Skip to main content
SpringerLink
Log in

In Vitro and In Vivo Models for Evaluation of GDEPT

Quantifying Bystander Killing in Cell Cultures and Tumors

  • Protocol
Suicide Gene Therapy

Part of the book series: Methods in Molecular Medicine™ ((MIMM,volume 90))

Abstract

The vectors currently available for gene therapy of cancer rarely achieve expression of therapeutic genes in more than a small fraction of the cells in solid tumors. This makes therapeutic success critically dependent on secondary events, known as bystander effects, by which transgene expression leads to the death of nontransduced tumor cells. An efficient bystander effect has the potential to compensate for spatially nonuniform expression of therapeutic genes, and its optimization is therefore an important goal in gene therapy of cancer. Here, we describe protocols for quantifying bystander effects using in vitro and in vivo experimental models.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Culver, K. W., Ram, Z., Wallbridge, S., Ishii, H., Oldfield, E. H., and Blaese, R. M. (1992) In vivo gene transfer with retroviral vector-producer cells for treatment of experimental brain tumors. Science 256, 1550–1552.

    Article  PubMed  CAS  Google Scholar 

  2. Mesnil, M. and Yamasaki, H. (2000) Bystander effect in herpes simplex virus-thymidine kinase/ganciclovir cancer gene therapy: role of gap-junctional intercellular communication. Cancer Res. 60, 3989–3999.

    PubMed  CAS  Google Scholar 

  3. Huber, B. E., Austin, E. A., Good, S. S., Knick, V. C., Tibbels, S., and Richards, C. A. (1993) In vivo antitumor activity of 5-fluorocytosine on human colorectal carcinoma cells genetically modified to express cytosine deaminase. Cancer Res. 53, 4619–4626.

    PubMed  CAS  Google Scholar 

  4. Kievit, E., Nyati, M. K., Ng, E., Stegman, L. D., Parsels, J., Ross, B. D., Rehemtulla, A., and Lawrence, T. S. (2000) Yeast cytosine deaminase improves radiosensitization and bystander effect by 5-fluorocytosine of human colorectal cancer xenografts. Cancer Res. 60, 6649–6655.

    PubMed  CAS  Google Scholar 

  5. Waxman, D. J., Chen, L., Hecht, J. E., and Jounaidi, Y. (1999) Cytochrome P450-based cancer gene therapy: recent advances and future prospects. Drug Metab. Rev. 31, 503–522.

    Article  PubMed  CAS  Google Scholar 

  6. Springer, C. J., Antoniw, P., Bagshawe, K. D., Searle, F., Bisset, G. M., and Jarman, M. (1990) Novel prodrugs which are activated to cytotoxic alkylating agents by carboxypeptidase G2. J. Med. Chem. 33, 677–681.

    Article  PubMed  CAS  Google Scholar 

  7. Marais, R., Spooner, R. A., Light, Y., Martin, J., and Springer, C. J. (1996) Gene-directed enzyme prodrug therapy with a mustard prodrug/carboxypeptidase G2 combination. Cancer Res. 56, 4735–4742.

    PubMed  CAS  Google Scholar 

  8. Anlezark, G. M., Melton, R. G., Sherwood, R. F., Coles, B., Friedlos, F., and Knox, R. J. (1992) The bioactivation of 5-(aziridin-1-yl)-2,4-dinitrobenzamide (CB1954)—I. Purification and properties of a nitroreductase enzyme from Escherichia coli—a potential enzyme for antibody-directed enzyme prodrug therapy (ADEPT). Biochem. Pharmacol. 44, 2289–2295.

    Article  PubMed  CAS  Google Scholar 

  9. Anlezark, G. M., Melton, R. G., Sherwood, R. F., et al. (1995) Bioactivation of dinitrobenzamide mustards by an E. coli B nitroreductase. Biochem. Pharmacol. 50, 609–618.

