Perspective on Transgenic Animal Drug Production

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The pharmaceutical industry has successfully commercialized two transgenic animal drug products, ATRYN and RUCONEST.1,2 More than 40 additional drugs are candidates for production through this technology.3

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ATRYN (recombinant antithrombin) was approved in 2009 for prevention of thromboembolic events in antithrombin-deficient patients.4 It is produced using genetically engineered goats who carry the DNA coding sequence for human antithrombin linked to a DNA sequence limiting expression to milk.4 The ATRYN amino acid sequence is identical to that of human plasma-derived antithrombin and both proteins are configured with six cysteine residues, three disulfide bridges, and 3-4 N-linked carbohydrate moieties.4 While the two versions of the protein are equipotent,4 the glycosylation profile of ATRYN differs slightly from plasma-derived antithrombin. ATRYN contains less sialic acid than its native counterpart and also contains non-native N-glycosylneuraminic acid;5 however, these differences don’t appear to be clinically important.

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RUCONEST (human c1 esterase inhibitor analog) was approved by FDA in July 2014 for acute attacks in patients with hereditary angioedema.6 RUCONEST is produced from the milk of genetically engineered rabbits and is identical to the human plasma protein except that it is not as fully glycosylated.5,6 Consequently, compared to human plasma-derived C1 esterase inhibitor, the engineered protein has a lower molecular mass (68 kDa vs 105 kDa) and a shorter half-life (~3 hours vs ~18 hours).6,7 Like ATRYN, the glycosylation differences don’t appear to be clinically important and 1 unit of RUCONEST is equivalent to the activity of C1 esterase inhibitor contained in 1 mL of human plasma.6,8

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The story behind the science of transgenic drug development is worthwhile reading.5,9  Dating back to the early 1980’s transgenic animals were originally developed for the study of human diseases.10 Progression to commercial transgenic drug production required the perseverance to overcome many challenging technical and regulatory barriers.1,11

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In the United States, transgenic animals are regulated and approved by the FDA along with their drug products.12 For transgenic drug production “founder” animals are typically engineered by combining microinjection of recombinant DNA with in vitro fertilization procedures.3,5,13 Grown to maturity, transgenic founder animals produce the target protein and also pass along the novel manufacturing ability to their offspring. Compared with traditional mammalian cell “bioreactor” methods of production, the main advantages of transgenic animal production are: 1) retention of posttranslational protein modification processes required for the biological activity of complex proteins (e.g., folding, subunit association, and glycosylation), 2) ease of scalability, and 3) low production costs.5,14

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Milk-based synthesis is the most mature system for producing transgenic animal drugs.5 For the production of monoclonal antibodies, however, transgenic chicken eggs, specifically the egg whites, are a more viable production system.5,14 Milk-based production is attractive because, sequestered within mammary tissues, most therapeutic proteins are essentially devoid of systemic pharmacologic activity within the transgenic host (growth hormone is a notable exception).5 Milk-based systems also have the advantage of ease of harvest through the use of highly automated milking machines.15

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Cows, sheep, goats, pigs, rabbits, and camels are all candidate species for milk-based transgenic drug production and the choice between them is largely based on economic considerations.5,16 These include the relative costs associated with keeping livestock (i.e., housing and feed costs), the natural “richness” (protein content) of the milk, gestation period, and number of offspring.5 In the case of RUCONEST, given that each New Zealand rabbit is capable of producing 500 g of therapeutic protein/year (120 mL of milk/day containing 12 g of human c1 esterase inhibitor analog/liter),17 and given that the species is noted for its vigorous reproduction capacity, the use of dairy rabbits seems a logical choice for the transgenic production of this drug product.

