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.

  • Original Article
  • Published:

Evaluation of in vivo antitumor effects of ANT2 shRNA delivered using PEI and ultrasound with microbubbles

Gene Therapy volumeĀ 22,Ā pages 325ā€“332 (2015)Cite this article

Subjects

Abstract

Gene therapy using RNA interference can be directed against tumors through various strategies, but has been hindered owing to the inefficiency of non-viral delivery. To evaluate the antitumor effects of adenine nucleotide translocase-2 (ANT2) short hairpin RNA (shRNA) by intraperitoneal injection using the polyethylenimine (PEI) and an ultrasound gene delivery method, human breast carcinoma MDA-MB-231 cells were injected subcutaneously into NOG (NOD/Shi-scid/IL-2RĪ³null) mice. The results showed greater tumor regression (*P<0.05) as well as an increased survival rate in the group receiving ANT2 shRNA+two types of enhancer relative to the groups receiving ANT2 shRNA without enhancer. These findings demonstrate that the introduction of PEI and ultrasound with SonoVue exerted enhanced antitumor effects in vivo. Although the combination of jet-PEI and ultrasound provided the best results with respect to tumor regression, the antitumor effects from the individual enhancers were approximately equivalent. In addition, we confirmed that there was no toxicity on aspartate aminotransferase and alanine aminotransferase levels in the liver and albumin, blood urea nitrogen or creatine kinase levels in the kidney following the various gene delivery methods.

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

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

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

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6

Similar content being viewed by others

References

  1. Lin C-R, Chen K-H, Yang C-H, Cheng J-T, Sheen-Chen S-M, Wu C-H et al. Sonoporation-mediated gene transfer into adult rat dorsal root ganglion cells. J Biomed Sci 2010; 17: 44.

    ArticleĀ  Google ScholarĀ 

  2. Bekeredjian R, Grayburn PA, Shohet RV . Use of ultrasound contrast agents for gene or drug delivery in cardiovascular medicine. J Am Coll Cardiol 2005; 45: 329ā€“335.

    ArticleĀ  CASĀ  Google ScholarĀ 

  3. Shen Z-Y, Hu B . Low-frequency low-intensity ultrasound with contrast agent for the treatment of subcutaneous tumors in mice. Sci Res Essays 2011; 6: 5579ā€“5585.

    Google ScholarĀ 

  4. Zhang L, Yang N, Mohamed-Hadley A, Rubin SC, Coukos G . Vector-based RNAi a novel tool for isoform-specific knock-down of VEGF and anti-angiogenesis gene therapy of cancer. Biochem Biophys Res Commun 2003; 303: 1169ā€“1178.

    ArticleĀ  CASĀ  Google ScholarĀ 

  5. Urban-Klein B, Werth S, Abuharbeid S, Czubayko F, Aigner A . RNAi-mediated gene-targeting through systemic application of polyethylenimine (PEI)-complexed siRNA in vivo. Gene Therapy 2005; 12: 461ā€“466.

    ArticleĀ  CASĀ  Google ScholarĀ 

  6. Jang J-Y, Choi Y, Jeon Y-K, Kim C-W . Suppression of adenine nucleotide translocase-2 by vector-based siRNA in human breast cancer cells induces apoptosis and inhibits tumor growth in vitro and in vivo. Breast Cancer Res 2008; 10: R11.

    ArticleĀ  Google ScholarĀ 

  7. Jang J-Y, Jeon Y-K, Kim C-W . Degradation of HER2/neu by ANT2 shRNA suppresses migration and invasiveness of breast cancer cells. BMC Cancer 2010; 10: 391.

    ArticleĀ  Google ScholarĀ 

  8. Choi Y, Jeon YH, Jang J-Y, Chung J-K, Kim C-W . Treatment With mANT2 shRNA enhances antitumor therapeutic effects induced by MUC1 DNA vaccination. Mol Ther 2011; 19: 979ā€“989.

    ArticleĀ  CASĀ  Google ScholarĀ 

  9. JƤnicke RU . MCF-7 breast carcinoma cells do not express caspase-3. Breast Cancer Res Treat 2009; 117: 219ā€“221.

    ArticleĀ  Google ScholarĀ 

  10. Doerner A, Pauschinger M, Badorff A, Noutsias M, Giessen S, Schulze K et al. Tissue-specific transcription pattern of the adenine nucleotide translocase isoforms in humans. FEBS Lett 1997; 414: 258ā€“262.

    ArticleĀ  CASĀ  Google ScholarĀ 

  11. Luciakova K, Barath P, Poliakova D, Persson A, Nelson BD . Repression of the human adenine nucleotide translocase-2 gene in growth-arrested human diploid cells the role of nuclear factor-1. J Biol Chem 2003; 278: 30624ā€“30633.

    ArticleĀ  CASĀ  Google ScholarĀ 

  12. Miller DL, Pislaru SV, Greenleaf JF . Sonoporation: mechanical DNA delivery by ultrasonic cavitation. Somatic Cell Mol Genet 2002; 27: 115ā€“134.

    ArticleĀ  CASĀ  Google ScholarĀ 

  13. Thomas M, Klibanov A . Non-viral gene therapy: polycation-mediated DNA delivery. Appl Microbiol Biotechnol 2003; 62: 27ā€“34.

    ArticleĀ  CASĀ  Google ScholarĀ 

  14. Zhang Y, Satterlee A, Huang L . In vivo gene delivery by nonviral vectors: overcoming hurdles & quest. Mol Ther 2012; 20: 1298ā€“1304.

    ArticleĀ  CASĀ  Google ScholarĀ 

  15. Li S, Huang L . Nonviral gene therapy: promises and challenges. Gene Therapy 2000; 7: 31ā€“34.

    ArticleĀ  CASĀ  Google ScholarĀ 

  16. Aihara H, Miyazaki J-i . Gene transfer into muscle by electroporation in vivo. Nat Biotechnol 1998; 16: 867ā€“870.

    ArticleĀ  CASĀ  Google ScholarĀ 

  17. Yoshida A, Nagata T, Uchijima M, Higashi T, Koide Y . Advantage of gene gun-mediated over intramuscular inoculation of plasmid DNA vaccine in reproducible induction of specific immune responses. Vaccine 2000; 18: 1725ā€“1729.

    ArticleĀ  CASĀ  Google ScholarĀ 

  18. Huth S, Lausier J, Gersting SW, Rudolph C, Plank C, Welsch U et al. Insights into the mechanism of magnetofection using PEIā€based magnetofectins for gene transfer. J Gene Med 2004; 6: 923ā€“936.

    ArticleĀ  CASĀ  Google ScholarĀ 

  19. Newman C, Bettinger T . Gene therapy progress and prospects: ultrasound for gene transfer. Gene Therapy 2007; 14: 465ā€“475.

    ArticleĀ  CASĀ  Google ScholarĀ 

  20. Erbacher P, Bettinger T, Belguiseā€Valladier P, Zou S, Coll JL, Behr JP et al. Transfection and physical properties of various saccharide, poly (ethylene glycol), and antibodyā€derivatized polyethylenimines (PEI). J Gene Med 1999; 1: 210ā€“222.

    ArticleĀ  CASĀ  Google ScholarĀ 

  21. Aoki K, Furuhata S, Hatanaka K, Maeda M, Remy J, Behr J et al. Polyethylenimine-mediated gene transfer into pancreatic tumor dissemination in the murine peritoneal cavity. Gene Therapy 2001; 8: 504ā€“514.

    ArticleĀ  Google ScholarĀ 

  22. Werth S, Urban-Klein B, Dai L, Hƶbel S, Grzelinski M, Bakowsky U et al. A low molecular weight fraction of polyethylenimine (PEI) displays increased transfection efficiency of DNA and siRNA in fresh or lyophilized complexes. J Control Rel 2006; 112: 257ā€“270.

    ArticleĀ  CASĀ  Google ScholarĀ 

  23. Moret I, Esteban Peris J, Guillem VM, Benet M, Revert F, DasĆ­ F et al. Stability of PEIā€“DNA and DOTAPā€“DNA complexes: effect of alkaline pH, heparin and serum. J Control Rel 2001; 76: 169ā€“181.

    ArticleĀ  CASĀ  Google ScholarĀ 

  24. Saito M, Mazda O, Takahashi KA, Arai Y, Kishida T, Shinā€Ya M et al. Sonoporation mediated transduction of pDNA/siRNA into joint synovium in vivo. J Orthop Res 2007; 25: 1308ā€“1316.

    ArticleĀ  CASĀ  Google ScholarĀ 

  25. Xenariou S, Griesenbach U, Liang H, Zhu J, Farley R, Somerton L et al. Use of ultrasound to enhance nonviral lung gene transfer in vivo. Gene Therapy 2007; 14: 768ā€“774.

    ArticleĀ  CASĀ  Google ScholarĀ 

  26. Dang S-p, Wang R-x, Qin M-d, Zhang Y, Gu Y-z, Wang M-y et al. A novel transfection method for eukaryotic cells using polyethylenimine coated albumin microbubbles. Plasmid 2011; 66: 19ā€“25.