    Article  PubMed  CAS  Google Scholar 

  10. Grove, J. I., Searle, P. F., Weedon, S. J., Green, N. K., McNeish, I. A., and Kerr, D. J. (1999) Virus-directed enzyme prodrug therapy using CB 1954. Anti-Cancer Drug Des. 14, 461–472.

    CAS  Google Scholar 

  11. Bridgewater, J. A., Springer, C. J., Knox, R. J., Minton, N. P., Michael, N. P., and Collins, M. K. (1995) Expression of the bacterial nitroreductase enzyme in mammalian cells renders them selectively sensitive to killing by the prodrug CB1954. Eur. J. Cancer 31A, 2362–2370.

    Article  PubMed  CAS  Google Scholar 

  12. Green, N. K., Youngs, D. J., Neoptolemos, J. P., et al. (1997) Sensitization of colorectal and pancreatic cancer cell lines to the prodrug 5-(aziridin-1-yl)-2,4-dinitrobenzamide (CB1954) by retroviral transduction and expression of the E. coli nitroreductase gene. Cancer Gene Ther. 4, 229–238.

    PubMed  CAS  Google Scholar 

  13. Friedlos, F., Court, S., Ford, M., Denny, W. A., and Springer, C. (1998) Gene-directed enzyme prodrug therapy: quantitative bystander cytotoxicity and DNA damage induced by CB1954 in cells expressing bacterial nitroreductase. Gene Ther. 5, 105–112.

    Article  PubMed  CAS  Google Scholar 

  14. McNeish, I. A., Green, N. K., Gilligan, M. G., et al. (1998) Virus directed enzyme prodrug therapy for ovarian and pancreatic cancer using retrovirally delivered E. coli nitroreductase and CB1954. Gene Ther. 5, 1061–1069.

    Article  PubMed  CAS  Google Scholar 

  15. Djeha, A. H., Hulme, A., Dexter, M. T., et al. (2000) Expression of Escherichia coli B nitroreductase in established human tumor xenografts in mice results in potent antitumoral and bystander effects upon systemic administration of the prodrug CB1954. Cancer Gene Ther. 7, 721–731.

    Article  PubMed  CAS  Google Scholar 

  16. Wilson, W. R., Pullen, S. M., Hogg, A., et al. (2002) Quantitation of bystander effects in nitroreductase suicide gene therapy using three-dimensional cell cultures. Cancer Res. 62, 1425–1432.

    PubMed  CAS  Google Scholar 

  17. Chung-Faye, G., Palmer, D., Anderson, D., et al. (2001) Virus-directed, enzyme prodrug therapy with nitroimidazole reductase: a phase I and pharmacokinetic study of its prodrug, CB1954. Clin. Cancer Res. 7, 2662–2668.

    PubMed  CAS  Google Scholar 

  18. Freeman, S. M., Abboud, C. N., Whartenby, K. A., et al. (1993) The “bystander effect”: tumor regression when a fraction of the tumor mass is genetically modified. Cancer Res. 53, 5274–5283.

    PubMed  CAS  Google Scholar 

  19. Sawant, S. G., Randers-Pehrson, G., Metting, N. F., and Hall, E. J. (2001) Adaptive response and the bystander effect induced by radiation in C3H 10T(1/2) cells in culture. Radiat. Res. 156, 177–180.

    Article  PubMed  CAS  Google Scholar 

  20. Kanwar, J. R., Kanwar, R. K., Pandey, S., Ching, L. M., and Krissansen, G. W. (2001) Vascular attack by 5,6-dimethylxanthenone-4-acetic acid combined with B7.1 (CD80)-mediated immunotherapy overcomes immune resistance and leads to the eradication of large tumors and multiple tumor foci. Cancer Res. 61, 1948–1956.

    PubMed  CAS  Google Scholar 

  21. Ali, S. A., McLean, C. S., Boursnell, M. E., et al. (2000) Preclinical evaluation of “whole” cell vaccines for prophylaxis and therapy using a disabled infectious single cycle-herpes simplex virus vector to transduce cytokine genes. Cancer Res. 60, 1663–1670.

