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Systemic genetic transfer of p21WAF−1 and GM-CSF utilizing of a novel oligopeptide-based EGF receptor targeting polyplex

Cancer Gene Therapy volume 10, pages 529–539 (2003)Cite this article

Abstract

Based on the fact that aberrant overexpression of some growth factor receptors was observed in a variety of human cancer cells, a novel nonviral gene delivery system GE7, which contains a 16-amino-acid ligand for identifying EGF receptor was constructed for tumor-targeted gene therapy. Intravenous administration of GE7 system revealed that it has the ability to target β-galactosidase (β-gal) reporter gene into murine hepatoma (Hepa) cells. Owing to the limited antitumor effects elicited by a single-gene transfer, recent efforts to treat malignancy using combined gene therapy have been accomplished with varying degrees of success. In this study, the human cyclin-dependent kinase inhibitor gene p21WAF−1 and the murine cytokine gene granulocyte–macrophage colony-stimulating factor (GM-CSF) were used simultaneously for in vivo gene therapy through systemic injection of the EGF R targeted GE7/DNA complex into murine hepatoma-bearing mice. The results demonstrated that combined administration of p21WAF−1 and GM-CSF could remarkably inhibit the growth of subcutaneously transplanted hepatoma Hepa cells, and significantly increase the survival rate of tumor-bearing mice. The activities of natural killer (NK) cells and specific cytotoxic T lymphocytes (CTL) were clearly enhanced after combined gene therapy. In vitro experiments showed that p21WAF−1 gene transfer exhibited a suppressive function on the growth of Hepa cells and the expression of H-2Kb and B7-1 molecules on Hepa cells increased significantly after combined genes delivery. All these results suggested that the GE7 system was able to target therapeutic genes efficiently to cancer cells, which showed high EGF R expression. The cotransfer of p21WAF−1 and GM-CSF genes apparently inhibited the growth of tumors through (a) the arrest of tumor cell growth and (b) the enhancement of systemic antitumor immunity.

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Abbreviations

EGF R:

epidermal growth factor receptor

GM-CSF:

granulocyte–macrophage colony-stimulating factor

CKIs:

cyclin-dependent kinase inhibitors

NK:

natural killer

CTL:

specific cytotoxic T lymphocyte

GE7-PL:

GE7-poly-L-lysine

HA20-PL:

HA20-poly-L-lysine

NS:

normal saline

References

  1. Cheng S, Merlino GT, Pastan IH . A versatile method for the coupling of protein to DNA: synthesis of alpha 2-macroglobulin-DNA conjugates. Nucleic Acids Res. 1983;11:659–669.

    Article  CAS  Google Scholar 

  2. Wu CH, Wilson JM, Wu GY . Targeting genes: delivery and persistent expression of a foreign gene driven by mammalian regulatory elements in vivo. J Biol Chem. 1989;264:16985–16987.

    CAS  PubMed  Google Scholar 

  3. Wagner E . Ligand–polycation conjugates for receptor-targeted gene transfer. In: Leaf Huang, Mien-chie Hung, Ernst Wagner, eds. Non-viral Vectors for Gene Therapy. London, UK: Academic Press, 1999:207–277.

    Chapter  Google Scholar 

  4. Curiel DT, Agarwal S, Wagner E, Cotten M . Adenovirus enhancement of transferrin–polylysine-mediated gene delivery. Proc Natl Acad Sci USA. 1991;88:8850–8854.

    Article  CAS  Google Scholar 

  5. Cotten M, Wagner E, Zatloukal K, et al. High-efficiency receptor-mediated delivery of small and large (48 kilobase) gene constructs using the endosome-disruption activity of defective or chemically inactivated adenovirus particles. Proc Natl Acad Sci USA. 1992;89:6094–6098.

    Article  CAS  Google Scholar 

  6. Wagner E, Plank C, Zatloukal K, et al. Influenza virus hemagglutinin HA-2 N-terminal fusogenic peptides augment gene transfer by transferrin–polylysine–DNA complexes: toward a synthetic virus-like gene-transfer vehicle. Proc Natl Acad Sci USA. 1992;89:7934–7938.

    Article  CAS  Google Scholar 

  7. Wagner E, Zatloukal K, Cotten M, et al. Coupling of adenovirus to transferrin–polylysine/DNA complexes greatly enhances receptor-mediated gene delivery and expression of transfected genes. Proc Natl Acad Sci USA. 1992;89:6099–6103.

    Article  CAS  Google Scholar 

  8. Huang SM, Harari PM . Epidermal growth factor receptor inhibition in cancer therapy: biology, rationale and preliminary clinical results. Invest New Drugs. 1999;17:259–269.

