title: Improved production of monoclonal antibodies through oxygen-limited cultivation of glycoengineered yeast.
abstract: Glycoengineering technology can elucidate and exploit glycan related structure-function relationships for therapeutic proteins. Glycoengineered yeast has been established as a safe, robust, scalable, and economically viable expression platform. It has been found that specific productivity of antibodies in glycoengineered Pichia pastoris is a non-linear function of specific growth rate that is dictated by a limited methanol feed rate. The optimal carbon-limited cultivation requires an exponential methanol feed rate with an increasing biomass concentration and more significantly an increase in heat and mass transfer requirements that often become the limiting factor in scale-up. Both heat and mass transfer are stoichiometrically linked to the oxygen uptake rate. Consequently an oxygen-limited cultivation approach was evaluated to limit the oxygen uptake rate and ensure robust and reliable scale-up. The oxygen-limited process not only limited the maximum oxygen uptake rate (and consequently the required heat removal rate) in mut+ P. pastoris strains but also enabled extension of the induction phase leading to an increased antibody concentration (1.9gL(-1) vs. 1.2gL(-1)), improved N-glycan composition and galactosylation, and reduced antibody fragmentation. Furthermore, the oxygen-limited process was successfully scaled to manufacturing pilot scale and thus presents a promising process option for the glycoengineered yeast protein expression platform.
authors: Biologics New & Enabling Technologies, Merck & Co. Inc., 126 E. Lincoln Ave, Rahway, NJ 07065, United States. marina.berdichevsky@merck.com
authors: Marina
authors: M
authors: Berdichevsky
authors: Marc
authors: M
authors: d'Anjou
authors: Muralidhar R
authors: MR
authors: Mallem
authors: Seemab S
authors: SS
authors: Shaikh
authors: Thomas I
authors: TI
authors: Potgieter
mesh_terms: D000911:Antibodies, Monoclonal / Q000096:biosynthesis*; D018533:Biomass; D019149:Bioreactors; D006358:Hot Temperature; D000432:Methanol; D010101:Oxygen Consumption / Q000502:physiology; D010843:Pichia / Q000254:growth & development / Q000378:metabolism*; D011134:Polysaccharides / Q000378:metabolism; D015202:Protein Engineering / Q000379:methods*
chemical_list: D000911:Antibodies, Monoclonal; D011134:Polysaccharides; D000432:Methanol