July 16-20, 2017
Therapeutic antibodies have become one of the most effective therapeutics for human diseases such as cancer, inflammation and viral infection. The production of antibody-based drugs using microbial expression systems is more cost effective with ease of gene manipulation compared to mammalian expression systems. In our team, antibody fragments (ex: BsAb, scFv and Fab) were produced from methylotrophic yeast Pichia pastoris secretion expression system with the AOX1 as driven promoter or E. coli secretion expression system. To achieve high production yield for both system, we investigated fermentation parameter such as base medium, induction medium, induction condition, feeding strategy and pH. For the 250 ml fermentor Pichia system, the nitrogen have been add into glycerol fed medium and/or methanol induction medium and also compared base-medium, buffered glycerol-complex medium (BMGY) and basal salt medium (BS). The highest scFv production was yielded from the basal salt medium as base medium, glycerol fed medium plus nitrogen and multiple carbon source methanol induction medium. This process can yielded over 500 mg/L scFv. After scale-up from 250 ml fermentor to 5L fermentor, the methanol fed-back control system also applied on the 5 L fermentor, can achieve 1.7 g/L scFv in 5 days. The E. coli expression process has passed through screening for high production yield clones in 2 ml deep-well then confirmed by using 250 ml flask scale. Feeding medium, DO, pH etc, parameters were investigated by parallel 250 ml-fermenter. The parameters from 250 ml fermentor were validated by using 5 L fermenter. Under this scale-up procedure, the antibody Fab was 100 folds production yield, production deep well stage at 1 mg/L, production from 250 ml fermentor stage is 50-100 mg/L and production 5 L fermentor stage is over 35-90 mg/L. Although different antibodies will result in different production yield, building a reliable platform to predict production yield from antibody cell clones under deep well and shake flask stage serves a good scale-down model for future scale-up prediction.
Jen-Wei Chang, Wei-Kuang Chi, Neng-Hsien Chang, Wei-Hong Chen, Yi-Hua Huang, and Chih-Hsi Fan, "The microbial antibodies secretion expression platform with scale down fermentors" in "Biochemical and Molecular Engineering XX", Wilfred Chen, University of Delaware, USA Nicole Borth, Universität für Bodenkultur, Vienna, Austria Stefanos Grammatikos, UCB Pharma, Belgium Eds, ECI Symposium Series, (2017). https://dc.engconfintl.org/biochem_xx/37