Abstract
In vivo electroporation (EP) is an efficient method for effective gene transfer and is highly expected for application in anticancer gene therapy. Non-invasive monitoring of gene transfer/expression is critical for optimal gene therapy. Here we report in vivo optical and high-field magnetic resonance imaging (MRI) of EP-mediated transgene expression in a tumor model. Initially, we observed spatio-temporal change in in vivo EP-mediated transgene expression by optical imaging using red fluorescence protein (RFP) as a reporter gene. Next, we constructed a dual-reporter plasmid carrying a gene-encoding MRI reporter ferritin heavy chain and RFP gene to visualize the intratumoral transgene expression by dual modality. Cells transfected with this plasmid showed lower signal intensity on in vitro T2-weighted cellular MRI and quantitatively increased the transverse relaxation rate (1/T2) compared with control cells. After conducting in vivo EP in an experimental tumor, the plasmid-injected region showed both fluorescent emissions in optical imaging and detectably lowered signal on T2-weighted MRI. The correlative immunohistological findings confirmed that both the reporter transgenes were co-expressed in this region. Thus, our strategy provides a platform for evaluating EP-mediated cancer gene therapy easily and safely without administering contrast agent or substrate.
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References
Alexander BL, Ali RR, Alton EW, Bainbridge JW, Braun S, Cheng SH et al. Progress and prospects: gene therapy clinical trials (part 1). Gene Therapy 2007; 14: 1439–1447.
Aiuti A, Bachoud-Levi AC, Blesch A, Brenner MK, Cattaneo F, Chiocca EA et al. Progress and prospects: gene therapy clinical trials (part 2). Gene Therapy 2007; 14: 1555–1563.
Porteus MH, Connelly JP, Pruett SM . A look to future directions in gene therapy research for monogenic diseases. PLoS Genet 2006; 2: e133.
Payen E, Bettan M, Rouyer-Fessard P, Beuzard Y, Scherman D . Improvement of mouse beta-thalassemia by electrotransfer of erythropoietin cDNA. Exp Hematol 2001; 29: 295–300.
Argyros O, Wong SP, Niceta M, Waddington SN, Howe SJ, Coutelle C et al. Persistent episomal transgene expression in liver following delivery of a scaffold/matrix attachment region containing non-viral vector. Gene Therapy 2008; 15: 1593–1605.
Daud AI, DeConti RC, Andrews S, Urbas P, Riker AI, Sondak VK et al. Phase I trial of interleukin-12 plasmid electroporation in patients with metastatic melanoma. J Clin Oncol 2008; 26: 5896–5903.
Shimao K, Takayama T, Enomoto K, Saito T, Nagai S, Miyazaki J et al. Cancer gene therapy using in vivo electroporation of Flt3-ligand. Int J Oncol 2005; 27: 457–463.
Kaiga T, Sato M, Kaneda H, Iwakura Y, Takayama T, Tahara H . Systemic administration of IL-23 induces potent antitumor immunity primarily mediated through Th1-type response in association with the endogenously expressed IL-12. J Immunol 2007; 178: 7571–7580.
Golzio M, Mazzolini L, Ledoux A, Paganin A, Izard M, Hellaudais L et al. In vivo gene silencing in solid tumors by targeted electrically mediated siRNA delivery. Gene Therapy 2007; 14: 752–759.
Isaka Y, Imai E . Electroporation-mediated gene therapy. Expert Opin Drug Deliv 2007; 4: 561–571.
Goto T, Nishi T, Tamura T, Dev SB, Takeshima H, Kochi M et al. Highly efficient electro-gene therapy of solid tumor by using an expression plasmid for the herpes simplex virus thymidine kinase gene. Proc Natl Acad Sci USA 2000; 97: 354–359.
Lohr F, Lo DY, Zaharoff DA, Hu K, Zhang X, Li Y et al. Effective tumor therapy with plasmid-encoded cytokines combined with in vivo electroporation. Cancer Res 2001; 61: 3281–3284.
Tamura T, Nishi T, Goto T, Takeshima H, Dev SB, Ushio Y et al. Intratumoral delivery of interleukin 12 expression plasmids with in vivo electroporation is effective for colon and renal cancer. Hum Gene Therapy 2001; 12: 1265–1276.
Lucas ML, Heller L, Coppola D, Heller R . IL-12 plasmid delivery by in vivo electroporation for the successful treatment of established subcutaneous B16.F10 melanoma. Mol Ther 2002; 5: 668–675.
Matsubara H, Mizutani Y, Hongo F, Nakanishi H, Kimura Y, Ushijima S et al. Gene therapy with TRAIL against renal cell carcinoma. Mol Cancer Ther 2006; 5: 2165–2171.
Massoud TF, Gambhir SS . Molecular imaging in living subjects: seeing fundamental biological processes in a new light. Genes Dev 2003; 17: 545–580.
Golzio M, Rols MP, Gabriel B, Teissie J . Optical imaging of in vivo gene expression: a critical assessment of the methodology and associated technologies. Gene Ther 2004; 11 (Suppl 1): S85–S91.
Inubushi M, Wu JC, Gambhir SS, Sundaresan G, Satyamurthy N, Namavari M et al. Positron-emission tomography reporter gene expression imaging in rat myocardium. Circulation 2003; 107: 326–332.