    Article  CAS  Google Scholar 

  9. Yanagihara K, Cheng H, Cheng PW . Effects of epidermal growth factor, transferrin, and insulin on lipofection efficiency in human lung carcinoma cells. Cancer Gene Ther. 2000;7:59–65.

    Article  CAS  Google Scholar 

  10. Chen J, Gamou S, Takayanagi A, Shimizu N . A novel gene delivery system using EGF receptor-mediated endocytosis. FEBS Lett. 1994;338:167–169.

    Article  CAS  Google Scholar 

  11. Tian PK, Ren SJ, Ren CC, et al. A novel receptor-targeted gene delivery system for cancer gene therapy. Sci China (Ser C). 1999;42:216–224.

    Article  CAS  Google Scholar 

  12. Li JM, Han JS, Huang Y, et al. A novel gene delivery system targeting cells expressing VEGF receptors. Cell Res. 1999;9:11–25.

    Article  CAS  Google Scholar 

  13. Ren CC, Tian PK, Qu SM, et al. Expression of cyclin-dependent kinase inhibitor genes induces apoptosis in human hepatoma cell line. Chin Sci Bull. 1997;42:2000–2005.

    Article  CAS  Google Scholar 

  14. Liu X, Tian P, Yu Y, et al. Enhanced antitumor effect of EGF R-targeted p21 and GM-CSF gene transfer in the established murine hepatoma by peritumoral injection. Cancer Gene Ther. 2001;9:100–108.

    Article  Google Scholar 

  15. Li Z, Rakkar A, Katayose Y, et al. Efficacy of multiple administrations of a recombinant adenovirus expressing wild-type p53 in an immune-competent mouse tumor model. Gene Therapy. 1998;5:605–613.

    Article  CAS  Google Scholar 

  16. Song YK, Liu F, Chu S, Liu D . Characterization of cationic liposome-mediated gene transfer in vivo by intravenous administration. Hum Gene Ther. 1997;8:1585–1594.

    Article  CAS  Google Scholar 

  17. Liu F, Qi H, Huang L, Liu D . Factors controlling the efficiency of cationic lipid-mediated transfection in vivo via intravenous administration. Gene Therapy. 1997;4:517–523.

    Article  CAS  Google Scholar 

  18. Kircheis R, Schuller S, Brunner S, et al. Polycation-based DNA complexes for tumor-targeted gene delivery in vivo. J Gene Med. 1999;1:111–120.

    Article  CAS  Google Scholar 

  19. Rogulski KR, Zhang K, Kolozsvary A, et al. Pronounced antitumor effects and tumor radiosensitization of double suicide gene therapy. Clin Cancer Res. 1997;3:2081–2088.

    CAS  PubMed  Google Scholar 

  20. Aruga A, Tanigawa K, Aruga E, et al. Enhanced adjuvant effect of granulocyte–macrophage colony-stimulating factor plus interleukin-12 compared with either alone in vaccine-induced tumor immunity. Cancer Gene Ther. 1999;6:89–95.

    Article  CAS  Google Scholar 

  21. Sandig V, Brand K, Herwig S, et al. Adenovirally transferred p16INK4/CDKN2 and p53 genes cooperate to induce apoptotic tumor cell death. Nat Med. 1997;3:313–319.

    Article  CAS  Google Scholar 

  22. Putzer BM, Bramson JL, Addison CL, et al. Combination therapy with interleukin-2 and wild-type p53 expressed by adenoviral vectors potentiates tumor regression in a murine model of breast cancer. Hum Gene Ther. 1998;9:707–718.

    Article  CAS  Google Scholar 

  23. Cao X, Huang X, Ju DW, et al. Enhanced antitumoral effect of adenovirus-mediated cytosine deaminase gene therapy by induction of antigen-presenting cells through stem cell factor/granulocyte–macrophage colony-stimulating factor gene transfer. Cancer Gene Ther. 2000;7:177–186.

    Article  CAS  Google Scholar 

  24. Fan W, Richter G, Cereseto A, et al. Cytokine response gene 6 induces p21 and regulates both cell growth and arrest. Oncogene. 1999;18:6573–6582.

    Article  CAS  Google Scholar 

  25. Eastham JA, Hall SJ, Sehgal I, et al. In vivo gene therapy with p53 or p21 adenovirus for prostate cancer. Cancer Res. 1995;55:5151–5155.