    PubMed  CAS  Google Scholar 

  22. Freeman, S. M. (2000) Suicide gene therapy. Adv. Exp. Med. Biol. 465, 411–422.

    Article  PubMed  CAS  Google Scholar 

  23. Fife, K., Bower, M., Cooper, R. G., et al. (1998) Endothelial cell transfection with cationic liposomes and herpes simplex-thymidine kinase mediated killing. Gene Ther. 5, 614–620.

    Article  PubMed  CAS  Google Scholar 

  24. Jager, U., Zhao, Y., and Porter, C. D. (1999) Endothelial cell-specific transcriptional targeting from a hybrid long terminal repeat retrovirus vector containing human preproendothelin-1 promoter sequences. J. Virol. 73, 9702–9709.

    PubMed  CAS  Google Scholar 

  25. Ram, Z., Walbridge, S., Shawker, T., Culver, K. W., Blaese, R. M., and Oldfield, E. H. (1994) The effect of thymidine kinase transduction and ganciclovir therapy on tumor vasculature and growth of 9L gliomas in rats. J. Neurosurg. 81, 256–260.

    Article  PubMed  CAS  Google Scholar 

  26. Trinh, Q. T., Austin, E. A., Murray, D. M., Knick, V. C., and Huber, B. E. (1995) Enzyme/prodrug gene therapy: comparison of cytosine deaminase/5-fluorocytosine versus thymidine kinase/ganciclovir enzyme/prodrug systems in a human colorectal carcinoma cell line. Cancer Res. 55, 4808–4812.

    PubMed  CAS  Google Scholar 

  27. Denning, C. and Pitts, J. D. (1997) Bystander effects of different enzyme-prodrug systems for cancer gene therapy depend on different pathways for intercellular transfer of toxic metabolites, a factor that will govern clinical choice of appropriate regimes. Hum. Gene Ther. 8, 1825–1835.

    Article  PubMed  CAS  Google Scholar 

  28. Nishihara, E., Nagayama, Y., Narimatsu, M., Namba, H., Watanabe, M., Niwa, M., and Yamashita, S. (1998) Treatment of thyroid carcinoma cells with four different suicide gene/prodrug combinations in vitro. Anticancer Res. 18, 1521–1526.

    PubMed  CAS  Google Scholar 

  29. Chen, L. and Waxman, D. J. (1995) Intratumoral activation and enhanced chemotherapeutic effect of oxazaphosphorines following cytochrome P-450 gene transfer: development of a combined chemotherapy/cancer gene therapy strategy. Cancer Res. 55, 581–589.

    PubMed  CAS  Google Scholar 

  30. Hughes, B. W., Wells, A. H., Bebok, Z., et al. (1995) Bystander killing of melanoma cells using the human tyrosinase promoter to express the Escherichia coli purine nucleoside phosphorylase gene. Cancer Res. 55, 3339–3345.

    PubMed  CAS  Google Scholar 

  31. Patterson, A. V., Zhang, H., Moghaddam, A., et al. (1995) Increased sensitivity to the prodrug 5′-deoxy-5-fluorouridine and modulation of 5-fluoro-2′-deoxyuridine sensitivity in MCF-7 cells transfected with thymidine phosphorylase. Br. J. Cancer 72, 669–675.

    Article  PubMed  CAS  Google Scholar 

  32. Bridgewater, J. A., Knox, R. J., Pitts, J. D., Collins, M. K., and Springer, C. J. (1997) The bystander effect of the nitroreductase/CB1954 enzyme/prodrug system is due to a cell-permeable metabolite. Hum. Gene Ther. 8, 709–717.

    Article  PubMed  CAS  Google Scholar 

  33. Sutherland, R. M. (1988) Cell and environment interactions in tumor microregions: the multicell spheroid model. Science 240, 177–184.

    Article  PubMed  CAS  Google Scholar 

  34. O’Connor, K. C. (1999) Three-dimensional cultures of prostatic cells: tissue models for the development of novel anti-cancer therapies. Pharm. Res. 16, 486–493.

