Production of Recombinant Therapeutic Proteins in the Milk of Transgenic Animals - - BioPharm International


Production of Recombinant Therapeutic Proteins in the Milk of Transgenic Animals

BioPharm International
Volume 19, Issue 8


Table 1. List of therapeutic proteins produced in the milk of transgenic animals that are currently in commercial development.
Here is the method to achieve milk-specific recombinant protein production. Fuse an expression vector, comprising a gene that is encoded for the human or humanized target protein with mammary gland-specific regulatory sequences, and then insert into the germline of the selected production species. When integrated, the milk-specific expression construct becomes a dominant genetic characteristic that is inherited by the progeny of the founder animal (Figure 1). This general strategy makes it possible to harness the ability of dairy animal mammary glands to produce large quantities of complex proteins.

GTC Biotherapeutics and other have generated transgenic animal herds that yield large amounts of proteins as diverse as: human antithrombin (AT), alpha1-antitrypsin (AAT), C1 esterase inhibitor, fibrinogen, albumin, and monoclonal antibodies (Table 1). Technologies that permit the clinical-grade purification of recombinant therapeutic proteins from the milk of transgenic dairy animals have been developed and implemented.

Limitations of the transgenic expression systems are related to potential adverse effects of bioactive heterologous proteins on the health of the production animals and, to a lesser extent, to initial timelines. Although transgenic expression systems are able to perform complex post-translational modifications, such as γ-carboxylation, β-hydroxylation or N- and O-linked glycosylation, there are species- and tissue-specific characteristics for these modifications that may affect the appropriateness of a given system for the expression of specific proteins. This is also a challenge found with mammalian cell culture, microbial expression systems, or transgenic plants.


The targeting of heterologous proteins to the mammary gland of transgenic mice was independently reported by several groups during the late 1980s.1,2,3 These initial successes were followed by reports relating the generation of transgenic sheep, goats, cows, and pigs with milk-specific transgenes with the ultimate objective of producing recombinant proteins for clinical use (reviews by Clark,4 Meade et al.,5 Pollock et al.6 ).

Figure 1. Schematic representation of the transgenic production process, using the production of rhAT in the milk of transgenic goats as an example.
The aim was to target recombinant proteins to the mammary gland of transgenic farm animals to solve problems associated with either microbial or animal cell expression systems. Bacteria often improperly fold complex proteins, leading to involved and expensive refolding processes, and both bacteria and yeast lack adequate post-translational modification machinery for mammalian-specific N- and O-linked glycosylation, γ-carboxylation, and proteolytic processing. Cell culture systems require high initial capital expenditures, lack scale-up (or down) flexibility, and use large volumes of culture media. On the other hand, transgenic livestock can be maintained and scaled-up in relatively inexpensive facilities, use animal feed as raw material, and can achieve impressive yields of recombinant proteins.

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