermina. Universidad Nacional del Litoral; Argentina. Zelltek; Argentina
Fil: Mauro, Laura Valeria. Zelltek; Argentina
Fil: Kratje, Ricardo Bertoldo. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Cultivos Celulares; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina
Fil: Oggero Eberhardt, Marcos Rafael. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Cultivos Celulares; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina
Materia
Antitumoral Activity
Cho Cells
Glycoproteins
Hek293 Cells
Hyperglycosylated Ifn-Α2b
N-Glycosylation
Pharmacokinetics
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-nd/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/70926
Acceder
id CONICETDig_78c5ffc8efb05325924deda1127dc834
oai_identifier_str oai:ri.conicet.gov.ar:11336/70926
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Glycosylation and antiproliferative activity of hyperglycosylated IFN-α2 potentiate HEK293 cells as biofactoriesGugliotta, AgustinaCeaglio, Natalia AnaliaRaud, BrendaForno, Angela GuillerminaMauro, Laura ValeriaKratje, Ricardo BertoldoOggero Eberhardt, Marcos RafaelAntitumoral ActivityCho CellsGlycoproteinsHek293 CellsHyperglycosylated Ifn-Α2bN-GlycosylationPharmacokineticshttps://purl.org/becyt/ford/3.4https://purl.org/becyt/ford/3Both CHO and HEK cells are interesting hosts for the production of biotherapeutics due to their ability to introduce post-translational modifications such as glycosylation. Even though oligosaccharide structures attached to proteins are conserved among eukaryotes, many differences have been found between therapeutic glycoproteins expressed in hamster and human derived cells. In this work, a hyperglycosylated IFN-α2b mutein (IFN4N) was produced in CHO and HEK cell lines and an extensive characterization of their properties was performed. IFN4NCHO exhibited a higher average molecular mass and more acidic isoforms compared to IFN4NHEK. In agreement with these results, a 2-times higher sialic acid content was found for IFN4NCHO in comparison with the HEK-derived protein. This result was in agreement with monosaccharide quantification and glycan's analysis using WAX chromatography and HILIC coupled to mass spectrometry; all methods supported the existence of highly sialylated and also branched structures for IFN4NCHO glycans, in contrast with smaller and truncated structures among IFN4NHEK glycans. Unexpectedly, those remarkable differences in the glycosylation pattern had not a considerable impact on the clearance rate of both molecules in rats. In fact, although IFN4NHEK reached maximum plasma concentration 3-times faster than IFN4NCHO, their elimination profile did not differ significantly. Also, despite the in vitro antiviral specific biological activity of both proteins was the same, IFN4NHEK was more efficient as an antiproliferative agent in different tumor-derived cell lines. Accordingly, IFN4NHEK showed a higher in vivo antitumor activity in animal models. Our results show the importance of an appropriate host selection to set up a bioprocess and potentiate the use of HEK293 cells for the production of a new hyperglycosylated protein-based pharmaceutical.Fil: Gugliotta, Agustina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Cultivos Celulares; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; ArgentinaFil: Ceaglio, Natalia Analia. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Cultivos Celulares; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; ArgentinaFil: Raud, Brenda. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Cultivos Celulares; ArgentinaFil: Forno, Angela Guillermina. Universidad Nacional del Litoral; Argentina. Zelltek; ArgentinaFil: Mauro, Laura Valeria. Zelltek; ArgentinaFil: Kratje, Ricardo Bertoldo. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Cultivos Celulares; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; ArgentinaFil: Oggero Eberhardt, Marcos Rafael. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Cultivos Celulares; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; ArgentinaElsevier Science2017-03info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/70926Gugliotta, Agustina; Ceaglio, Natalia Analia; Raud, Brenda; Forno, Angela Guillermina; Mauro, Laura Valeria; et al.; Glycosylation and antiproliferative activity of hyperglycosylated IFN-α2 potentiate HEK293 cells as biofactories; Elsevier Science; European Journal Of Pharmaceutics And Biopharmaceutics; 112; 3-2017; 119-1310939-6411CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/doi/10.1016/j.ejpb.2016.11.012info:eu-repo/semantics/altIdentifier/url/https://www.sciencedirect.com/science/article/pii/S0939641116308098info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-nd/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-10-22T11:04:45Zoai:ri.conicet.gov.ar:11336/70926instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-10-22 11:04:46.193CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Glycosylation and antiproliferative activity of hyperglycosylated IFN-α2 potentiate HEK293 cells as biofactories
