title: Optimization of cultivation conditions in spin tubes for Chinese hamster ovary cells producing erythropoietin and the comparison of glycosylation patterns in different cultivation vessels.
abstract: This article describes the optimization of cultivation factor settings, that is the shaking rate and working volume in 50 mL spin tubes for a Chinese hamster ovary cell line expressing recombinant human alpha-erythropoietin, using a response D-optimal surface method. The main objectives of the research were, firstly, to determine a setting in which the product titer and product quality attributes in spin tubes are equivalent to those in 250 mL shake flasks in a seven day batch and, secondly, to find a setting in which the product titer is maximal. The model for product titer prediction as a function of shaking rate and working volume in the defined design space was successfully applied to the optimization of cultivation conditions in spin tubes for the tested cell line. Subsequently, validation experiments were carried out simultaneously in spin tubes, shake flasks and bench scale bioreactors to compare cell culture performance parameters such as growth, productivity and product quality attributes in the form of isoform profiles and glycan antennarity structures. The results of the experiments showed that similar cell culture performance and product quality could be achieved in spin tubes when compared to shake flasks. Additionally, bioreactor titers could be reproduced in spin tubes at high shaking rates and low working volumes, but with differing product quality. Cultivation at lower shaking rates in spin tubes and shake flasks produced a glycoprotein with a product quality slightly comparable to that from bioreactors, but with titers being only two thirds.
authors.affiliation: Dept. of Bioprocess Development, Lek Pharmaceuticals d.d., a Sandoz Company, Biopharmaceuticals, Kolodvorska 27, SI-1234 Menges, Slovenia.
authors.forename: Jure
authors.initials: J
authors.lastname: Strnad
authors.forename: Matjaz
authors.initials: M
authors.lastname: Brinc
authors.forename: Vatroslav
authors.initials: V
authors.lastname: Spudić
authors.forename: Nadja
authors.initials: N
authors.lastname: Jelnikar
authors.forename: Lidija
authors.initials: L
authors.lastname: Mirnik
authors.forename: Barbara
authors.initials: B
authors.lastname: Carman
authors.forename: Zdravko
authors.initials: Z
authors.lastname: Kravanja
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D019149:Bioreactors; D016466:CHO Cells / Q000166:cytology* / Q000378:metabolism*; D018929:Cell Culture Techniques / Q000295:instrumentation / Q000379:methods*; D049109:Cell Proliferation; D006224:Cricetinae; D003412:Cricetulus; D000068817:Epoetin Alfa; D004921:Erythropoietin / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D008954:Models, Biological; D011134:Polysaccharides / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins; D015203:Reproducibility of Results; D049490:Systems Biology
chemical_list: D011134:Polysaccharides; D011994:Recombinant Proteins; D004921:Erythropoietin; D000068817:Epoetin Alfa