siRNA and miRNA Gene Silencing pp 1-27

Part of the Methods in Molecular Biology book series (MIMB, volume 487)

Bacterial Delivery of siRNAs: A New Approach to Solid Tumor Therapy

  • De-Qi Xu
  • Ling Zhang
  • Dennis J Kopecko
  • Lifang Gao
  • Yueting Shao
  • Baofeng Guo
  • Lijing Zhao
Protocol

Abstract

RNAi is a powerful research tool for specific gene silencing and may also lead to promising novel therapeutic strategies. However, the development of RNAi-based therapies has been slow due to the lack of targeted delivery methods. The biggest challenge in the use of siRNA-based therapies is delivery to target cells. There are many additional obstacles to in vivo delivery of siRNAs, such as degradation by endogenous enzymes and interaction with blood components leading to nonspecific uptake into cells, which govern biodistribution and availability of siRNA in the body. Naked unmodified synthetic siRNA including plasmid-carried-shRNA-expression constructs cannot penetrate cellular membranes, and therefore, systemic application is unlikely to be successful. The success of gene therapy by siRNAs relies on the development of safe, economical, and efficacious in vivo delivery systems into the target cells. Attenuated Salmonella have been employed recently as vectors to deliver silencing hairpin RNA (shRNA) expression plasmids into mammalian cells. This approach has achieved gene silencing in vitro and in vivo. The facultative anaerobic, invasive Salmonella have a natural tropism for solid tumors including metastatic tumors. Genetically modified, attenuated Salmonella have been used recently both as potential antitumor agents by themselves, and to deliver specific tumoricidal therapies. This chapter describes the use of attenuated bacteria as tumor-targeting delivery systems for cancer therapy.

