For Amgen’s purposes, this usually involves isolating a human gene with therapeutic potential, or genetically engineering a potential therapeutic, then introducing it into bacteria, yeast or an animal cell line. The recombinant systems are induced to produce the protein in high quantities under controlled conditions. In the end, we can produce large quantities of a highly purified protein for clinical and ultimately commercial therapeutic use.
The technique of recombinant DNA, illustrated above, is fairly easy to grasp. Using proteins called restriction enzymes, individual genes from human DNA or genetically engineered in the laboratory, are isolated and inserted into small circular pieces of DNA, known as plasmids. Once inserted into a plasmid, the gene can be glued in place using another enzyme called DNA ligase. Restriction enzymes and DNA ligase are the scissors and glue of recombinant DNA technology.
Once constructed in this way, the recombinant plasmid is inserted into a bacterial, yeast or cultured animal cell in a process called transformation. At Amgen, we use the bacteria Escherichia coli (E. coli), baker's yeast, and a number of mammalian cell lines, including the Chinese Hamster Ovary cell line (CHO cells) we use to produce Aranesp® (Darbepoetin alfa). Transformed cells are separated from non-transformed cells in a selection procedure that takes advantage of drug-resistance genes also found on the plasmid. A pure population of recombinant cells is then established through the process of cloning. In cloning, a single cell is selected, and it gives rise to a whole population of identical cells, or clones, by normal cell division. In this process, all of the resultant cells are expected to contain a copy of the plasmid carrying the inserted DNA sequence, designed to produce a potential therapeutic.
Once the gene has been inserted and the cell line cloned, the cells are then coaxed to turn on, or "express," the DNA sequence. Depending upon the cell system selected, the recombinant protein that is produced may be found inside the cells or outside in the surrounding medium.
