Dependence of bleomycin cytotoxicity on plasmid size using concomitant bleomycin and plasmid DNA electrotransfer

Electroporation is a process, when applied short and intense electric pulses that transiently permeabilize the cell plasma membrane, thus allowing transport of molecules otherwise non-permeant through the membrane. The method has been applied for anticancer treatment, as anticancer drugs, can be easier transported into electroporated tumour cells. Currently, the method is used in clinics and termed as electrochemotherapy (ECT). At this moment, various methods are being explored to increase the impact of ETC, such as gene transfer. Preliminary studies have shown that plasmid DNA and bleomycin (BLM) in the medium allows simultaneous transport of both molecules. Also, we have shown that the presence of DNA in the medium can increase bleomycin transport and thereby cytotoxicity These effects can be used to increase the efficiency of ECT and to regulate the immune response, by delivering genes, encoding specific cytokines. Chinese Hamster Ovary cells were used for bleomycin and plasmid DNA electrotransfer experiments. BLM was used at the concentrations ranging from 0.1 to 10 nM. pEGFP (4.7 kb) and piggyBac (7.1 kb) plasmids in concentrations of 200 μg/ml were used. Cells were suspended in electroporation medium (conductivity 0.1 S/m, osmolarity 270 mOsm, pH 7.1). Cell electroporation was performed by using combination of 1 electric pulse of 1400 V/cm pulse strength and 100 μs pulse duration. Our study has shown that a combination of plasmid DNA, bleomycin, and electroporation increases the cytotoxic effect of the anticancer drug. Furthermore, the results have shown that the cytotoxic effects of anticancer drug (BLM) are dependent on plasmid DNA sizeBiologijos katedraGamtos mokslų fakultetasVytauto Didžiojo universiteta