Upstream and downstream issues in the production of recombinant vitamin K-dependent proteins in the milk of transgenic pigs

• Main Author: Van Cott, Kevin E.
• Other Authors: Chemical Engineering
• Format: Others
• Language: en
• Published: Virginia Tech 2014
• Subjects: LD5655.V856 1996.V363
• Online Access: http://hdl.handle.net/10919/39172; http://scholar.lib.vt.edu/theses/available/etd-08142006-110134/

The short supply and pathogen safety concerns for plasma-derived therapeutic proteins, such as protein C and Factor IX, have spurred the development of recombinant DNA technology for large-scale, economically feasible production of therapeutic proteins. The mammary gland is unique in its ability to synthesize and secrete large amounts of protein into a harvestable medium, so transgenic livestock secreting recombinant proteins into their milk can be a viable method for the large-scale production of certain proteins. Pigs offer several distinct advantages over other dairy livestock for production of certain recombinant proteins in milk: rapid herd development since pigs have a one year generation time and sows average about two litters per year (about 21 offspring per year), and lactating sows produce an average of about 10 kg of milk/day. The above factors, combined with the excellent safety record of parenteral porcine derivatives, have been the basis for our work on the development of recombinant vitamin K-dependent protein production in porcine milk. This thesis contains the first collection of articles meant to specifically address issues that will affect FDA clinical trials and approval of recombinant VKD proteins from the milk of transgenic pigs. We report on the genotypic and phenotypic stability in lines of transgenic pigs secreting recombinant protein C. In addition, this work details the first reported effects of recombinant protein production on the endogenous milk protein composition of livestock, the first reported production of high levels of biologically active recombinant Factor IX, and the purification of biologically active and inactive subpopulations of recombinant protein C. Our observations suggest that pigs may be used as transgenic bioreactors for large-scale production of protein C and Factor IX. === Ph. D.