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Construction, Purification, and Characterization of a Homodimeric Granulocyte Colony-Stimulating Factor

Author:
Mickiene, Gitana, Dalgediene, Indre, Dapkunas, Zilvinas, Zvirblis, Gintautas, Pesliakas, Henrikas, Kaupinis, Algirdas, Valius, Mindaugas, Mistiniene, Edita, Pleckaityte, Milda
Source:
Molecular biotechnology 2017 v.59 no.9-10 pp. 374-384
ISSN:
1073-6085
Subject:
bioaccumulation, blood serum, chemical bonding, drugs, granulocyte colony-stimulating factor, half life, immune response, neutrophils, pharmacokinetics, polyethylene glycol, proteins
Abstract:
Granulocyte colony-stimulating factor (G-CSF) has found widespread clinical application, and modified forms with improved biopharmaceutical properties have been marketed as well. PEGylation, the covalent modification of G-CSF with polyethylene glycol (PEG), has a beneficial effect on drug properties, but there are concerns connected to the immunogenicity of PEGylated compounds and bioaccumulation of the synthetic polymer. To overcome challenges connected with chemical modifications, we developed fusion proteins composed of two G-CSF molecules connected via different peptide linkers. Three different homodimeric G-CSF proteins were purified, and their in vitro and in vivo activities were determined. A G-CSF dimer, GCSF-Lα, was constructed using an alpha-helix-forming peptide linker, and it demonstrated an extended half-life in serum with a stronger neutrophil response as compared to the monomeric G-CSF protein. The GCSF-Lα protein, therefore, might be selected for further studies as a potential drug candidate.
Agid:
5807633