    CAS  PubMed  Google Scholar 

  26. Joshi US, Chen YQ, Kalemkerian GP, et al. Inhibition of tumor cell growth by p21WAF1 adenoviral gene transfer in lung cancer. Cancer Gene Ther. 1998;5:183–191.

    CAS  PubMed  Google Scholar 

  27. 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;90:3539–3543.

    Article  CAS  Google Scholar 

  28. Burger JA, Baird SM, Powell HC, et al. Local and systemic effects after adenoviral transfer of the murine granulocyte–macrophage colony-stimulating factor gene into mice. Br J Haematol. 2000;108:641–652.

    Article  CAS  Google Scholar 

  29. Simons JW, Mikhak B, Chang JF, et al. Induction of immunity to prostate cancer antigens: results of a clinical trial of vaccination with irradiated autologous prostate tumor cells engineered to secrete granulocyte–macrophage colony-stimulating factor using ex vivo gene transfer. Cancer Res. 1999;59:5160–5168.

    CAS  PubMed  Google Scholar 

  30. Waldman T, Zhang Y, Dillehay L, et al. Cell-cycle arrest versus cell death in cancer therapy. Nat Med. 1997;3:1034–1036.

    Article  CAS  Google Scholar 

  31. Lowe SW, Ruley HE, Jacks T, Housman DE . p53-Dependent apoptosis modulates the cytotoxicity of anticancer agents. Cell. 1993;74:957–967.

    Article  CAS  Google Scholar 

  32. Symonds H, Krall L, Remington L, et al. p53-Dependent apoptosis suppresses tumor growth and progression in vivo. Cell. 1994;78:703–711.

    Article  CAS  Google Scholar 

  33. Fisher DE . Apoptosis in cancer therapy: crossing the threshold. Cell. 1994;78:539–542.

    Article  CAS  Google Scholar 

  34. Barratt-Boyes SM, Vlad A, Finn OJ . Immunization of chimpanzees with tumor antigen MUC1 mucin tandem repeat peptide elicits both helper and cytotoxic T-cell responses. Clin Cancer Res. 1999;5:1918–1924.

    CAS  PubMed  Google Scholar 

  35. Hiltbold EM, Alter MD, Ciborowski P, Finn OJ . Presentation of MUC1 tumor antigen by class I MHC and CTL function correlate with the glycosylation state of the protein taken up by dendritic cells. Cell Immunol. 1999;194:143–149.

    Article  CAS  Google Scholar 

  36. Tamada K, Shimozaki K, Chapoval AI, et al. Modulation of T-cell-mediated immunity in tumor and graft-versus-host disease models through the LIGHT co-stimulatory pathway. Nat Med. 2000;6:283–289.

    Article  CAS  Google Scholar 

  37. Cao X, Ju DW, Tao Q, et al. Adenovirus-mediated GM-CSF gene and cytosine deaminase gene transfer followed by 5-fluorocytosine administration elicit more potent antitumor response in tumor-bearing mice. Gene Therapy. 1998;5:1130–1136.

    Article  CAS  Google Scholar 

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Acknowledgements

We thank Mr Ming Yao for his help on animal manipulation, Dr Jingjun Li and Ruijiao Zhao for the immunohistochemistry analysis, Min Bai and Hongmei Song (Department of Immunology, Second Military Medical University, Shanghai, China) for their excellent technical assistance in animal experiments and Mr Zhu Tengfang (Department of Pathology, Shanghai Medical University, Shanghai, China) for his assistance with frozen section preparation. We are also grateful to Professor Jianyun Dong (Department of Immunology, Medical University of South Carolina, SC) and Dr William M Shannon (Senior Vice President, GenPhar Inc., SC) for critical reading of the manuscript and helpful suggestion and comments. This work was supported by grants from the Biotechnology Project, National High Technology Program (Project No. Z-20-01-01).

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Authors and Affiliations

  1. National Laboratory for Oncogenes and Related Genes, Shanghai Cancer Institute, Shanghai, China

    Xiang Liu, Pei-Kun Tian, Ming Yao & Jian-Ren Gu

  2. Department of Immunology, Second Military Medical University, Shanghai, China

    Dian-Wen Ju, Min-Hui Zhang & Xue-Tao Cao

  3. Neworgen Limited,

    Jian-Ren Gu

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  1. Xiang Liu
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Correspondence to Jian-Ren Gu.

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Liu, X., Tian, PK., Ju, DW. et al. Systemic genetic transfer of p21WAF−1 and GM-CSF utilizing of a novel oligopeptide-based EGF receptor targeting polyplex. Cancer Gene Ther 10, 529–539 (2003). https://doi.org/10.1038/sj.cgt.7700596

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