Serganova I, Blasberg R . Reporter gene imaging: potential impact on therapy. Nucl Med Biol 2005; 32: 763–780.
Furukawa T, Lohith TG, Takamatsu S, Mori T, Tanaka T, Fujibayashi Y . Potential of the FES-hERL PET reporter gene system—basic evaluation for gene therapy monitoring. Nucl Med Biol 2006; 33: 145–151.
Siddiqui F, Barton KN, Stricker HJ, Steyn PF, Larue SM, Karvelis KC et al. Design considerations for incorporating sodium iodide symporter reporter gene imaging into prostate cancer gene therapy trials. Hum Gene Ther 2007; 18: 312–322.
Niwa H, Yamamura K, Miyazaki J . Efficient selection for high-expression transfectants with a novel eukaryotic vector. Gene 1991; 108: 193–199.
Brooks AR, Harkins RN, Wang P, Qian HS, Liu P, Rubanyi GM . Transcriptional silencing is associated with extensive methylation of the CMV promoter following adenoviral gene delivery to muscle. J Gene Med 2004; 6: 395–404.
Maruyama H, Ataka K, Higuchi N, Sakamoto F, Gejyo F, Miyazaki J . Skin-targeted gene transfer using in vivo electroporation. Gene Therapy 2001; 8: 1808–1812.
Takei Y, Nemoto T, Mu P, Fujishima T, Ishimoto T, Hayakawa Y et al. In vivo silencing of a molecular target by short interfering RNA electroporation: tumor vascularization correlates to delivery efficiency. Mol Cancer Ther 2008; 7: 211–221.
Pedron-Mazoyer S, Plouet J, Hellaudais L, Teissie J, Golzio M . New anti angiogenesis developments through electro-immunization: optimization by in vivo optical imaging of intradermal electro gene transfer. Biochim Biophys Acta 2007; 1770: 137–142.
Bloquel C, Trollet C, Pradines E, Seguin J, Scherman D, Bureau MF . Optical imaging of luminescence for in vivo quantification of gene electrotransfer in mouse muscle and knee. BMC Biotechnol 2006; 6: 16.
Heller LC, Ugen K, Heller R . Electroporation for targeted gene transfer. Expert Opin Drug Deliv 2005; 2: 255–268.
Genove G, DeMarco U, Xu H, Goins WF, Ahrens ET . A new transgene reporter for in vivo magnetic resonance imaging. Nat Med 2005; 11: 450–454.
Cohen B, Dafni H, Meir G, Harmelin A, Neeman M . Ferritin as an endogenous MRI reporter for noninvasive imaging of gene expression in C6 glioma tumors. Neoplasia 2005; 7: 109–117.
Cohen B, Ziv K, Plaks V, Israely T, Kalchenko V, Harmelin A et al. MRI detection of transcriptional regulation of gene expression in transgenic mice. Nat Med 2007; 13: 498–503.
Aung W, Hasegawa S, Furukawa T, Saga T . Potential role of ferritin heavy chain in oxidative stress and apoptosis in human mesothelial and mesothelioma cells: implications for asbestos-induced oncogenesis. Carcinogenesis 2007; 28: 2047–2052.
Gilad AA, Winnard Jr PT, van Zijl PC, Bulte JW . Developing MR reporter genes: promises and pitfalls. NMR Biomed 2007; 20: 275–290.
Ngoi SM, Chien AC, Lee CG . Exploiting internal ribosome entry sites in gene therapy vector design. Curr Gene Ther 2004; 4: 15–31.
Wilkinson JT, Di X, Schonig K, Buss JL, Kock ND, Cline JM et al. Tissue-specific expression of ferritin H regulates cellular iron homoeostasis in vivo. Biochem J 2006; 395: 501–507.
Weissleder R, Pittet MJ . Imaging in the era of molecular oncology. Nature 2008; 452: 580–589.
Acknowledgements
We thank Dr Paolo Arosio (University of Brescia, Italy) for the generous gift of ferritin cDNA; Dr Jun-ichi Miyazaki (Osaka University, Japan) for the generous donation of pCAGGS vector; Kosho Suzuki (NEPA GENE) for technical suggestions for in vivo EP; Michiko Narazaki, Misao Yoneyama, Sayaka Shibata and Shigeyoshi Saito for skillful assistance with the MRI experiments; Hikaru Takenaka for preparation of the paper; members in our group for helpful discussions; Drs Yasuhito Sasaki, Yoshiharu Yonekura and Shuji Tanada for encouragements for this study. This study was supported in part by a Grant-in-Aid for Scientific Research for Exploratory Research from the Ministry of Education, Culture, Sports, Science and Technology, Japan (18659357: S Hasegawa); Intramural Research Grant from the National Institute of Radiological Sciences (S Hasegawa).
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Aung, W., Hasegawa, S., Koshikawa-Yano, M. et al. Visualization of in vivo electroporation-mediated transgene expression in experimental tumors by optical and magnetic resonance imaging. Gene Ther 16, 830–839 (2009). https://doi.org/10.1038/gt.2009.55
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DOI: https://doi.org/10.1038/gt.2009.55
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