Recurring genomic structural variation leads to clonal instability and loss of productivity

Chinese hamster ovary cells, commonly used in the production of therapeutic proteins, are aneuploid. Their chromosomes bear structural abnormality and undergo changes in structure and number during cell proliferation. Some production cell lines are unstable and lose their productivity over time in the manufacturing process and during the product’s life cycle. To better understand the link between genomic structural changes and productivity stability, an immunoglobulin G producing cell line was successively single‐cell cloned to obtain subclones that retained or lost productivity, and their genomic features were compared. Although each subclone started with a single karyotype, the progeny quickly diversified to a population with a distribution of chromosome numbers that is not distinctive from the parent and among subclones. The comparative genomic hybridization (CGH) analysis showed that the extent of copy variation of gene coding regions among different subclones stayed at levels of a few percent. Genome regions that were prone to loss of copies, including one with a product transgene integration site, were identified in CGH. The loss of the transgene copy was accompanied by loss of transgene transcript level. Sequence analysis of the host cell and parental producing cell showed prominent structural variations within the regions prone to loss of copies. Taken together, we demonstrated the transient nature of clonal homogeneity in cell line development and the retention of a population distribution of chromosome numbers; we further demonstrated that structural variation in the transgene integration region caused cell line instability. Future cell line development may target the transgene into structurally stable regions. A CHO cell line producing IgG was successively single cell cloned to obtain subclones of high and low productivities, and the resulting genomic heterogeneity was investigated. The clonally derived cell lines had a wide range of chromosome numbers, and certain genome regions were prone to loss in low producing cells, including one of the transgene integration sites. This work demonstrates the transient nature of clonal homogeneity and that structural variation in the transgene integration region led to cell line instability.