Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Rapid, in vivo, evaluation of antiangiogenic and antineoplastic gene products by nonviral transfection of tumor cells

Cancer Gene Therapy volume 11, pages 346–353 (2004)Cite this article

Abstract

Using a nonviral, electroporation-based gene transfection approach, we demonstrate the efficient and consistent transfection of two poorly immunogenic tumor cell lines: B16F10 melanoma and renal carcinoma (RENCA). Three genes, IL-12, angiostatin (AS), and an endostatin:angiostatin fusion protein (ES:AS) were subcloned into a DNA plasmid containing EBNA1-OriP, which was then transfected into B16F10 and RENCA cells. Significant levels of protein were secreted into the culture supernatants of transfected cells in vitro. Transfected tumor cells were injected subcutaneously into mice. All the three transgenes were capable of significantly delaying and reducing the formation of primary B16F10 and RENCA tumors, as well as B16F10 lung metastases. By day 11 post-injection, all control mice that received either mock-transfected or empty vector DNA-transfected B16F10 tumor cells had developed large primary tumors. In contrast, mice that received IL-12-transfected B16F10 cells did not develop appreciable tumors until day 17, and these were significantly smaller than controls. Similar results were observed for the RENCA model, in which only one of the IL-12 mice had developed tumors out to day 31. Expression of AS or ES:AS also significantly delayed and reduced primary tumors. Overall, ES:AS was more effective than AS alone. Furthermore, 25% of the AS mice and 33% of the ES:AS mice remained tumor-free at day 17, by which point all control mice had significant tumors. Mouse survival rates also correlated with the extent of tumor burden. Importantly, no lung metastases were detected in the lungs of mice that had received either AS or ES:AS-transfected B16F10 tumor cells and significantly fewer metastases were found in the IL-12 group. The consistency of our transfection results highlight the feasibility of directly electroporating tumor cells as a means to screen, identify, and validate in vivo potentially novel antiangiogenic and/or antineoplastic genes.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Learn more

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
Figure 8

Similar content being viewed by others

References

  1. 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 

  2. Yannelli JR, Hyatt C, Johnson S, et al. Characterization of human tumor cell lines transduced with the cDNA encoding either tumor necrosis factor alpha (TNF-a) or interleukin-2 (IL-2). J Immunol Methods. 1993;161:77–90.

    Article  CAS  Google Scholar 

  3. Feldman E, Ahmed T, Lutton JD, et al. Adenovirus mediated alpha interferon (IFN-alpha) gene transfer into CD34+ cells and CML mononuclear cells. Stem Cells. 1997;15:386–395.

    Article  CAS  Google Scholar 

  4. Shiloni E, Karp SE, Custer MC, et al. Retroviral transduction of interferon-gamma cDNA into a nonimmunogenic murine fibrosarcoma: generation of T cells in draining lymph nodes capable of treating established parental metastatic tumor. Cancer Immunol Immunother. 1993;37:286–292.

    Article  CAS  Google Scholar 

  5. Nagai H, Hara I, Horikawa T, et al. Gene transfer of secreted-type modified interleukin-18 gene to B16F10 melanoma cells suppresses in vivo tumor growth through inhibition of tumor vessel formation. J Invest Dermatol. 2002;119:541–548.

    Article  CAS  Google Scholar 

  6. Maehara N, Nagai E, Mizumoto K, et al. Gene transduction of NK4, HGF antagonist, inhibits in vitro invasion and in vivo growth of human pancreatic cancer. Clin Exp Metastasis. 2002;19:417–426.

    Article  CAS  Google Scholar 

  7. Indraccolo S, Gola E, Rosato A, et al. Differential effects of angiostatin, endostatin and interferon-alpha(1) gene transfer on in vivo growth of human breast cancer cells. Gene Therapy. 2002;9:867–878.

    Article  CAS  Google Scholar 

  8. Feldman AL, Alexander HR, Hewitt SM, et al. Effect of retroviral endostatin gene transfer on subcutaneous and intraperitoneal growth of murine tumors. J Natl Cancer Inst. 2001;93:1014–1020.

    Article  CAS  Google Scholar 

  9. Scappaticci FA, Smith R, Pathak A, et al. Combination angiostatin and endostatin gene transfer induces synergistic antiangiogenic activity in vitro and antitumor efficacy in leukemia and solid tumors in mice. Mol Ther. 2001;3:186–196.

    Article  CAS  Google Scholar 

  10. Sauter BV, Martinet O, Zhang WJ, et al. Adenovirus-mediated gene transfer of endostatin in vivo results in high level of transgene expression and inhibition of tumor growth and metastases. Proc Natl Acad Sci USA. 2000;97:4802–4807.

    Article  CAS  Google Scholar 

  11. Scholz C, Starck L, Willimsky G, et al. Adenoviral transduction of tumor cells induces apoptosis in co-cultured T lymphocytes. Gene Therapy. 2002;9:1438–1446.

    Article  CAS  Google Scholar 

  12. Steiner MS, Barrack ER . Transforming growth factor-beta 1 overproduction in prostate cancer: effects on growth in vivo and in vitro. Mol Endocrinol. 1992;6:15–25.

    CAS  PubMed  Google Scholar 

  13. van Deventer HW, Serody JS, McKinnon KP, et al. Transfection of macrophage inflammatory protein 1 alpha into B16 F10 melanoma cells inhibits growth of pulmonary metastases but not subcutaneous tumors. J Immunol. 2002;169:1634–1639.

