ABSTRACT
Interstitial transport is a crucial step in plasmid DNA-based gene therapy. However, interstitial diffusion of large nucleic acids is prohibitively slow. Therefore, we proposed to facilitate interstitial transport of DNA via pulsed electric fields. To test the feasibility of this approach to gene delivery, we developed an ex vivo technique to quantify the magnitude of DNA movement due to pulsed electric fields in two tumor tissues: B16.F10 (a mouse melanoma) and 4T1 (a mouse mammary carcinoma). When the pulse duration and strength were 50 ms and 233 V/cm, respectively, we found that the average plasmid DNA movements per 10 pulses were 1.47 microm and 0.35 microm in B16.F10 and 4T1 tumors, respectively. The average plasmid DNA movements could be approximately tripled, ie to reach 3.69 microm and 1.01 microm, respectively, when the pulse strength was increased to 465 V/cm. The plasmid DNA mobility was correlated with the tumor collagen content, which was approximately eight times greater in 4T1 than in B16.F10 tumors. These data suggest that electric field can be a powerful driving force for improving interstitial transport of DNA during gene delivery.
Subject(s)
Electroporation/methods , Gene Transfer Techniques , Mammary Neoplasms, Experimental/therapy , Melanoma, Experimental/therapy , Plasmids/pharmacokinetics , Animals , Collagen/analysis , Extracellular Space/metabolism , Feasibility Studies , Female , Genetic Therapy , Genetic Vectors , Mammary Neoplasms, Experimental/chemistry , Mammary Neoplasms, Experimental/metabolism , Melanoma, Experimental/chemistry , Melanoma, Experimental/metabolism , Mice , Mice, Inbred BALB C , Mice, Inbred C57BLABSTRACT
Plasmids may have unique advantages as a gene delivery system. However, a major obstacle is the low in vivo transduction efficiency. In this study, an electroporation-based gene transduction approach was taken to study the effect of interleukin (IL)-2 or IL-12 gene transduction on the growth of experimental murine tumors. Significant intratumoral gene transduction was achieved by electroporation of tumors that had been injected with naked plasmids encoding reporter genes and cytokine genes (IL-2 and IL-12) under the control of a constitutive cytomegalovirus promoter. In addition, significant tumor growth delay could be achieved in a murine melanoma line B16.F10 with the cytokine genes. Most importantly, systemic transgene levels were negligible when compared with intratumoral adenovirus-mediated IL-12 gene delivery, which leads to significantly higher systemic cytokine levels. Therefore, naked plasmid- and in vivo electroporation-mediated cancer gene therapy may be therapeutically efficacious while maintaining low systemic toxicity.
