Study On Amplification Of Upstream Production Of Anti CD20 Monoclonal Antibody

Background and aim:The clinical application of antitumor is the most important aspect of antibody drug.In order to meet the clinical needs,the global pharmaceutical research and development,production,engineering,clinical and other aspects are committed to the developing and producing high quality antibody drugs with low price.One of the important production links is the production of monoclonal antibody drugs and the scaling-up and optimization of the production process,which is an essential key step to ensure that antibody drugs can be developed and marketed.This thesisreviews the development and market of antibody drugs,illustrates the importance of process amplification in the production process,and then summarizes the data obtained from the experimental results,analyzes the product quality under the scale of 200L,500L and 2000L to evaluate the effect of the process transfer and amplification.Through the trend chart of the experimental data,the key parameters of process amplification in the production scale of antibody drugs are obtained,so as to be used as a reference for scientific research and engineering technicians in the field of biomedicine.Methods:We took rituximab biosimilar product as an example to study the engineering scale-up and technology/process transferusing Mabthera（Rituxan,rituximab）as a positive control.CHO cell lines,which have been improved and can express human mouse chimeric antibody gene efficiently,were cultured and produced in different scales.It evaluates the effectiveness of process amplification by analyzing and comparing the quality attributes and process attributes of production process and cell culture harvest.This study is divided into two stages,the first stage is from 200L cell culture production process to 500L production process;the second stage is further from 500L to 2000L cell culture production scale.In the research process,we focused on the viable cell density（VCD）,cell viability（Via）,antibody concentration（Titer）,product quality analysis（ion exchange column chromatography,CEX）,glycosylation analysis（Glycon）,pH,partial pressure of oxygen（p O₂）/partial pressure of carbon dioxide（p CO₂）,nutrition and metabolite analysis（Gluc,Lac）.These key parameters can be used as the basis to evaluate the scale-up effect of the process.Results:In the first phase of the study,the components of the medium were adjusted and the equipment with better performance was used in the process of scale-up from 200L to 500L.12 batches of 200L scale cell culture and 19batches of 500L scale cell culture were carried out.The data show that these changes can make the production have better repeatability.Compared with200L,the culture time of 500L was shortened by one day.Although Titer was lower,CEX and Glycan of the product were more similar to the originator.Combined with the detection results of VCD,Via,pH,p O₂,Gluc and Lac,the cell culture process can be successfully enlarged from 200L to500L,and the adjustment in the process makes the product more similar to Rituxan.In the second phase of the study,the production process was directly scaled up from 500L to 2000L without major changes.56 batches were carried out in 2000L scale.The performance of the equipment used in cell culture was good,and the culture time of one day was shortened.Although the impurity of the product from ion-exchang chromatography（CEX）analysis was slightly higher than that of 500L,expression and product quality（Titer and Glycan）were better.Combined with the process data of VCD,Via,pH,p CO₂ and the detection results of cell metabolism（Gluc and Lac）,the adjustment made in the process of enlarging 500L to 2000L is more conducive to the production of biosimilar of rituximab.