    Article  CAS  Google Scholar 

  14. Klenchin VA, Sukharev SI, Serov SM, et al. Electrically induced DNA uptake by cells is a fast process involving DNA electrophoresis. Biophys J. 1991;60:804–811.

    Article  CAS  Google Scholar 

  15. Wu H, Ceccarelli DF, Frappier L . The DNA segregation mechanism of Epstein–Barr virus nuclear antigen 1. EMBO Rep. 2000;1:140–144.

    Article  CAS  Google Scholar 

  16. Cruickshank J, Shire K, Davidson AR, Edwards AM, Frappier L . Two domains of the Epstein–Barr virus origin DNA-binding protein, EBNA1, orchestrate sequence-specific DNA binding. J Biol Chem. 2000;275:22273–22277.

    Article  CAS  Google Scholar 

  17. Li LH, Shivakumar R, Feller S, et al. Highly efficient, large volume flow electroporation. Technol Cancer Res Treat. 2002;1:341–350.

    Article  CAS  Google Scholar 

  18. Iwai M, Harada Y, Ishii M, et al. Suicide gene therapy of human hepatoma and its peritonitis carcinomatosis by a vector of replicative-deficient herpes simplex virus. Biochem Biophys Res Commun. 2002;291:855–860.

    Article  CAS  Google Scholar 

  19. Wilasrusmee C, Da Silva M, Singh B, et al. A new in vitro model to study endothelial injury. J Surg Res. 2002;104:131–136.

    Article  Google Scholar 

  20. Tomayko MM, Reynolds CP . Determination of subcutaneous tumor size in athymic (nude) mice. Cancer Chemother Pharmacol. 1989;24:148–154.

    Article  CAS  Google Scholar 

  21. Heller LC, Ingram SF, Lucas ML, et al. Effect of electrically mediated intratumor and intramuscular delivery of a plasmid encoding IFN alpha on visible B16 mouse melanomas. Technol Cancer Res Treat. 2002;1:205–209.

    Article  CAS  Google Scholar 

  22. Rusciano D, Lorenzoni P, Burger M . Murine models of liver metastasis. Invas Metastasis. 1994;14:349–361.

    Google Scholar 

  23. Cichon T, Jamrozy L, Glogowska J, et al. Electrotransfer of gene encoding endostatin into normal and neoplastic mouse tissues: inhibition of primary tumor growth and metastatic spread. Cancer Gene Ther. 2002;9:771–777.

    Article  CAS  Google Scholar 

  24. Cemazar M, Sersa G, Wilson J, et al. Effective gene transfer to solid tumors using different nonviral gene delivery techniques: electroporation, liposomes, and integrin-targeted vector. Cancer Gene Ther. 2002;9:399–406.

    Article  CAS  Google Scholar 

  25. Sakurai F, Terada T, Maruyama M, et al. Therapeutic effect of intravenous delivery of lipoplexes containing the interferon-beta gene and poly I: poly C in a murine lung metastasis model. Cancer Gene Ther. 2003;10:661–668.

    Article  CAS  Google Scholar 

  26. Chen ZY, He CY, Ehrhardt A, et al. Minicircle DNA vectors devoid of bacterial DNA result in persistent and high-level transgene expression in vivo. Mol Ther. 2003;8:495–500.

    Article  CAS  Google Scholar 

  27. Li LH, Wang S, Wood M, et al. Non-viral gene transfer into resting primary lymphocytes: small and large volume. Mol Ther. 2001;3:S69.

    Google Scholar 

  28. Li LH, Feller S, Fratantoni JC, et al. Single base gene conversion by single strand DNA oligo delivered by electroporation. Mol Ther. 2002;5:S334.

    Google Scholar 

Download references

Acknowledgements

We thank Nicholas Chopas, Nat Forgotson and Sarah Wang for instrumentation support. We thank JoAnn Murphy-Roach and Bobby Mercer for assistance with the animal studies and caring for the animals. We would like to acknowledge Dr Osam Mazda at Kyoto Prefectural University of Medicine in Japan for providing the pGEG and pGEG.mIL-12 plasmid. We would also like to thank Drs John Holaday, Kim Lee Sim, and Carolyn Sidor for helpful discussions.

Author information

Authors and Affiliations

  1. MaxCyte, Inc., 9640 Medical Center Drive, Rockville, 20850, Maryland, USA

    Jonathan M Weiss, Rama Shivakumar, Stephanie Feller, Lin-Hong Li, Joseph C Fratantoni & Linda N Liu

  2. EntreMed, Inc., 9640 Medical Center Drive, Rockville, 20850, Maryland, USA

    Art Hanson & William E Fogler

Authors
  1. Jonathan M Weiss
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Rama Shivakumar
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Stephanie Feller
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Lin-Hong Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Art Hanson
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. William E Fogler
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Joseph C Fratantoni
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Linda N Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Linda N Liu.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Weiss, J., Shivakumar, R., Feller, S. et al. Rapid, in vivo, evaluation of antiangiogenic and antineoplastic gene products by nonviral transfection of tumor cells. Cancer Gene Ther 11, 346–353 (2004). https://doi.org/10.1038/sj.cgt.7700686

Download citation

  • Received:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/sj.cgt.7700686

Keywords

This article is cited by

Search

Advanced search

Quick links