title Glycosylation and antiproliferative activity of hyperglycosylated IFN-α2 potentiate HEK293 cells as biofactories
spellingShingle Glycosylation and antiproliferative activity of hyperglycosylated IFN-α2 potentiate HEK293 cells as biofactories
Gugliotta, Agustina
Antitumoral Activity
Cho Cells
Glycoproteins
Hek293 Cells
Hyperglycosylated Ifn-Α2b
N-Glycosylation
Pharmacokinetics
title_short Glycosylation and antiproliferative activity of hyperglycosylated IFN-α2 potentiate HEK293 cells as biofactories
title_full Glycosylation and antiproliferative activity of hyperglycosylated IFN-α2 potentiate HEK293 cells as biofactories
title_fullStr Glycosylation and antiproliferative activity of hyperglycosylated IFN-α2 potentiate HEK293 cells as biofactories
title_full_unstemmed Glycosylation and antiproliferative activity of hyperglycosylated IFN-α2 potentiate HEK293 cells as biofactories
title_sort Glycosylation and antiproliferative activity of hyperglycosylated IFN-α2 potentiate HEK293 cells as biofactories
dc.creator.none.fl_str_mv Gugliotta, Agustina
Ceaglio, Natalia Analia
Raud, Brenda
Forno, Angela Guillermina
Mauro, Laura Valeria
Kratje, Ricardo Bertoldo
Oggero Eberhardt, Marcos Rafael
author Gugliotta, Agustina
author_facet Gugliotta, Agustina
Ceaglio, Natalia Analia
Raud, Brenda
Forno, Angela Guillermina
Mauro, Laura Valeria
Kratje, Ricardo Bertoldo
Oggero Eberhardt, Marcos Rafael
author_role author
author2 Ceaglio, Natalia Analia
Raud, Brenda
Forno, Angela Guillermina
Mauro, Laura Valeria
Kratje, Ricardo Bertoldo
Oggero Eberhardt, Marcos Rafael
author2_role author
author
author
author
author
author
dc.subject.none.fl_str_mv Antitumoral Activity
Cho Cells
Glycoproteins
Hek293 Cells
Hyperglycosylated Ifn-Α2b
N-Glycosylation
Pharmacokinetics
topic Antitumoral Activity
Cho Cells
Glycoproteins
Hek293 Cells
Hyperglycosylated Ifn-Α2b
N-Glycosylation
Pharmacokinetics
purl_subject.fl_str_mv https://purl.org/becyt/ford/3.4
https://purl.org/becyt/ford/3
dc.description.none.fl_txt_mv Both CHO and HEK cells are interesting hosts for the production of biotherapeutics due to their ability to introduce post-translational modifications such as glycosylation. Even though oligosaccharide structures attached to proteins are conserved among eukaryotes, many differences have been found between therapeutic glycoproteins expressed in hamster and human derived cells. In this work, a hyperglycosylated IFN-α2b mutein (IFN4N) was produced in CHO and HEK cell lines and an extensive characterization of their properties was performed. IFN4NCHO exhibited a higher average molecular mass and more acidic isoforms compared to IFN4NHEK. In agreement with these results, a 2-times higher sialic acid content was found for IFN4NCHO in comparison with the HEK-derived protein. This result was in agreement with monosaccharide quantification and glycan's analysis using WAX chromatography and HILIC coupled to mass spectrometry; all methods supported the existence of highly sialylated and also branched structures for IFN4NCHO glycans, in contrast with smaller and truncated structures among IFN4NHEK glycans. Unexpectedly, those remarkable differences in the glycosylation pattern had not a considerable impact on the clearance rate of both molecules in rats. In fact, although IFN4NHEK reached maximum plasma concentration 3-times faster than IFN4NCHO, their elimination profile did not differ significantly. Also, despite the in vitro antiviral specific biological activity of both proteins was the same, IFN4NHEK was more efficient as an antiproliferative agent in different tumor-derived cell lines. Accordingly, IFN4NHEK showed a higher in vivo antitumor activity in animal models. Our results show the importance of an appropriate host selection to set up a bioprocess and potentiate the use of HEK293 cells for the production of a new hyperglycosylated protein-based pharmaceutical.