    Article  PubMed  Google Scholar 

  35. Roskelley, C. D. and Bissell, M. J. (1995) Dynamic reciprocity revisited: a continuous, bidirectional flow of information between cells and the extracellular matrix regulates mammary epithelial cell function. Biochem. Cell Biol. 73, 391–397.

    Article  PubMed  CAS  Google Scholar 

  36. Cukierman, E., Pankov, R., Stevens, D. R., and Yamada, K. M. (2001) Taking cell-matrix adhesions to the third dimension. Science 294, 1708–1712.

    Article  PubMed  CAS  Google Scholar 

  37. Durand, R. E. and Olive, P. L. (2001) Resistance of tumor cells to chemo-and radiotherapy modulated by the three-dimensional architecture of solid tumors and spheroids. Methods Cell Biol. 64, 211–233.

    Article  PubMed  CAS  Google Scholar 

  38. Cowan, D. S., Hicks, K. O., and Wilson, W. R. (1996) Multicellular membranes as an in vitro model for extravascular diffusion in tumours. Br. J. Cancer 27(Suppl), S28–S31.

    CAS  Google Scholar 

  39. Hicks, K. O., Ohms, S. J., van Zijl, P. L., Denny, W. A., Hunter, P. J., and Wilson, W. R. (1997) An experimental and mathematical model for the extravascular transport of a DNA intercalator in tumours. Br. J. Cancer 76, 894–903.

    Article  PubMed  CAS  Google Scholar 

  40. Hicks, K. O., Fleming, Y., Siim, B. G., Koch, C. J., and Wilson, W. R. (1998) Extravascular diffusion of tirapazamine: effect of metabolic consumption assessed using the multicellular layer model. Int. J. Radiat. Oncol. Biol. Phys. 42, 641–649.

    Article  PubMed  CAS  Google Scholar 

  41. Hicks, K. O., Pruijn, F. B., Baguley, B. C., and Wilson, W. R. (2001) Extravascular transport of the DNA intercalator and topoisomerase poison N-[2-(dimethylamino)ethyl]acridine-4-carboxamide (DACA): diffusion and metabolism in multicellular layers of tumor cells. J. Pharmacol. Exp. Ther. 297, 1088–1098.

    PubMed  CAS  Google Scholar 

  42. Baguley, B.C., Hicks, K.O., and Wilson, W.R. (2002) Tumour cell cultures in drug development, in Anticancer Drug Development (Baguley, B. C. and Kerr, D.J., eds.), Academic, San Diego, CA, pp. 269–284.

    Chapter  Google Scholar 

  43. Minchinton, A. I., Wendt, K. R., Clow, K. A., and Fryer, K. H. (1997) Multilayers of cells growing on a permeable support. An in vitro tumour model. Acta Oncol. 36, 13–16.

    Article  PubMed  CAS  Google Scholar 

  44. Phillips, R. M., Loadman, P. M., and Cronin, B. P. (1998) Evaluation of a novel in vitro assay for assessing drug penetration into avascular regions of tumours. Br. J. Cancer 77, 2112–2119.

    Article  PubMed  CAS  Google Scholar 

  45. Topp, E. M., Kitos, P. A., Vijaykumar, V., DeSilva, B. S., and Hendrickson, T. L. (1998) Antibody transport in cultured tumor cell layers. J. Control. Release 53, 15–23.

    Article  PubMed  CAS  Google Scholar 

  46. Kyle, A. H. and Minchinton, A. I. (1999) Measurement of delivery and metabolism of tirapazamine to tumour tissue using the multilayered cell culture model. Cancer Chemother. Pharmacol. 43, 213–220.

    Article  PubMed  CAS  Google Scholar 

  47. Chen, T. R. (1977) In situ detection of mycoplasma contamination in cell cultures by fluorescent Hoechst 33258 stain. Exp. Cell Res. 104, 255–262.

