Half-life extension of efficiently produced DARPin serum albumin fusions as a function of FcRn affinity and recycling
Autor: | Fabian Brandl, Andreas Plückthun, Jan Terje Andersen, Jonas V. Schaefer, Uwe Zangemeister-Wittke, Martina Zimmermann, Kine Marita Knudsen Sand, Hannes Merten, Linda Irpinio, Jeannette Nilsen |
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Přispěvatelé: | University of Zurich, Zangemeister-Wittke, Uwe |
Jazyk: | angličtina |
Rok vydání: | 2021 |
Předmět: |
Transgene
Serum albumin 3003 Pharmaceutical Science Pharmaceutical Science Mice Transgenic Serum Albumin Human 610 Medicine & health Receptors Fc Pichia pastoris 03 medical and health sciences Mice 0302 clinical medicine Neonatal Fc receptor medicine 10019 Department of Biochemistry Animals Humans Designed Ankyrin Repeat Proteins Serum Albumin 030304 developmental biology 0303 health sciences biology Chemistry Histocompatibility Antigens Class I General Medicine biology.organism_classification Human serum albumin Fusion protein Receptor–ligand kinetics Cell biology DARPin 030220 oncology & carcinogenesis Saccharomycetales biology.protein 1305 Biotechnology 570 Life sciences Female Biotechnology medicine.drug Half-Life |
Zdroj: | Merten, Hannes; Brandl, Fabian; Zimmermann, Martina; Schaefer, Jonas V; Irpinio, Linda; Sand, Kine M K; Nilsen, Jeannette; Andersen, Jan Terje; Zangemeister-Wittke, Uwe; Plückthun, Andreas (2021). Half-life extension of efficiently produced DARPin serum albumin fusions as a function of FcRn affinity and recycling. European journal of pharmaceutics and biopharmaceutics, 167, pp. 104-113. Elsevier 10.1016/j.ejpb.2021.07.011 |
ISSN: | 0939-6411 |
Popis: | Serum albumin shows slow clearance from circulation due to neonatal Fc receptor (FcRn)-mediated recycling and has been used for half-life extension. We report here fusions to a high-affinity DARPin, binding to Epithelial Cell Adhesion Molecule (EpCAM). We developed a novel, efficient expression system for such fusion proteins in Pichia pastoris with titers above 300 mg/L of lab-scale shake-flask culture. Since human serum albumin (HSA) does not bind to the murine FcRn, half-lives of therapeutic candidates are frequently measured in human FcRn transgenic mice, limiting useable tumor models. Additionally, serum albumins with extended half-life have been designed. We tested HSA7, motivated by its previously claimed extraordinarily long half-life in mice, which we could not confirm. Instead, we determined a half-life of only 29 h for HSA7, comparable to MSA. The fusion of HSA7 to a DARPin showed a similar half-life. To rationalize these findings, we measured binding kinetics and affinities to murine and human FcRn. Briefly, HSA7 showed affinity to murine FcRn only in the micromolar range, comparable to MSA to its cognate murine FcRn, and an affinity in the nanomolar range only to the human FcRn. This explains the comparable half-life of MSA and HSA7 in mice, while wild-type-HSA has a half-life of only 21 h, as it does not bind the murine FcRn and is not recycled. Thus, HSA-fusions with improved FcRn-affinity, such as HSA7, can be used for preclinical experiments in mice when FcRn transgenes cannot be used, as they reflect better the complex FcRn-mediated recycling and distribution mechanisms. |
Databáze: | OpenAIRE |
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