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Increasing Transfection Efficiency of Lipoplexes by Modulating Complexation Solution for Transient Gene Expression
Jaemun Kim1,2, Ji Yul Kim1,2, Hyeonkyeong Kim2
1Department of Bioprocess Engineering, KRIBB School of Biotechnology, Korea University of Science and Technology (UST), 217 Gajeong-ro, Yuseong-gu, Daejeon 34113, Korea.
International Journal of Molecular Sciences
|November 27, 2021
Summary
This study developed an efficient transient gene expression method using cationic liposomes and plasmid DNA (pDNA) in Chinese hamster ovary cells. The optimized method achieved high yields of biopharmaceutical proteins, offering a rapid alternative for early-stage drug development.
Area of Science:
- Biotechnology
- Molecular Biology
- Biopharmaceutical Development
Background:
- Transient gene expression offers a rapid alternative to stable cell lines for early-stage biopharmaceutical production.
- Optimizing gene delivery methods is crucial for maximizing protein yields and ensuring cell viability.
Purpose of the Study:
- To establish an efficient transient gene expression methodology using DC-Chol/DOPE cationic liposomes and pDNA in Chinese hamster ovary (CHO) suspension cells.
- To optimize lipoplex formation conditions by modulating liposome and pDNA solutions for enhanced protein production and reduced cytotoxicity.
Main Methods:
- Screening of diverse lipoplex formation conditions, including varying pH of complexation solutions.
- Characterization of liposomes and lipoplexes using dynamic light scattering (DLS) to analyze hydrodynamic size, polydispersity index, and zeta potential.
- Evaluation of protein expression and cellular cytotoxicity post-transfection using fluorescence analysis to trace lipoplex transfer and conformation.
Main Results:
- Successfully produced up to 1785 mg/L of human Fc protein and 191 mg/L of immunoglobulin G (bevacizumab).
- Achieved high yields using a specific combination of acidic liposome formation and alkaline pDNA solutions.
- Demonstrated effective lipoplex transfer to the cytosol and maintained conformation during early transfection stages.
Conclusions:
- The developed lipoplex formation strategy using specific pH conditions is an effective methodology for transient gene delivery.
- This approach provides a promising and efficient gene delivery strategy for biopharmaceutical production in CHO cells.
- The optimized transient gene expression system accelerates the early-stage development of biopharmaceutical candidates.