Fil: Gugliotta, Agustina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Cultivos Celulares; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina
Fil: Ceaglio, Natalia Analia. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Cultivos Celulares; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina
Fil: Raud, Brenda. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Cultivos Celulares; Argentina
Fil: Forno, Angela Guillermina. Universidad Nacional del Litoral; Argentina. Zelltek; Argentina
Fil: Mauro, Laura Valeria. Zelltek; Argentina
Fil: Kratje, Ricardo Bertoldo. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Cultivos Celulares; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina
Fil: Oggero Eberhardt, Marcos Rafael. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Cultivos Celulares; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina
description Both CHO and HEK cells are interesting hosts for the production of biotherapeutics due to their ability to introduce post-translational modifications such as glycosylation. Even though oligosaccharide structures attached to proteins are conserved among eukaryotes, many differences have been found between therapeutic glycoproteins expressed in hamster and human derived cells. In this work, a hyperglycosylated IFN-α2b mutein (IFN4N) was produced in CHO and HEK cell lines and an extensive characterization of their properties was performed. IFN4NCHO exhibited a higher average molecular mass and more acidic isoforms compared to IFN4NHEK. In agreement with these results, a 2-times higher sialic acid content was found for IFN4NCHO in comparison with the HEK-derived protein. This result was in agreement with monosaccharide quantification and glycan's analysis using WAX chromatography and HILIC coupled to mass spectrometry; all methods supported the existence of highly sialylated and also branched structures for IFN4NCHO glycans, in contrast with smaller and truncated structures among IFN4NHEK glycans. Unexpectedly, those remarkable differences in the glycosylation pattern had not a considerable impact on the clearance rate of both molecules in rats. In fact, although IFN4NHEK reached maximum plasma concentration 3-times faster than IFN4NCHO, their elimination profile did not differ significantly. Also, despite the in vitro antiviral specific biological activity of both proteins was the same, IFN4NHEK was more efficient as an antiproliferative agent in different tumor-derived cell lines. Accordingly, IFN4NHEK showed a higher in vivo antitumor activity in animal models. Our results show the importance of an appropriate host selection to set up a bioprocess and potentiate the use of HEK293 cells for the production of a new hyperglycosylated protein-based pharmaceutical.
publishDate 2017
dc.date.none.fl_str_mv 2017-03
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/70926
Gugliotta, Agustina; Ceaglio, Natalia Analia; Raud, Brenda; Forno, Angela Guillermina; Mauro, Laura Valeria; et al.; Glycosylation and antiproliferative activity of hyperglycosylated IFN-α2 potentiate HEK293 cells as biofactories; Elsevier Science; European Journal Of Pharmaceutics And Biopharmaceutics; 112; 3-2017; 119-131
0939-6411
CONICET Digital
CONICET
url http://hdl.handle.net/11336/70926
identifier_str_mv Gugliotta, Agustina; Ceaglio, Natalia Analia; Raud, Brenda; Forno, Angela Guillermina; Mauro, Laura Valeria; et al.; Glycosylation and antiproliferative activity of hyperglycosylated IFN-α2 potentiate HEK293 cells as biofactories; Elsevier Science; European Journal Of Pharmaceutics And Biopharmaceutics; 112; 3-2017; 119-131
0939-6411
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.1016/j.ejpb.2016.11.012
info:eu-repo/semantics/altIdentifier/url/https://www.sciencedirect.com/science/article/pii/S0939641116308098
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-nd/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-nd/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
application/pdf
application/pdf
dc.publisher.none.fl_str_mv Elsevier Science
publisher.none.fl_str_mv Elsevier Science
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
collection CONICET Digital (CONICET)
instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
_version_ 1846781312006881280
score 12.982451

Publicaciones similares