Abstract
The full-length cDNA encoding human Granulocyte-macrophage colony-stimulating factor (GM-CSF) was cloned by RT-PCR, placed under the control of CMV promoter, and inserted into adenovirus vector of El-substitution type, pAxlcw. Subsequently, the cassette cosmid was cotransfected into 293 cells together with EcoT22I-digested Ad5-TPC, and the replication-deficient recombinant adenoviruses(Ad) of human GM-CSF were generated efficiently by homologous recombination, with the titers of 1.51×l09pfu/ml. 48 hours after infection with prepared human GM-CSF recombinant adenoviruses in vitro, HeLa cells and primary human skin fibroblasts expressed high levels of human GM-CSF (80⊃400ng/106cells/24hr). These suggest that the recombinant Ad of human GM-CSF prepared by COS/TPC method is effective in mediating GM-CSF gene transfer and might be used in cancer gene therapy.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Roth J and Cristiano R. Gene therapy for Cancer: What have we done and where are we going? J Natl Cancer Inst 1997; 89:21.
Dranoff G, Jaffee E, Lazenby A, et al. Vaccination with irradiated tumor cells engineered to secrete murine granulocyte-macrophage colony-stimulating factor stimulates potent, specific, and long-lasting anti-tumor immunity. Proc Natl Acad Sci USA 1993; 90: 3539.
Miyake S, Makimura M, Kanegae Y, et al. Efficient generation of recombinant adenoviruses using adenovirus DNA-terminal protein complex and a cosmid bearing the full-length virus genome. Proc Natl Acad Sci USA 1996; 93:1320.
Wong G, Witek J, Temple P, et al. Human GM-CSF: Molecular cloning of the complementary DNA and purification of the natural and recombinant proteins. Science 1985; 228:810.
Caux C., Dezutter-Dambuyant C, Schmitt D, et al. GM-CSF and TNF-α cooperate in the generation of dendritic Langerhans cells. Nature 1992; 360:258.
Sallusto F and A. Lanzavecchia. Efficient presentation of soluble antigen by cultured human dendritic cells is maintained by Granulocyte-macrophage colony-stimulating factor plus interleukin 4 and down-regulated by tumor necrosis factor α. J Exp Med 1994; 179:1109.
Cao XT, Zhang WP, Ma SH, et al. Enhanced antitumor effects of tumor antigen-pulsed dendritic cells by their transfection with GM-CSF gene. Science in China 1997; 40:539.
Horwitz M. Adenoviridae and their replication. In Fundamental Virolgy, 2nd edition, edited by Fields B, Knipe D et al. Raven Press, Ltd. New York, 1991; 771:804.
Author information
Authors and Affiliations
Additional information
This work was supported by National High Biotechnology Foundation (Z20-01-03).
Rights and permissions
About this article
Cite this article
Zhang, W., Cao, X., Tao, Q. et al. Construction and expression of the replication-deficient adenonirus vector of human GM-CSF. Chin J Cancer Res 9, 304–308 (1997). https://doi.org/10.1007/BF02974980
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF02974980