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With the “boost” from the successful commercialization of the first two transgenic animal drug products for human use, it is expected that the technology eventually will be extended to “improve” human foodstuffs, for example, through the marketing of dairy products transgenically “fortified” with healthful molecules.5

Reference(s) +
 1. Kling J. First US approval for transgenic animal drug. Nat Biotechnol 2009 Apr; 27(4): 302-4.   [PubMed: 19352350]
 2. FDA approves second transgenic milk drug [News in Brief]. Nat Rev Drug Discov 2014 Sept 1; 13: 644.
 3. Dunn DA, Pinkert CA, Kooyman DL. Foundation Review: Transgenic animals and their impact on the drug discovery industry. Drug Discov Today 2005 Jun 1; 10(11): 757-67.   [PubMed: 15922934]
 4. ATryn, Antithrombin (Recombinant) [prescribing information]. Dated February 3, 2009. GTC Biotherapeutics, Inc., Framingham, MA, U.S.A. Available from http://www.fda.gov/downloads/BiologicsBloodVaccines/BloodBloodProducts/ApprovedProducts/LicensedProductsBLAs/FractionatedPlasmaProducts/UCM134045.pdf Accessed 12/9/14.
 5. Houdebine L-M. Production of pharmaceutical proteins by transgenic animals. Comp Immunol Microbiol Infect Dis 2009 Mar; 32(2):107-121.   [PubMed: 18243312]
 6. Ruconest (C1 esterase inhibitor [recombinant]) [prescribing information]. Revised August 2014. Pharming Group NV, The Netherlands. Available from https://81b77e9a9bc9711e90b1-f416dd7f832b6e1ad8c969a90667ca99.ssl.cf1.rackcdn.com/shared/pi/ruconest-pi.pdf Accessed 12/9/14.
 7. Berinert [C1 Esterase Inhibitor (Human)] [prescribing information]. CSL Behring LLC, Kankakee, IL, U.S.A. Available from http://www.fda.gov/downloads/biologicsbloodvaccines/bloodbloodproducts/approvedproducts/licensedproductsblas/fractionatedplasmaproducts/ucm186268.pdf Last accessed 12/9/14.
 8. Davis B, Bernstein JA. Conestat alfa for the treatment of angioedema attacks. Ther Clin Risk Manag 2011; 7: 265-73. PMCID: PMC3132097.
 9. Velander WH. Lubon H, Drohan WN. Transgenic Livestock as Drug Factories. Sci Am 1997 Jan; 276 (1): 70-74.   [PubMed: 8972619]
10. Houdebine LM. The methods to generate transgenic animals and to control transgene expression. J Biotechnol 2002 Sep 25 ;98 (2-3): 145-60.   [PubMed: 12141984]
11. United States Food and Drug Administration. Points to Consider in the Manufacture and Testing of Therapeutic Products for Human Use Derived from Transgenic Animals. 1995. Available from http://www.fda.gov/downloads/BiologicsBloodVaccines/GuidanceComplianceRegulatoryInformation/OtherRecommendationsforManufacturers/UCM153306.pdf Accessed 12/9/14.
12. Pollack A. F.D.A. Approves Drug from Gene-Altered Goats. The New York Times. February 6, 2009. Available from http://www.nytimes.com/2009/02/07/business/07goatdrug.html?pagewanted=all&_r=0  Accessed 12/9/14.
13. Wheeler MB, Walters EM, Clark SG. Transgenic animals in biomedicine and agriculture: outlook for the future. Anim Reprod Sci 2003 Dec 15; 79 (3-4): 265-89.   [PubMed: 14643108]
14. Choi CQ. Old MacDonald's Pharm: First drug from transgenic goats nears approval. Sci Am 2009 Jan 10. Available from http://www.scientificamerican.com/article/atryn-pharming-goats-transgenic/ Accessed 12/9/14.
15. Houdebine L-M, Fan J (Eds.) Rabbit Biotechnology: Rabbit genomics, transgenesis, cloning and models. New York, NY: Springer; 2009.  [PubMed: ] [[XSLOpenURL/]]
16. Almohsen R. First GM camels to be engineered for drug production. Sci Dev Net. March 9, 2012. Available from http://www.scidev.net/global/genomics/news/first-gm-camels-to-be-engineered-for-drug-production.html  Accessed 12/9/14.
17. Owen J. Rabbits Milked for Human Protein; Drug Soon for Sale?[National Geographic News] December 1, 2009. Available from http://news.nationalgeographic.com/news/2009/12/091201-rabbits-milk-human-protein-drug . Accessed 12/9/14.