Transcriptional and post-transcriptional targeting for enhanced transient gene expression in CHO cells.

Abstract

To develop a simple approach to increase titers of transient gene expression in CHO cells without relying on host cell line engineering as recent reports suggest that for PEI-mediated transfections, under optimized conditions, DNA delivery into cells and nuclei is not the limiting factor. N, N-Dimethyl acetamide (DMA) was utilized to enhance transcription. To target post-transcriptional events, we evaluated the co-expression of various genes involved in the unfolded protein response, namely XBP1S, ATF4, CHOP and HSPA5. XBP1S overexpression led to a 15-85% increase in titer for multiple therapeutic proteins. Mechanistic studies confirmed that addition of 0.125% DMA increased transgene mRNA levels as expected. However, overexpression of XBP1S had no effect on transgene mRNA levels, indicating that it influenced post-transcriptional events. Since DMA and XBP1S targeted different pathways, the combination of the two approaches led to an additive improvement in protein titer (150-250% titer increase). Transcriptional and post-transcriptional pathways of transient gene expression can be targeted to increase titers without resorting to host cell line engineering in a simple, short, 7day production process.