All currently marketed therapeutic Mabs are produced in mammalian cell cultures, a process that is prohibitively expensive for large-scale production. ApoLife (www.apolife.com) has perfected an alternative system that uses baker’s yeast (Saccharomyces cerevisiae) to make low-cost, high-volume recombinant proteins and functional human antibodies, according to the company.
Pharmaceutical companies use S. cerevisiae to produce recombinant insulin, vaccines, and other therapeutic proteins and peptides. “You can make several products in yeast,” says immunologist Nalini Motwani, Ph.D., founder and president of ApoLife.
Dr. Motwani started ApoLife in 1997 as a spinoff of Apex Bioscience (now part of Curacyte), which developed a recombinant hemoglobin product in yeast as a blood substitute. Although the recombinant hemoglobin is not yet commercialized, Dr. Motwani applied the yeast technology to the manufacturing of Mabs. Now ApoLife licenses its yeast platform to pharmaceutical and biotechnology companies to make therapeutic Mabs and recombinant proteins. “We are a technology-based company,” says Dr. Motwani, “not a product company.”
Twin Cassette Expression Technology
Because baker’s yeast has been used to manufacture several FDA-approved therapeutic products since the 1980s, the method offers regulatory advantages over other expression systems, such as other yeasts, algae, plants, or transgenic animals.
However, “antibodies are harder to make than insulin,” says Dr. Motwani. Hormone peptides like insulin are made by one gene that is inserted into a plasmid and cloned. Antibodies are more complicated because they consist of two chains: a heavy and light chain coded by different genes.
Many researchers use two plasmids, one for each gene, to generate antibodies. But one gene may prove faulty, resulting in the disproportionate overproduction of one of the proteins. In contrast, ApoLife’s Twin Cassette expression technology inserts both genes into one plasmid to insure equimolar expression of both antibody chains. “If something goes wrong, the yeast stops growing, so you don’t make just one gene product,” says Dr. Motwani.
This one-plasmid system offers a better way to obtain a functional molecule. ApoLife’s scientists have worked out conditions to optimize the production of different antibody chains by using promoters and plasmids, yeast strains, media formulations, and batch fermentation processes.
“We know what plasmid orientations give optimal amounts of the two chains,” says Dr. Motwani. In addition, ApoLife has a large collection of yeast strains to choose from to develop the best expression levels of particular antibodies. This special know-how guarantees high yields of pure Mabs, notes Dr. Motwani.
ApoLife’s yeast system can lower by half the cost of discovering and making Mabs or other biotherapeutics, according to the company. Compared to CHO cells, yeast cells grow three times more rapidly and require less expensive culture media, which is advantageous for screening antibody applications.
For instance, Dr. Motwani reports that the ApoLife method makes 100 milligrams of an antibody in eight days, compared to 21 to 30 days in Chinese hamster ovary (CHO) cell cultures. The faster growth rate of yeast cells translates into smaller production facilities, thereby reducing capital investment.
Animal serum or animal-derived components often are added to mammalian cell cultures, raising safety and regulatory issues about contamination by animal viruses or prions. Rather than using expensive antibiotics for the selection of plasmids, the yeast system uses inexpensive nutritional markers. “It all adds up,” says Dr. Motwani, making yeast production methods a cheaper alternative.
ApoLife is seeking collaborators with antibodies that have been expressed in CHO cells to demonstrate expression in the yeast system. Early in 2006, ApoLife started making Mabs for companies that currently rely on CHO or other mammalian cell culture methods. “People are starting to pay attention,” says Dr. Motwani, and three major biotechnology companies signed on. The customers, which cannot be named, are performing characterization and validation of their yeast-generated products.
Researchers can send their antibodies to ApoLife, where experts adapt the yeast system to efficiently produce a particular Mab. The recombinant proteins are returned to the collaborators for evaluation and comparison to antibodies generated in CHO cells. Then clients can transfer the yeast protocol to their own facility for production processes. Yeast-based manufacturing processes are readily transferable.
Mabs have proven to be highly successful and profitable therapies for cancer, arthritis, and other diseases. The combined sales of Mab products reached $10 billion in 2004 and are expected to grow to $30 billion by 2010. This creates a demand for manufacturing technologies that are cost-effective and fast.
ApoLife’s yeast system can be applied to antibody screening and discovery from hybridoma subclones or phage display libraries, as well as cost-effective Mab manufacturing. The yeast platform offers advantages ranging from early-stage discovery through commercial manufacturing, which can shorten development cycles from years to months.