Gene copy number, gene configuration and <scp>LC</scp> / <scp>HC mRNA</scp> ratio impact on antibody productivity and product quality in targeted integration <scp>CHO</scp> cell lines
Zion Lee, Jun Wan, Amy Shen, Gavin C. Barnard
Abstract
The augmentation of transgene copy numbers is a prevalent approach presumed to enhance transcriptional activity and product yield. CHO cell lines engineered via targeted integration (TI) offer an advantageous platform for investigating the interplay between gene copy number, mRNA abundance, product yield, and product quality. Our investigation revealed that incrementally elevating the gene copy numbers of both IgG heavy chain (HC) and light chain (LC) concurrently resulted in the attainment of plateaus in mRNA levels and product titers, notably occurring beyond four to five gene copies integrated at the same TI site. Furthermore, maintaining a fixed gene copy number while varying the position of genes within the vector influenced the LC/HC mRNA ratio, which subsequently exerted a substantial impact on product titer. Moreover, manipulation of the LC/HC gene ratio through the introduction of surplus LC gene copies led to heightened LC mRNA expression and a reduction in the levels of high molecular weight species. It is noteworthy that the effects of excess LC on product titer were dependent on the specific molecule under consideration. The strategic utilization of PCR tags enabled precise quantification of transcription from each expression slot within the vector, facilitating the identification of highly expressive and less expressive slots. Collectively, these findings significantly enhance our understanding of stable antibody production in TI CHO cell lines.