Keywords

Bacterial delivery vector Salmonella siRNA shRNA RNAi solid tumor 

References

  1. 1.
    Hannon, G.J. (2002). RNA interference. Nature 418, 244–251.CrossRefGoogle Scholar
  2. 2.
    Antoszczyk, S., Taira, K., and Kato, Y. (2006). Correlation of structure and activity of short hairpin RNA. Nucleic Acids Symp Ser (Oxf) 50, 295–296.CrossRefGoogle Scholar
  3. 3.
    Hiroaki, K.H. and Taira, K. (2003). Short hairpin type of dsRNAs that are controlled by tRNAVal. promoter significantly induce RNAi-mediated gene silencing in the cytoplasm of human cells Nucleic Acids Res. 31, 700–707.CrossRefGoogle Scholar
  4. 4.
    Zhang, L., Gao, L., Guo, B., et al. (2007). Intratumoral delivery and suppression of prostate tumor growth by attenuated Salmonella enterica. serovar Typhimurium carrying plasmid- based siRNAs Cancer Res. 67, 5859–5864.CrossRefGoogle Scholar
  5. 5.
    Bermudes, D., Zheng, L.M., and King, L.C. (2002). Live bacteria as anticancer agents and tumor-selective protein deliver vectors. Curr. Opin. Drug. Discov. Devel. 5, 194–199.Google Scholar
  6. 6.
    Tjuvajev, J, Blasberg, R, Luo, X, et al. (2001). Salmonella-based tumor-targeted cancer therapy: tumor amplified protein expression therapy (TAPET) for diagnostic imaging J. Control Release 74, 313–315.CrossRefGoogle Scholar
  7. 7.
    Zheng, L., Luo, X., Feng, M., et al. (2000). Tumor amplified protein expression therapy: Salmonella as a tumor-selective protein delivery vector. Oncol. Res. 12, 127–135.Google Scholar
  8. 8.
    Forbes, N.S., Munn, L.L., Fukumura, D., et al. (2003). Sparse initial entrapment of systemically injected Salmonella typhimurium. leads to heterogeneous accumulation within tumors Cancer Res. 63, 5188–5193.Google Scholar
  9. 9.
    Pawelek, J.M., Low, K.B., and Bermudes, D. (2003). Bacteria as tumour-targeting vectors. Lancet Oncol. 4, 548–556.CrossRefGoogle Scholar
  10. 10.
    Grosshans, H. and Slack, F.J. (2002). Micro-RNAs: small is plentiful. J. Cell Biol. 156, 17–21.CrossRefGoogle Scholar
  11. 11.
    Amarzguioui, M. and Prydz, H. (2004). An algorithm for selection of functional siRNA sequences. Biochem. Biophys. Res. Commun. 316, 1050–1058.CrossRefGoogle Scholar
  12. 12.
    Paul, C.P., Good, P.D., Winer, I., et al. (2002). Effective expression of small interfering RNA in human cells. Nat. Biotechnol. 20, 505–508.CrossRefGoogle Scholar
  13. 13.
    Brummelkamp, T.R., Bernards, R., and Agami, R. (2002). A system for stable expression of short interfering RNAs in mammalian cells. Science 296, 550–553.CrossRefGoogle Scholar
  14. 14.
    Brummelkamp, T.R., Bernards, R., and Agami, R (2002). Stable suppression of tumorigenicity by virus-mediated RNA interference. Cancer Cell 2, 243–247.CrossRefGoogle Scholar
  15. 15.
    Lambin, P., Theys, J., Landuyt, W., et al. (1998). Colonization of Clostridium in the body is restricted to hypoxic and necrotic areas of tumours. Anaerobe 4, 183–188.CrossRefGoogle Scholar
  16. 16.
    Luo, X., Li, Z., Lin, S., et al. (2000). Antitumor effect of VNP20009, an attenuated Salmonella in murine tumor models. Ocol. Res. 12, 501–508.Google Scholar
  17. 17.
    Jazowiecka-Rakus, J and Szala, S (2004). Antitumour activity of Salmonella typhimurium. VNP20047 in B16(F10) murine melanoma model Acta Biochim. Pol. 51, 851–856.Google Scholar
  18. 18.
    Khan, S.A., Everest, P., and Servos, S (1998). A lethal role for lipid A in Salmonella infections. Mol. Microbiol. 29, 571–579.CrossRefGoogle Scholar
  19. 19.
    Low, K.B., Ittensohn, M., Le, T., et al. (1999). Lipid A mutant salmonella with suppressed virulence and TNF-α induction retain tumor-targeting in vivo. Nat. Biotech. 17, 3–41.CrossRefGoogle Scholar
  20. 20.
    Miller, S.I., Kukral, A.M., and Mekalanos, J.J. (1989). A two-component regulatory system (phoP phoQ) controls Salmonella-typhimurium virulence. Proc. Natl. Acad. Sci. USA 86, 5054–5058.CrossRefGoogle Scholar
  21. 21.
    Ronson, C.W., Nixon, B.T., and Ausubel, F.M. (1987). Conserved domains in Bacterial regulatory proteins that respond to environmental stimuli. Cell 49, 579–581.CrossRefGoogle Scholar
  22. 22.
    Ernst, R.K., Guina, T., and Miller, S.I. (1999). How intracellular bacteria, survive: surface modifications that promote resistance to host innate immune responses. J. Infect. Dis. 179, S326–S330.CrossRefGoogle Scholar
  23. 23.
    VanCott, J.L., Chatfield, S.N., Roberts, M., et al. (1998). Regulation of host immune responses by modification of Salmonella virulence genes. Nat. Med. 4, 1247–1252.CrossRefGoogle Scholar
  24. 24.
    Guo, L., Lim, K.B., Gunn, J.S., et al. (1997). Regulation of lipid A modifications by Salmonella typhimurium. virulence genes phoP-phoQ Science 276, 250–253.CrossRefGoogle Scholar
  25. 25.
    Dang, L.H., Bettegowda, C., Huso, D.L., et al. (2001). Combination bacteriolytic therapy for the treatment of experimental tumors. Proc. Natl. Acad. Sci. U S A 98, 15155–15160.CrossRefGoogle Scholar
  26. 26.
    Low, K.B., Ittensohn, M., Luo, X., et al. (2004). Construction of VNP20009: A novel, genetically stable antibiotic-sensitive strain of tumor-targeting Salmonella. for parenteral administation in humans Methods Mol. Med. 90, 47–60.Google Scholar
  27. 27.
    Hohmann, E.L, Oletta, C.A., Killeen, K.P., et al. (1996). phoP/phoQ-deleted Salmonella typhi. (Ty800) is a safe and immunogenic single-dose typhoid fevr vaccine in volunteers J. Infect. Dis. 173, 1948–1014.CrossRefGoogle Scholar
  28. 28.
    Hohmann, E.L., Oletta, C.A., and Miller, S.I. (1996). Evaluation of a phoP/phoQ-deleted, aroA-deleted live oral Salmonella typhi vaccine strain in human volunteers. Vaccine 14, 19–24.CrossRefGoogle Scholar
  29. 29.
    Koslowski, J.M., Fidler, I.J., Campbell, D., et al. (1984). Metastatic behavior of human tumor cell lines grown in the nude mice. Cancer Res. 44, 3522–3529.Google Scholar
  30. 30.
    Hoffman, R.M. (1999). Orthotopic metastatic mouse models for anticancer discovery and evaluation: a bridge to the clinic. Invest. New Drugs 17, 343–359.CrossRefGoogle Scholar
  31. 31.
    Davies, J. and Jimenez, A. (1980). A new selective agent for eukayotic cloning vector. Am. J. Trop. Med. Hyg. 29, (5 Suppl.)1089–1092.Google Scholar
  32. 32.
    Bar-Nun, S., et al. (1983). G-418, an enlongation inhibitor of 80S ribosomes. Biochem. Biophys. Acta 741, 123–127.Google Scholar

Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • De-Qi Xu
    • 1
  • Ling Zhang
    • 1
  • Dennis J Kopecko
    • 1
  • Lifang Gao
    • 1
  • Yueting Shao
    • 1
  • Baofeng Guo
    • 1
  • Lijing Zhao
    • 1
  1. 1.FDA HFM 440, Bldg. 29/424NIH CampusBethesdaMD

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