[Construction of two robust CHO cell lines resistant to apoptosis and adapted to protein-free medium by over-expression of Igf-1/bcl-2 or bcl-2/cyclin E genes].

Serum used widely in mammalian cell culture is also a potential source of bacterial, mycoplasmal and viral contaminations. In addition, the complex biological components in serum make harder the subsequent product recovery process. High cost, high batch variation and potential source limitation are among the other shortcomings.

[Construction of CHO-IVB, A serum-independent, apoptosis-resistant cell line that can grow in adherence].

Without serum to provide adherent factors, CHO-dhfr- cells grow in suspension when cultured in serum-free medium. Although this offers advantages in some applications, in most production systems adherent cell growth is preferable. Gene transfection, clonal selection and amplification can be easier for adherent cells; the density of immobilized cells is often higher than those in suspension culture, which results in a higher protein productivity; washout of cells by perfused medium during continuous fermentation can be avoided; for high-throughput microplate assays, adherent cells are preferred to facilitate medium changes and cell washing.

[Construction of an anti-apoptosis CHO cell line for biopharmaceutical production].

Mammalian cells are prone to apoptosis when cultured in large scale for production of biopharmaceuticals. And this will reduce production duration and result in high cost of production. Apoptosis is triggered by various factors, and delicately regulated by a set of genes.

Small antisense RNA to cyclin D1 generated by pre-tRNA splicing inhibits growth of human hepatoma cells.

Introns are present in some human pre-tRNAs. They are spliced out during the maturation processes of pre-tRNAs in a way that is irrelevant to their specific nucleotide sequences. This unique characteristic of tRNA splicing can be used for generation of small antisense RNAs by replacing the intron sequences with corresponding antisense sequences.

Blocking caspase-3 activity with a U6 SnRNA promoter-driven ribozyme enhances survivability of CHO cells cultured in low serum medium and production of interferon-beta.

Apoptosis responding to various insults in bioreactors was observed to reduce cell viability and prevent cells from growing to high density. Inhibition of apoptosis in different ways has proved to be effective in keeping cells viable in high density and result in higher recombinant protein production. In apoptosis, death signals activate a family of proteinases, namely caspases, in a cascade and ultimately activate the final effector proteinase, caspase-3, which cleaves various substrates and drives the cells irreversibly to death.

[Modification of Chinese hamster ovary cells].

Chinese hamster ovary cells (CHO) are preferable to prokaryotic, yeast or insect cells as hosts for biopharmaceutical production due to the products are more similar to their natural conformation. However, CHO cells confront tremendous difficulties when cultured in large scale such as mal-adaptation to serum-free medium, apoptosis and over-growth without limitation. So in addition to optimizing CHO system in respect of medium, environment and expression vector, modification of CHO cells themselves has drawn more and more attention.