    Article  PubMed  CAS  Google Scholar 

  48. Kyle, A. H., Chan, C. T., and Minchinton, A. I. (1999) Characterization of three-dimensional tissue cultures using electrical impedance spectroscopy. Biophys. J. 76, 2640–2648.

    Article  PubMed  CAS  Google Scholar 

  49. Thanou, M., Nihot, M. T., Jansen, M., Verhoef, J. C., and Junginger, H. E. (2001) Mono-N-carboxymethyl chitosan (MCC), a polyampholytic chitosan derivative, enhances the intestinal absorption of low molecular weight heparin across intestinal epithelia in vitro and in vivo. J. Pharm. Sci. 90, 38–46.

    Article  PubMed  CAS  Google Scholar 

  50. Siim, B. G., Denny, W. A., and Wilson, W. R. (1997) Nitro reduction as an electronic switch for bioreductive drug activation. Oncol. Res. 9, 357–369.

    PubMed  CAS  Google Scholar 

  51. Friedlos, F., Denny, W. A., Palmer, B. D., and Springer, C. J. (1997) Mustard prodrugs for activation by Escherichia coli nitroreductase in gene-directed enzyme prodrug therapy. J. Med. Chem. 40, 1270–1275.

    Article  PubMed  CAS  Google Scholar 

  52. Kyriazis, A. A. and Kyriazis, A. P. (1980) Preferential sites of growth of human tumors in nude mice following subcutaneous transplantation. Cancer Res. 40, 4509–4511.

    PubMed  CAS  Google Scholar 

  53. Dipersio, L. P. (1981) Regional growth differences of human tumour xenografts in nude mice. Lab. Anim. 15, 179–180.

    Article  PubMed  CAS  Google Scholar 

  54. Wilson, K. M. and Lord, E. M. (1987) Specific (EMT6) and non-specific (WEHI-164) cytolytic activity by host cells infiltrating tumour spheroids. Br. J. Cancer 55, 141–146.

    Article  PubMed  CAS  Google Scholar 

  55. Hobbs, S., Jitrapakdee, S., and Wallace, J. C. (1998) Development of a bicistronic vector driven by the human polypeptide chain elongation factor 1alpha promoter for creation of stable mammalian cell lines that express very high levels of recombinant proteins. Biochem. Biophys. Res. Commun. 252, 368–372.

    Article  PubMed  CAS  Google Scholar 

  56. Skehan, P., Storeng, R., Scudiero, D., et al. (1990) New colorimetric cytotoxicity assay for anticancer-drug screening. J. Natl. Cancer Inst. 82, 1107–1112.

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Auckland Cancer Society Research Centre, The University of Auckland, Auckland, New Zealand

    William R. Wilson, Susan M. Pullen, Alison Hogg, Frederik B. Pruijn & Kevin O. Hicks

  2. Institute of Cancer Research, Sutton, Surrey, UK

    Stephen M. Hobbs

Authors
  1. William R. Wilson
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Susan M. Pullen
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Alison Hogg
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Stephen M. Hobbs
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Frederik B. Pruijn
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Kevin O. Hicks
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. Cancer Research UK Centre for Cancer Therapeutics at the Institute of Cancer Research, Sutton, Surrey, UK

    Caroline J. Springer

Rights and permissions

Reprints and permissions

Copyright information

© 2004 Humana Press Inc.

About this protocol

Cite this protocol

Wilson, W.R., Pullen, S.M., Hogg, A., Hobbs, S.M., Pruijn, F.B., Hicks, K.O. (2004). In Vitro and In Vivo Models for Evaluation of GDEPT. In: Springer, C.J. (eds) Suicide Gene Therapy. Methods in Molecular Medicine™, vol 90. Humana Press. https://doi.org/10.1385/1-59259-429-8:403

Download citation

  • DOI: https://doi.org/10.1385/1-59259-429-8:403

  • Publisher Name: Humana Press

  • Print ISBN: 978-0-89603-971-1

  • Online ISBN: 978-1-59259-429-0

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation