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Inhibition of nuclear delivery of plasmid DNA and transcription by interferon γ: hurdles to be overcome for sustained gene therapy

Abstract

Sustained expression of murine interferon (IFN)-γ (Muγ) was found to be effective in preventing tumor metastasis and atopic dermatitis in mouse models. However, our preliminary experiments suggested that the time-dependent decrease in the Muγ expression was not compensated for by repeated injections of Muγ-expressing plasmid. To identify the mechanism underlying this observation, a reporter plasmid was hydrodynamically injected into mice and the levels of the plasmid, mRNA and reporter protein were measured in mice receiving a pre- or co-administration of Muγ-expressing plasmid. Co-injection of Muγ-expressing plasmid had no significant effects on transgene expression from the reporter plasmid. In contrast, pre-injection of Muγ-expressing plasmid greatly inhibited the expression of the reporter protein. Moreover, pre-injection of Muγ-expressing plasmid also reduced the amount of the reporter plasmid in the nuclear fraction of mouse liver to <10%, and that of reporter mRNA to <1%. The degree of reduction in the expression of reporter protein was comparable with the reduction in mRNA. These results indicate that the difficulty in regaining the expression level of IFN-γ is due to the impaired delivery of plasmid to the nucleus and to the suppression of transcription from the plasmid.

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References

  1. Goodbourn S, Didcock L, Randall RE . Interferons: cell signalling, immune modulation, antiviral response and virus countermeasures. J Gen Virol 2000; 81: 2341–2364.

    Article  CAS  PubMed  Google Scholar 

  2. Samuel CE . Antiviral actions of interferons. Clin Microbiol Rev 2001; 14: 778–809.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  3. Grassegger A, Hopfl R . Significance of the cytokine interferon gamma in clinical dermatology. Clin Exp Dermatol 2004; 29: 584–588.

    Article  CAS  PubMed  Google Scholar 

  4. Smyth MJ, Cretney E, Kershaw MH, Hayakawa Y . Cytokines in cancer immunity and immunotherapy. Immunol Rev 2004; 202: 275–293.

    Article  CAS  PubMed  Google Scholar 

  5. Borden EC, Sen GC, Uze G, Silverman RH, Ransohoff RM, Foster GR et al. Interferons at age 50: past, current and future impact on biomedicine. Nat Rev Drug Discov 2007; 6: 975–990.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  6. Pepinsky RB, LePage DJ, Gill A, Chakraborty A, Vaidyanathan S, Green M et al. Improved pharmacokinetic properties of a polyethylene glycol-modified form of interferon-beta-1a with preserved in vitro bioactivity. J Pharmacol Exp Ther 2001; 297: 1059–1066.

    CAS  PubMed  Google Scholar 

  7. Palumbo E . PEG-interferon alpha-2b for acute hepatitis C: a review. Mini Rev Med Chem 2007; 7: 839–843.

    Article  CAS  PubMed  Google Scholar 

  8. Mata-Espinosa DA, Mendoza-Rodríguez V, Aguilar-León D, Rosales R, López-Casillas F, Hernández-Pando R . Therapeutic effect of recombinant adenovirus encoding interferon-gamma in a murine model of progressive pulmonary tuberculosis. Mol Ther 2008; 16: 1065–1072.

    Article  CAS  PubMed  Google Scholar 

  9. Shiau AL, Lin CY, Tzai TS, Wu CL . Postoperative immuno-gene therapy of murine bladder tumor by in vivo administration of retroviruses expressing mouse interferon-gamma. Cancer Gene Ther 2001; 8: 73–81.

    Article  CAS  PubMed  Google Scholar 

  10. Kobayashi N, Kuramoto T, Chen S, Watanabe Y, Takakura Y . Therapeutic effect of intravenous interferon gene delivery with naked plasmid DNA in murine metastasis models. Mol Ther 2002; 6: 737–744.

    Article  CAS  PubMed  Google Scholar 

  11. Kawano H, Nishikawa M, Mitsui M, Takahashi Y, Kako K, Yamaoka K et al. Improved anti-cancer effect of interferon gene transfer by sustained expression using CpG-reduced plasmid DNA. Int J Cancer 2007; 121: 401–406.

    Article  CAS  PubMed  Google Scholar 

  12. Mitsui M, Nishikawa M, Zang L, Ando M, Hattori K, Takahashi Y et al. Effect of the content of unmethylated CpG dinucleotides in plasmid DNA on the sustainability of transgene expression. J Gene Med 2009; 11: 435–443.

    Article  CAS  PubMed  Google Scholar 

  13. Liu F, Song Y, Liu D . Hydrodynamics-based transfection in animals by systemic administration of plasmid DNA. Gene Therapy 1999; 6: 1258–1266.

    Article  CAS  PubMed  Google Scholar 

  14. Hattori K, Nishikawa M, Watcharanurak K, Ikoma A, Kabashima K, Toyota H et al. Sustained exogenous expression of therapeutic levels of IFN-gamma ameliorates atopic dermatitis in NC/Nga mice via Th1 polarization. J Immunol 2010; 184: 2729–2735.

    Article  CAS  PubMed  Google Scholar 

  15. Nishikawa M, Nakayama A, Takahashi Y, Fukuhara Y, Takakura Y . Reactivation of silenced transgene expression in mouse liver by rapid, large-volume injection of isotonic solution. Hum Gene Ther 2008; 19: 1009–1020.

    Article  CAS  PubMed  Google Scholar 

  16. Qin L, Ding Y, Pahud DR, Chang E, Imperiale MJ, Bromberg JS . Promoter attenuation in gene therapy: interferon-gamma and tumor necrosis factor-alpha inhibit transgene expression. Hum Gene Ther 1997; 8: 2019–2029.

    Article  CAS  PubMed  Google Scholar 

  17. Harms JS, Splitter GA . Interferon-gamma inhibits transgene expression driven by SV40 or CMV promoters but augments expression driven by the mammalian MHC I promoter. Hum Gene Ther 1995; 6: 1291–1297.

    Article  CAS  PubMed  Google Scholar 

  18. Ghazizadeh S, Carroll JM, Taichman LB . Repression of retrovirus-mediated transgene expression by interferons: implications for gene therapy. J Virol 1997; 71: 9163–9169.

    CAS  PubMed  PubMed Central  Google Scholar 

  19. Acsadi G, O'Hagan D, Lochmüller H, Prescott S, Larochelle N, Nalbantoglu J et al. Interferons impair early transgene expression by adenovirus-mediated gene transfer in muscle cells. J Mol Med 1998; 76: 442–450.

    Article  CAS  PubMed  Google Scholar 

  20. Lui VW, Falo Jr LD, Huang L . Systemic production of IL-12 by naked DNA mediated gene transfer: toxicity and attenuation of transgene expression in vivo. J Gene Med 2001; 3: 384–393.

    Article  CAS  PubMed  Google Scholar 

  21. Sung RS, Qin L, Bromberg JS . TNFalpha and IFNgamma induced by innate anti-adenoviral immune responses inhibit adenovirus-mediated transgene expression. Mol Ther 2001; 3: 757–767.

    Article  CAS  PubMed  Google Scholar 

  22. Luo P, Reed BD, Tsang TC, Harris DT, Flavell RA . A self-augmenting gene expression cassette for enhanced and sustained transgene expression in the presence of proinflammatory cytokines. DNA Cell Biol 2006; 25: 659–667.

    Article  CAS  PubMed  Google Scholar 

  23. Reboredo M, Zabala M, Mauleon I, DeLas Rivas J, Kreppel F, Kochanek S et al. Interleukin-12 inhibits liver-specific drug-inducible systems in vivo. Gene Therapy 2008; 15: 277–288.

    Article  CAS  PubMed  Google Scholar 

  24. 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  PubMed  Google Scholar 

  25. Li S, Wu SP, Whitmore M, Loeffert EJ, Wang L, Watkins SC et al. Effect of immune response on gene transfer to the lung via systemic administration of cationic lipidic vectors. Am J Physiol 1999; 276: 796–804.

    Article  Google Scholar 

  26. Yasuda S, Yoshida H, Nishikawa M, Takakura Y . Comparison of the type of liposome involving cytokine production induced by non-CpG Lipoplex in macrophages. Mol Pharm 2010; 7: 533–542.

    Article  CAS  PubMed  Google Scholar 

  27. Yoshida H, Nishikawa M, Yasuda S, Mizuno Y, Toyota H, Kiyota T et al. TLR9-dependent systemic interferon-beta production by intravenous injection of plasmid DNA/cationic liposome complex in mice. J Gene Med 2009; 11: 708–717.

    Article  CAS  PubMed  Google Scholar 

  28. Yew NS, Zhao H, Wu IH, Song A, Tousignant JD, Przybylska M et al. Reduced inflammatory response to plasmid DNA vectors by elimination and inhibition of immunostimulatory CpG motifs. Mol Ther 2000; 1: 255–262.

    Article  CAS  PubMed  Google Scholar 

  29. Smolarczyk R, Cichoń T, Sochanik A, Szala S . Negligible induction of IFN-gamma, IL-12 and TNF-alpha by DNA-PEI 750 kDa/albumin complexes. Cytokine 2005; 29: 283–287.

    Article  CAS  PubMed  Google Scholar 

  30. Ramana CV, Grammatikakis N, Chernov M, Nguyen H, Goh KC, Williams BR et al. Regulation of c-myc expression by IFN-gamma through Stat1-dependent and -independent pathways. EMBO J 2000; 19: 263–272.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  31. Schaefer A, Unterberger C, Frankenberger M, Lohrum M, Staples KJ, Werner T et al. Mechanism of interferon-gamma mediated down-regulation of interleukin-10 gene expression. Mol Immunol 2009; 46: 1351–1359.

    Article  CAS  PubMed  Google Scholar 

  32. Kobayashi N, Nishikawa M, Hirata K, Takakura Y . Hydrodynamics-based procedure involves transient hyperpermeability in the hepatic cellular membrane: implication of a nonspecific process in efficient intracellular gene delivery. J Gene Med 2004; 6: 584–592.

    Article  CAS  PubMed  Google Scholar 

  33. Zhang G, Gao X, Song YK, Vollmer R, Stolz DB, Gasiorowski JZ et al. Hydroporation as the mechanism of hydrodynamic delivery. Gene Therapy 2004; 11: 675–682.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  34. Dean DA, Dean BS, Muller S, Smith LC . Sequence requirements for plasmid nuclear import. Exp Cell Res 1999; 253: 713–722.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  35. Roeder RG . The role of general initiation factors in transcription by RNA polymerase II. Trends Biochem Sci 1996; 21: 327–335.

    Article  CAS  PubMed  Google Scholar 

  36. Deng WG, Montero AJ, Wu KK . Interferon-γ suppresses cyclooxygenase-2 promoter activity by inhibiting C-Jun and C/EBPβ binding. Arterioscler Thromb Vasc Biol 2007; 27: 1752–1759.

    Article  CAS  PubMed  Google Scholar 

  37. Ganster RW, Guo Z, Shao L, Geller DA . Differential effects of TNF-alpha and IFN-gamma on gene transcription mediated by NF-kappaB-Stat1 interactions. J Interferon Cytokine Res 2005; 25: 707–719.

    Article  CAS  PubMed  Google Scholar 

  38. Shao L, Guo Z, Geller DA . Transcriptional suppression of cytokine-induced iNOS gene expression by IL-13 through IRF-1/ISRE signaling. Biochem Biophys Res Commun 2007; 362: 582–586.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  39. Nomura T, Yasuda K, Yamada T, Okamoto S, Mahato RI, Watanabe Y et al. Gene expression and antitumor effects following direct interferon (IFN)-gamma gene transfer with naked plasmid DNA and DC-chol liposome complexes in mice. Gene Therapy 1999; 6: 121–129.

    Article  CAS  PubMed  Google Scholar 

  40. Mir LM, Bureau MF, Gehl J, Rangara R, Rouy D, Caillaud JM et al. High-efficiency gene transfer into skeletal muscle mediated by electric pulses. Proc Natl Acad Sci USA 1999; 96: 4262–4267.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  41. Ochiai H, Harashima H, Kamiya H . Intranuclear disposition of exogenous DNA in vivo: silencing, methylation and fragmentation. FEBS Lett 2006; 580: 918–922.

    Article  CAS  PubMed  Google Scholar 

  42. Ochiai H, Fujimuro M, Yokosawa H, Harashima H, Kamiya H . Transient activation of transgene expression by hydrodynamics-based injection may cause rapid decrease in plasmid DNA expression. Gene Therapy 2007; 14: 1152–1159.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This work was supported in part by Grants-in-Aid for Scientific Research (B) and for Challenging Exploratory Research from the Ministry of Education, Culture, Sports, Science and Technology, Japan.

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Correspondence to Y Takakura.

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Zang, L., Nishikawa, M., Machida, K. et al. Inhibition of nuclear delivery of plasmid DNA and transcription by interferon γ: hurdles to be overcome for sustained gene therapy. Gene Ther 18, 891–897 (2011). https://doi.org/10.1038/gt.2011.35

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