Abstract
While many perceive mammalian cell culture-based manufacturing for biopharmaceuticals an established technology, numerous open questions remain to be solved. Genetic diversity and mutation rates in CHO cells have been underestimated since progeny of a clonal CHO cell become genetically diverse with each cell division. This is an important issue since products are made in bioreactors containing up to 1014 cells that have divided for weeks. Regulatory interest in “Proof of Clonality” is a misguided and misunderstood concern in this context. We revisit decades of research on scope and rate of genetic changes in CHO cells and suggest approaches to minimize trends for genomic instability when establishing reliable manufacturing processes. A concept is proposed for transfection-derived cell populations containing relatively stable (The term “stable” will always be used in a context of relative stability—considering time frames of weeks to months at best.) “CHO master sequence” genomes (containing the desired DNAs of interest). Stable cell populations are to be selected for and maintained for the various phases of manufacturing under specific culture conditions reducing trends for the selection of diverse subpopulations. Such conditions are based on insights gained from population genetics, evolutionary landscape fitness principles, and a 40-year old model for evolution of error prone replicating systems—the Quasi-Species concept.
Highlights
The sheer diversity of genomic and phenotypic variation of CHO cell populations is poorly understood and possibly underestimated
Identical genetics among the cells of a cell population derived from a single cell is not supported by any data
There are no data to demonstrate that cloning enhances the genetic homogeneity of emergent cell populations
Summary
The sheer diversity of genomic and phenotypic variation of CHO cell populations (and of other immortalized cell lines) is poorly understood and possibly underestimated. Mechanisms resulting in mutational evolution of populations in cell culture in general have not been appreciated This is surprising, since numerous aspects of this diversity, as seen on the basis of chromosome structures and karyotypes, were established already five to six decades ago. The well-intended but incorrect regulatory suggestion is that cell line and process stability are linked to clonality Most likely this is understood in a sense that the singularity of a cell and the derivation of dividing daughter cells from such a single cell would provide a (more) robust genetic foundation for reliable manufacture of recombinant proteins using cells. It is hoped, provide a path forward to further improve quality and consistency of products derived from recombinant CHO or other cell lines
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