MAMMALIAN-CELL CULTURES .2. GENETIC-ENGINEERING, PROTEIN GLYCOSYLATION, FERMENTATION AND PROCESS-CONTROL

For expression of human genes in mammalian cell culture regulatory seq uences such as promotor or terminator region of viral origin are requi red The most widely used expression system uses dihydrofolic acid redu ctase (DHFR) as a selection marker in conjunction with a DHFR deficien t Chinese hamster ovary (CHO) cell using methotrexat as selection pres sure. Alternatively the glutamine synthetase amplification system seem s to be one of the most efficient expression systems using methionine sulphoximine (MSX) as selection pressure. Folding and glycosylation ta kes place in mammalian cell cultures at the sites of endoplasmatic ret iculum and the Golgi apparatus and is comparable to synthesis in human cells. Most large scale manufacturing processes for products derived from mammalian cell cultures are fed batch suspension culture processe s up to 15, 000 l. Important factors for productivity are media compos ition and feeding strategies. Sterility of the entire system during th e fermentation period is a decisive factor for success rate. Because m ammalian cell cultures reacting very sensitive to small changes in tem perature, pH, osmolality and agitation the fermentation system require s a reliable process control system. Validation of the entire manufact uring process is required to ensure consistent product quality which m eets predetermined specifications and to provide a basis for an econom ic process. In a joint effort equipment qualification process validati on in-process controls and quality controls provide the basis for prod uct consistency from batch to batch.