    ArticleĀ  CASĀ  Google ScholarĀ 

  27. Phillips LC, Klibanov AL, Wamhoff BR, Hossack JA . Targeted gene transfection from microbubbles into vascular smooth muscle cells using focused, ultrasound-mediated delivery. Ultrasound Med Biol 2010; 36: 1470ā€“1480.

    ArticleĀ  Google ScholarĀ 

  28. Liu J, Lewis TN, Prausnitz MR . Non-invasive assessment and control of ultrasound-mediated membrane permeabilization. Pharm Res 1998; 15: 918ā€“924.

    ArticleĀ  CASĀ  Google ScholarĀ 

  29. Wei W, Zheng-zhong B, Yong-jie W, Qing-wu Z, Ya-lin M . Bioeffects of low-frequency ultrasonic gene delivery and safety on cell membrane permeability control. J Ultrasound Med 2004; 23: 1569ā€“1582.

    ArticleĀ  Google ScholarĀ 

  30. Zhou Y, Cui J, Deng CX . Dynamics of sonoporation correlated with acoustic cavitation activities. Biophys J 2008; 94: L51ā€“L53.

    ArticleĀ  CASĀ  Google ScholarĀ 

  31. Aigner A, Fischer D, Merdan T, Brus C, Kissel T, Czubayko F . Delivery of unmodified bioactive ribozymes by an RNA-stabilizing polyethylenimine (LMW-PEI) efficiently down-regulates gene expression. Gene Therapy 2002; 9: 1700ā€“1707.

    ArticleĀ  CASĀ  Google ScholarĀ 

  32. Intra J, Salem AK . Characterization of the transgene expression generated by branched and linear polyethylenimine-plasmid DNA nanoparticles in vitro and after intraperitoneal injection in vivo. J Control Rel 2008; 130: 129ā€“138.

    ArticleĀ  CASĀ  Google ScholarĀ 

  33. Louis M, Dutoit S, Denoux Y, Erbacher P, Deslandes E, Behr J et al. Intraperitoneal linear polyethylenimine (L-PEI)-mediated gene delivery to ovarian carcinoma nodes in mice. Cancer Gene Ther 2006; 13: 367ā€“374.

    ArticleĀ  CASĀ  Google ScholarĀ 

  34. Dou S, Smith M, Wang Y, Rusckowski M, Liu G . Intraperitoneal injection is not always a suitable alternative to intravenous injection for radiotherapy. Cancer Biother Radiopharm 2013; 28: 335ā€“342.

    ArticleĀ  CASĀ  Google ScholarĀ 

  35. Boussif O, Lezoualc'h F, Zanta MA, Mergny MD, Scherman D, Demeneix B et al. A versatile vector for gene and oligonucleotide transfer into cells in culture and in vivo: polyethylenimine. Proc Natl Acad Sci USA 1995; 92: 7297ā€“7301.

    ArticleĀ  CASĀ  Google ScholarĀ 

  36. Demeneix B, Behr JP . Polyethylenimine (PEI). Adv Genet 2005; 53: 215ā€“230.

    ArticleĀ  Google ScholarĀ 

  37. Williams A . A possible alteration in the permeability of ascites cell membranes after exposure to acoustic microstreaming. J Cell Sci 1973; 12: 875ā€“885.

    CASĀ  PubMedĀ  Google ScholarĀ 

  38. Wu J, Nyborg WL . Ultrasound, cavitation bubbles and their interaction with cells. Adv Drug Deliv Rev 2008; 60: 1103ā€“1116.

    ArticleĀ  CASĀ  Google ScholarĀ 

  39. Okada K, Kudo N, Niwa K, Yamamoto K . A basic study on sonoporation with microbubbles exposed to pulsed ultrasound. J Med Ultrason 2005; 32: 3ā€“11.

    ArticleĀ  Google ScholarĀ 

  40. Forbes MM, Steinberg RL . Examination of inertial cavitation of Optison in producing sonoporation of Chinese hamster ovary cells. Ultrasound Med Biol 2008; 34: 2009ā€“2018.

    ArticleĀ  Google ScholarĀ 

  41. Niidome T, Huang L . Gene therapy progress and prospects: nonviral vectors. Gene Therapy 2002; 9: 1647ā€“1652.

    ArticleĀ  CASĀ  Google ScholarĀ 

  42. Herweijer H, Wolff J . Progress and prospects: naked DNA gene transfer and therapy. Gene Therapy 2003; 10: 453ā€“458.

