Utilization of sequence variants as biomarkers to analyze population dynamics in cloned cell lines. Issue 8 (18th May 2017)

• Record Type: Journal Article
• Authors: Lin, Tzihsuan Jennifer; Beal, Kathryn M.; DeGruttola, Heather S.; Brennan, Steven; Marzilli, Lisa A.; Anderson, Karin
• Is Part Of: Biotechnology and bioengineering. Volume 114:Issue 8(2017)
• Page Start: 1744
• Page End: 1752
• Publication Date: 2017-05-18
• Subjects: cell line development -- clonality -- sequence variants -- population dynamics -- heterogeneity; Biotechnology -- Periodicals; Bioengineering -- Periodicals; 660.6
• DOI: 10.1002/bit.26298
• Languages: English
• ISSNs: 0006-3592

Abstract

ABSTRACT: The inherent nature of cloned CHO cell lines includes the presence of genetic and phenotypic drift that leads to heterogeneous populations. The genetic heterogeneity exhibited by these cells can be exploited to understand the population dynamics of cloned cell lines. Understanding the interplay between heterogeneity, cell culture conditions, and population dynamics will allow for critical assessment of overarching cell line development methods and strategies in terms of population and monoclonality. Sequence variants (SVs) are protein isoforms of the gene‐of‐interest that contain unintended amino acid substitutions, extensions, or truncations that may contribute to heterogeneity. In this case, SVs are unique sequences in the genome of the integrated transgene that can be used as biomarkers to understand the heterogeneity of a monoclonal cell line and how production process conditions can impact population dynamics. In this study, orthogonal genetic and analytical methods were used to examine the variability of SV levels in four different SV‐containing cell lines under varied culture conditions and generational ages. Culture conditions tested had little to no impact on SV levels. However, generational age studies showed two distinct trends: stability of SV levels out to approximately 100 generations in cell lines with higher level SVs (>10%) and a progressive decrease of SV levels as the cells age to approximately 100 generations in cell lines with lower level SVs (<10%). The results suggest that the four SV‐containing cell lines fall into two distinct population models; SVs present in the whole population of cells and SVs present in only a sub‐population of cells. The data presented here are one of the first studies to not only analyze and compare SV levels in both genetic and protein material but also to utilize SVs as biomarkers to probe distinct populations of cloned cell lines. Biotechnol. Bioeng. 2017;114: 1744–1752. © 2017 Wiley Periodicals, Inc. Abstract : Sequence variants (SVs) are protein isoforms that contain unintended substitutions in the amino acid sequence. Different SV‐containing cell lines were examined at various culture conditions including scale, process, and generational age. DNA, RNA, and protein SV levels in all four cell lines were not affected by differing culture processes. However, generational age did show a change. These data allow us to start modeling population dynamics leading to better understanding of SVs and monoclonality of cell lines.