RVLP-Reduced CHO Cell Lines via ERV Locus Inactivation
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Solution Overview
Problem
There is a need for RVLP-reduced Chinese hamster ovary (CHO) cells to facilitate the commercial production of recombinant proteins, as existing CHO cells produce viral-like particles (VLPs) that can contaminate protein products.
Innovation Solution
Modified CHO cells are developed by inactivating two or more endogenous retrovirus (ERV) loci, such as CHERV-1b, CHERV-2g, ETC109F, and CHERV-3g, through methods like CRISPR/Cas9-based genome editing, resulting in reduced or eliminated expression of RVLPs.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If CHO cells are used for recombinant protein production, then productivity is improved, but RVLP contamination occurs reducing product purity
Solution Approach 1:
The patent extracts and removes the harmful ERV loci (CHERV-1b, CHERV-2g, ETC109F, CHERV-3g) from the CHO cell genome through CRISPR/Cas9-mediated gene editing. By specifically targeting and eliminating these retroviral elements, the patent removes the source of RVLP contamination while preserving the cell line's productivity for recombinant protein expression.
Solution Approach 2:
The patent converts the harmful effect of ERV presence (RVLP production) into a benefit by using the ERV sequences themselves as targets for CRISPR/Cas9 guide RNAs. The same genomic regions that cause contamination are exploited as precise targeting sites for gene editing, allowing selective elimination of the harmful elements while maintaining cell functionality.
2Manufacturing precision
If genome editing is applied to inactivate ERV loci, then RVLP production is reduced improving product purity, but cell line stability may be affected
Solution Approach 1:
The patent applies partial action by selectively targeting only specific ERV loci (CHERV-1b, CHERV-2g, ETC109F, CHERV-3g) rather than attempting to eliminate all retroviral elements or make extensive genomic modifications. This targeted approach removes the harmful components while minimizing disruptions to the overall genomic architecture and cell line stability.
Solution Approach 2:
The patent applies local quality changes by making specific, localized modifications at the ERV locus regions through CRISPR/Cas9 editing. The gene editing is confined to discrete genomic locations (insertions, deletions, or mutations within the ERV sequences) rather than global genomic changes, preserving the stability and integrity of the rest of the cell line genome.
Data Source
AI summary
The presently disclosed subject matter relates to retroviral-like particle (RVLP)-reduced Chinese hamster ovary (CHO) cells suitable to produce recombinant proteins, as well as methods of producing and using such RVLP-reduced CHO production cells.