    ArticleĀ  CASĀ  Google ScholarĀ 

  43. Atici S, Cinel I, Cinel L, Doruk N, Eskandari G, Oral U . Liver and kidney toxicity in chronic use of opioids: an experimental long term treatment model. J Biosci 2005; 30: 245ā€“252.

    ArticleĀ  CASĀ  Google ScholarĀ 

  44. Zhang L, Liu Y, Xiang G, Lv Q, Huang G, Yang Y et al. Ultrasound-triggered microbubble destruction in combination with cationic lipid microbubbles enhances gene delivery. J Huazhong Univ Sci Technol [Medical Sciences] 2011; 31: 39ā€“45.

    ArticleĀ  Google ScholarĀ 

  45. Shen Z, Brayman A, Chen L, Miao C . Ultrasound with microbubbles enhances gene expression of plasmid DNA in the liver via intraportal delivery. Gene Therapy 2008; 15: 1147ā€“1155.

    ArticleĀ  CASĀ  Google ScholarĀ 

  46. Ferrara K, Pollard R, Borden M . Ultrasound microbubble contrast agents: fundamentals and application to gene and drug delivery. Annu Rev Biomed Eng 2007; 9: 415ā€“447.

    ArticleĀ  CASĀ  Google ScholarĀ 

  47. Song S, Shen Z, Chen L, Brayman A, Miao C . Explorations of high-intensity therapeutic ultrasound and microbubble-mediated gene delivery in mouse liver. Gene Therapy 2011; 18: 1006ā€“1014.

    ArticleĀ  CASĀ  Google ScholarĀ 

  48. Ito M, Hiramatsu H, Kobayashi K, Suzue K, Kawahata M, Hioki K et al. NOD/SCID/Ī³ mouse: an excellent recipient mouse model for engraftment of human cells. Blood 2002; 100: 3175ā€“3182.

    ArticleĀ  CASĀ  Google ScholarĀ 

  49. Leighton T . The Acoustic Bubble. Academic Press: New York, NY, USA, 1994.

    Google ScholarĀ 

  50. Schneider M . Characteristics of SonoVueā„¢. Echocardiography 1999; 16: 743ā€“746.

    ArticleĀ  Google ScholarĀ 

Download references

Acknowledgements

This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Ministry of Science, ICT & Future Planning (MSIP) (NRF-2013R1A2A2A04016262), (2012-0001190), (M1AXA003-2011-0032035), and (2010-00757), Republic of Korea.

Author information

Author notes

  1. D H Park and B K Jung: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Pathology, Tumor Immunity Medical Research Center, Cancer Research Institute, Seoul National University College of Medicine, Seoul, Republic of Korea

    D H Park,Ā B K Jung,Ā Y S Lee,Ā J Y Jang,Ā M K Kim,Ā J K LeeĀ &Ā C W Kim

  2. Department of Parasitology and Tropical Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea

    B K Jung

  3. School of Electronic Electrical Engineering, Sungkyunkwan University, Suwon, Republic of Korea

    H Park

  4. Department of Biomedical Engineering, Yonsei University, Wonju, Gangwon-do, Republic of Korea

    J Seo

Authors
  1. D H Park
    View author publications

    You can also search for this author in PubMedĀ Google Scholar

  2. B K Jung
    View author publications

    You can also search for this author in PubMedĀ Google Scholar

  3. Y S Lee
    View author publications

    You can also search for this author in PubMedĀ Google Scholar

  4. J Y Jang
    View author publications

    You can also search for this author in PubMedĀ Google Scholar

  5. M K Kim
    View author publications

    You can also search for this author in PubMedĀ Google Scholar

  6. J K Lee
    View author publications

    You can also search for this author in PubMedĀ Google Scholar

  7. H Park
    View author publications

    You can also search for this author in PubMedĀ Google Scholar

  8. J Seo
    View author publications

    You can also search for this author in PubMedĀ Google Scholar

  9. C W Kim
    View author publications

    You can also search for this author in PubMedĀ Google Scholar

Corresponding authors

Correspondence to J Seo or C W Kim.

Ethics declarations

Competing interests

The authors declare no conflict of interest.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Park, D., Jung, B., Lee, Y. et al. Evaluation of in vivo antitumor effects of ANT2 shRNA delivered using PEI and ultrasound with microbubbles. Gene Ther 22, 325ā€“332 (2015). https://doi.org/10.1038/gt.2014.120

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/gt.2014.120

This article is cited by

Search

Advanced search

Quick links