: Interferons (IFN) are cytokines with antiviral, antigrowth and immuno-modulatory properties. IFNalpha 2 is employed as a therapy for leukemia, metastasizing carcinoma, kaposi sarcoma and viral hepatitis. The annual cost of IFNalpha 2 therapy is $4000, making it unavailable for most patients worldwide. Among 120 million people infected with hepatitis C virus, 70 percent have abnormal liver function and 33 percent have severe cirrhosis. IFNalpha 5 expressed in liver has the most potent antiviral activity, making it an appealing therapeutic candidate. Insulin-like Growth Factor I (IGF-I) is a potent multifunctional anabolic hormone produced by the liver, and its deficiency results in several systematic complications occurring in liver cirrhosis. The annual requirement of IGF-l per cirrhotic patient is 600 mg, and the cost per mg is $30,000. Transgenic chloroplast technology provides a novel solution to recombinant protein production because of hyper-expression capabilities and ability to fold and process eukaryotic proteins with disulfide bridges (thereby eliminating the need for expensive post-purification processing). Tobacco is an ideal choice because of its large biomass, ease of scale-up (million seeds pen plant), genetic manipulation and an impending need to explore alternative uses for this hazardous crop. Oral delivery of functional biopharmaceuticals reduces 90 percent of the production cost, eliminating the need for expensive purification. Furthermore, bio-encapsulation within plant cells offers protection against proteolytic degradation during digestion. Interferon given orally has biological activity in human and animals. Therefore, IFNalpha2, IFNalpha5 and IGF-l will be expressed as follows: a) Develop recombinant DNA vectors for enhanced expression or oral delivery b) Generate transgenic tobacco and tomato plants c) Characterize transgenic expression of proteins or fusion proteins using molecular and biochemical methods d) Purify therapeutic proteins from transgenic chloroplasts e) Characterize and compare therapeutic proteins (yield, purity, functionality) produced in yeast, E.coli or transgenic plants f) Study in vitro and in vivo (pre-clinical trials) protein bio-functionality, parenteral and oral delivery. Large scale and low cost production via transgenic chloroplasts should provide treatment to patients at an affordable cost and tobacco farmers alternate uses for this hazardous crop.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM063879-04
Application #
6883919
Study Section
Special Emphasis Panel (ZRG1-SSS-2 (01))
Program Officer
Jones, Warren
Project Start
2002-05-01
Project End
2006-06-30
Budget Start
2005-05-01
Budget End
2006-06-30
Support Year
4
Fiscal Year
2005
Total Cost
$210,270
Indirect Cost
Name
University of Central Florida
Department
Biochemistry
Type
Schools of Medicine
DUNS #
150805653
City
Orlando
State
FL
Country
United States
Zip Code
32826
Kwon, Kwang-Chul; Sherman, Alexandra; Chang, Wan-Jung et al. (2018) Expression and assembly of largest foreign protein in chloroplasts: oral delivery of human FVIII made in lettuce chloroplasts robustly suppresses inhibitor formation in haemophilia A mice. Plant Biotechnol J 16:1148-1160
Xiao, Yuhong; Daniell, Henry (2017) Long-term evaluation of mucosal and systemic immunity and protection conferred by different polio booster vaccines. Vaccine 35:5418-5425
Zhang, Bei; Shanmugaraj, Balamurugan; Daniell, Henry (2017) Expression and functional evaluation of biopharmaceuticals made in plant chloroplasts. Curr Opin Chem Biol 38:17-23
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Malhotra, Karan; Subramaniyan, Mayavan; Rawat, Khushboo et al. (2016) Compartmentalized Metabolic Engineering for Artemisinin Biosynthesis and Effective Malaria Treatment by Oral Delivery of Plant Cells. Mol Plant 9:1464-1477
Kwon, Kwang-Chul; Chan, Hui-Ting; León, Ileana R et al. (2016) Codon Optimization to Enhance Expression Yields Insights into Chloroplast Translation. Plant Physiol 172:62-77
Pasoreck, Elise K; Su, Jin; Silverman, Ian M et al. (2016) Terpene metabolic engineering via nuclear or chloroplast genomes profoundly and globally impacts off-target pathways through metabolite signalling. Plant Biotechnol J 14:1862-75

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