Biotechnology companies, pharmaceutical companies, and academic laboratories frequently require gram quantities of antibodies or proteins for research, diagnostic, or clinical purposes. The production of such large quantities of antibodies or proteins recombinantly in mammalian cells is a challenging obstacle. While cell culture media and culturing conditions, play a role of protein production levels, the essential driver for high production yield are the cells used to produce the recombinant protein. While several large biotechnology and pharmaceutical companies (e.g. Genentech/Roche) have been able to successfully develop and utilize high-yield stable cell lines, their cell line development technology is either secret or expensive to contract. Recognizing the market need in this area, LakePharma has set out to develop a process that can make high-yield mammalian stable cell lines relatively inexpensively and with the utmost efficiency, making the development of high-yield stable cell lines accessible to all.
One of the key innovations to improve stable cell line protein production was the development of LakePharma’s proprietary mammalian expression vector technology. LakePharma vectors were rationally designed and constructed by total gene synthesis. Because the vectors were created from scratch, using a deep analysis of previous successes and failures, the vectors are more compact, contain optimized expression cassettes, and have a better orientation of operons. Further these vectors can integrate into mammalian chromosomes with significantly higher efficiency. When tested directly against the vectors from another leading company, the LakePharma vectors yielded about 100 times more stable clones, and remarkably 100% of the stable clones tested expressed target protein at a yield greater than 50 PCD (picograms per cell per day). The stable pools made with LakePharma vectors produced 8 times higher target protein production than those from the comparison vectors.
LakePharma has developed proprietary bicistronic vectors for recombinant antibody production. These vectors contain uniquely positioned restriction enzyme sites that enable efficient cloning of antibody genes. Unlike the popular bicistronic vectors that employ two copies of identical CMV promoters and are susceptible for construct instability, LakePharma vectors do not use CMV promoters and do not have stability issues.
LakePharma uses a variety of gene amplification-compatible selectable markers, including a new patented and proprietary selectable marker system developed completely in house. This new selectable marker has radically improved the stable cell line generation process and has significantly increased the success rate of stable cell line development for difficult targets. Utilizing this new selection process, the expression levels and stability of target genes can now be tightly controlled.
By combining LakePharma’s proprietary antibody expression vectors and high-yield stable cell line development systems, LakePharma has engineered CHO stable lines that produce therapeutic proteins and antibodies at levels exceeding 50 picograms per cell per day. Using this cell line development process, the development path to 1 gram per liter production yield is as short as 2 months. With additional gene amplification and clone selection, the production yield can be further improved to 5 gram per liter, which will reduce the amount of media consumed and the number of bioreactors required. Such technology milestones enable efficient and inexpensive large scale mammalian protein and antibody production.