Poster: Metabolic engineering

Abs # 294: Transgenic Expression of Human Insulin-like Growth Factor Binding Protein-3 (IGFBP-3) in Tobacco Seeds

Presenter:	Cheung, Chun-Kai , stanleykai@mail.hongkong.com
Authors	Cheung, Chun-Kai  (A)   Sun, Samuel S. M. (B)   Tong, Peter C. Y. (A)
Affiliations:	(A): Department of Medicine and Therapeutics, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, N.T., Hong Kong (B): Department of Biology, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong

Insulin-like growth factor binding protein-3 (IGFBP-3) is one of the binding and transporting proteins for the Insulin-like growth factor-I (IGF-I). Co-administration of recombinant IGF-I with recombinant IGFBP-3 (rhIGFBP-3) has been shown to lower plasma glucose and insulin doses in diabetic patients and the side effects can be reduced without affecting the therapeutic efficacy. Furthermore, rhIGFBP-3 alone can be an anti-tumor agent and has demonstrated efficacy in anti-proliferation on numerous cancer cells. Tobacco plant has been demonstrated as bioreactor to produce pharmaceutical proteins at low costs. We plan to express and produce rhIGFBP-3 in tobacco seeds using different expression constructs. Seed-specific phaseolin promoter, with or without phaseolin signal peptide, was used to drive the expression of IGFBP-3. Other two constructs contained additional sequences including the targeting tetrapeptide signal (AFVY) of phaseolin protein and the winged bean lysine-rich protein (LRP) as fusion receptor protein, with an aim to enhance the expression level and increase the target protein stability. Comparing the four constructs, the highest expression level could amount to 0.04% of the total seed weight in the first generation seeds. The crude seed protein extract was subject to biological activity test with human breast cancer cell line (MCF-7). Results showed that the cell growth was inhibited by the addition of total transgenic seed protein consisting of 100-300 ng/ml of rhIGFBP-3 protein. Our results demonstrated that transgenic tobacco seeds can be developed as a bioreactor for the synthesis of human IGFBP-3. Future studies are required to scale up the production of rhIGFBP-3 in cereal crops, such as rice, and to explore the possibility of oral administration.