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Tracing production instability in a clonally derived CHO cell line using single-cell transcriptomics
Ioanna Tzani1, Nicholas Herrmann2, Sara Carillo1
1National Institute for Bioprocessing Research and Training, Co Dublin, Ireland.
Abstract:
A variety of mechanisms including transcriptional silencing, gene copy loss, and increased susceptibility to cellular stress have been associated with a sudden or gradual loss of monoclonal antibody (mAb) production in Chinese hamster ovary (CHO) cell lines. In this study, we utilized single-cell RNA-seq (scRNA-seq) to study a clonally derived CHO cell line that underwent production instability leading to a dramatic reduction of the levels of mAb produced. From the scRNA-seq data, we identified subclusters associated with variations in the mAb transgenes and observed that heavy chain gene expression was significantly lower than that of the light chain across the population. Using trajectory inference, the evolution of the cell line was reconstructed and was found to correlate with a reduction in heavy and light chain gene expression. Genes encoding for proteins involved in the response to oxidative stress and apoptosis were found to increase in expression as cells progressed along the trajectory. Future studies of CHO cell lines using this technology have the potential to dramatically enhance our understanding of the characteristics underpinning efficient manufacturing performance as well as product quality.
Insights
Chinese hamster ovary (CHO) cell lines experiencing monoclonal antibody (mAb) production loss were studied using single-cell RNA sequencing. Researchers found reduced heavy and light chain gene expression, linked to increased oxidative stress and apoptosis, impacting mAb yield.
Area of Science:
- Biotechnology
- Cell Biology
- Molecular Biology
Background:
- Monoclonal antibody (mAb) production in Chinese hamster ovary (CHO) cell lines can be lost through mechanisms like transcriptional silencing or gene copy loss.
- Production instability in CHO cell lines leads to reduced mAb levels, impacting biopharmaceutical manufacturing.
Purpose of the Study:
- To investigate the molecular mechanisms behind mAb production instability in a clonally derived CHO cell line using single-cell RNA sequencing (scRNA-seq).
- To identify cell subclusters and gene expression changes associated with reduced mAb production.
Main Methods:
- Utilized scRNA-seq to analyze a CHO cell line exhibiting production instability.
- Applied trajectory inference to reconstruct cell line evolution and correlate gene expression changes.
- Analyzed gene expression patterns related to mAb transgenes, oxidative stress, and apoptosis.
Main Results:
- Identified subclusters within the CHO cell line associated with variations in mAb transgenes.
- Observed significantly lower heavy chain gene expression compared to light chain expression across the cell population.
- Demonstrated a correlation between cell line evolution, reduced heavy and light chain gene expression, and increased expression of genes involved in oxidative stress and apoptosis.
Conclusions:
- scRNA-seq is a powerful tool for understanding CHO cell line production instability.
- Reduced mAb heavy and light chain expression, coupled with stress responses, contributes to production loss.
- Further research using scRNA-seq can enhance understanding of CHO cell manufacturing performance and product quality.
