Identification of Loci with High Transgene Expression in CHO Cells

Area of Science:

• Biotechnology
• Cell Biology
• Genomics

Background:

• Chinese Hamster Ovary (CHO) cells are crucial for biopharmaceutical protein production.
• Targeted integration (TI) into high-expressing loci simplifies cell line development for biologics.
• Identifying these optimal genomic loci is essential for efficient transgene expression.

Purpose of the Study:

• To demonstrate the utility of Thousands of Reporters Integrated in Parallel (TRIP) technology for identifying transcriptional hotspots in CHO cells.
• To establish a functional screening method for locating genomic regions that enhance transgene expression.
• To compare the efficiency of TRIP-identified loci with traditional methods for stable expression.

Main Methods:

• Utilized Thousands of Reporters Integrated in Parallel (TRIP) technology with randomly integrated barcoded reporters.
• Employed next-generation sequencing to track reporter genomic locations and activity.
• Analyzed reporter expression levels and correlated them with active chromatin marks.

Main Results:

• TRIP technology successfully identified transcriptional hotspots in CHO cells.
• These hotspots led to significant increases in mRNA levels (up to 9.4-fold) and protein titers (up to 5.6-fold).
• Single-copy expression from a TRIP-identified locus outperformed piggyBac-mediated stable expression (up to 1.6-fold higher titer).

Conclusions:

• TRIP is an effective functional screening tool for identifying CHO cell transcriptional hotspots.
• This method simplifies the discovery of high-expression loci without complex epigenomic analyses.
• TRIP technology advances cell line development for enhanced biologics manufacturing.