title: Controlling glycation of recombinant antibody in fed-batch cell cultures.
abstract: Protein glycation is a non-enzymatic glycosylation that can occur to proteins in the human body, and it is implicated in the pathogenesis of multiple chronic diseases. Glycation can also occur to recombinant antibodies during cell culture, which generates structural heterogeneity in the product. In a previous study, we discovered unusually high levels of glycation (>50%) in a recombinant monoclonal antibody (rhuMAb) produced by CHO cells. Prior to that discovery, we had not encountered such high levels of glycation in other in-house therapeutic antibodies. Our goal here is to develop cell culture strategies to decrease rhuMAb glycation in a reliable, reproducible, and scalable manner. Because glycation is a post-translational chemical reaction between a reducing sugar and a protein amine group, we hypothesized that lowering the concentration of glucose--the only source of reducing sugar in our fed-batch cultures--would lower the extent of rhuMAb glycation. When we decreased the supply of glucose to bioreactors from bolus nutrient and glucose feeds, rhuMAb glycation decreased to below 20% at both 2-L and 400-L scales. When we maintained glucose concentrations at lower levels in bioreactors with continuous feeds, we could further decrease rhuMAb glycation levels to below 10%. These results show that we can control glycation of secreted proteins by controlling the glucose concentration in the cell culture. In addition, our data suggest that rhuMAb glycation occurring during the cell culture process may be approximated as a second-order chemical reaction that is first order with respect to both glucose and non-glycated rhuMAb. The basic principles of this glycation model should apply to other recombinant proteins secreted during cell culture.
authors: Early Stage Cell Culture, Genentech, Inc., 1 DNA Way, South San Francisco, California 94080, USA. inn@gene.com
authors: Inn H
authors: IH
authors: Yuk
authors: Boyan
authors: B
authors: Zhang
authors: Yi
authors: Y
authors: Yang
authors: George
authors: G
authors: Dutina
authors: Kimberly D
authors: KD
authors: Leach
authors: Natarajan
authors: N
authors: Vijayasankaran
authors: Amy Y
authors: AY
authors: Shen
authors: Dana C
authors: DC
authors: Andersen
authors: Bradley R
authors: BR
authors: Snedecor
authors: John C
authors: JC
authors: Joly