THE JOURNAL OF GENE MEDICINE RESEARCH ARTICLE J Gene Med 2009; 11: 1125–1137. Published online 23 September 2009 in Wiley InterScience (www.interscience.wiley.com) DOI: 10.1002/jgm.1403 Controlled systemic release of interleukin-12 after gene electrotransfer to muscle for cancer gene therapy alone or in combination with ionizing radiation in murine sarcomas Gregor Tevz 1 Simona Kranjc 1 Maja Cemazar 1 Urska Kamensek 1 Andrej Coer 2 Mojca Krzan 3 Suzana Vidic 1 Darja Pavlin 4 Gregor Sersa 1 * 1 Institute of Oncology Ljubljana, Department of Experimental Oncology, Ljubljana, Slovenia 2 University of Primorska, College of Health Care Isola, Izola, Slovenia 3 University of Ljubljana, Faculty of Medicine, Ljubljana, Slovenia 4 University of Ljubljana, Veterinary Faculty, Ljubljana, Slovenia *Correspondence to: Gregor Sersa, Department of Experimental Oncology, Institute of Oncology Ljubljana, Zaloska cesta 2, SI-1000 Ljubljana, Slovenia. E-mail: gsersa@onko-i.si Received: 12 June 2009 Revised: 18 August 2009 Accepted: 24 August 2009 Abstract Background The present study aimed to evaluate the antitumor effective- ness of systemic interleukin (IL)-12 gene therapy in murine sarcoma models, and to evaluate its interaction with the irradiation of tumors and metastases. To avoid toxic side-effects of IL-12 gene therapy, the objective was to achieve the controlled release of IL-12 after intramuscular gene electrotransfer. Methods Gene electrotransfer of the plasmid pORF-mIL12 was performed into the tibialis cranialis in A/J and C57BL/6 mice. Systemic release of the IL-12 was monitored in the serum of mice after carrying out two sets of intramuscular IL-12 gene electrotransfer of two different doses of plasmid DNA. The antitumor effectiveness of IL-12 gene electrotransfer alone or in combination with local tumor or lung irradiation with X-rays, was evaluated on subcutaneous SA-1 and LPB tumors, as well as on lung metastases. Results A synergistic antitumor effect of intramuscular gene electrotransfer combined with local tumor irradiation was observed as a result of the systemic distribution of IL-12. The gene electrotransfer resulted in up to 28% of complete responses of tumors. In combination with local tumor irradiation, the curability was increased by up to 100%. The same effect was observed for lung metastases, where a potentiating factor of 1.3-fold was determined. The amount of circulating IL-12 was controlled by the number of repeats of gene electrotransfer and by the amount of the injected plasmid. Conclusions The present study demonstrates the feasibility of treatment by IL-12 gene electrotransfer combined with local tumor or lung metastases irradiation on sarcoma tumors for translation into the clinical setting. Copyright  2009 John Wiley & Sons, Ltd. Keywords gene electrotransfer; interleukin-12; irradiation; mice; sarcoma Introduction Gene therapy is receiving much attention as a local and systemic treatment for a variety of cancers. However, because of its limited impact on bulky local disease, its clinical applicability is foreseen in combination with other established cytotoxic treatment modalities (i.e. radiotherapy). The radiosensitizing role of different cytokines has already been examined, in particular, interleukin (IL)-12 has proven to be promising [1]. The discovery of IL-12 and its potent immunostimulatory effect on natu- ral killer (NK) cells of the innate immune system, as well as on cytotoxic Copyright  2009 John Wiley & Sons, Ltd.