Bleomycin electrotransfer enhancement by using Plasmid Dna

Abstract

Over the last decade a new cancer treatment method termed electrochemotherapy has emerged. Such anticancer therapy is based on the process of electroporation. This process initiates when applied electric field is increasing transmembrane voltage of the affected cells, hence increasing cell membrane permeability that leads to increased uptake of exogenous molecules such as anticancer drugs. Currently, the combination of anticancer drug (mainly bleomycin) and the method of electroporation is used in clinics. Once inside the cell, the anticancer drug bleomycin induces the cellular DNA cleavage, that in turn leads to cell death. Previously it has been shown that big molecules (dextrans) with high net charge increase electrotransfer of bleomycin into the cells. Here we show the enhanced bleomycin electrotransfer when plasmid DNA (as a big molecule with charge) was in the media during the electroporation We analysed bleomycin electrotransfer in Chinese Hamster Ovary (CHO) cells. For that different concentrations of anticancer drugs were used together with pDNA transfection. pMAX FGP coding plasmid in concentraton of 200 µg/ml was used. The anticancer drug bleomycin was used for electrotransfer experiments in the concentrations ranging from 0.1 to 20 ng/ml. Electroporation was performed by using combination of 1 electric pulse that induced electric fields at the amplitude of 1400 V/cm for the duration of 100 µs. Afterwards, comet assay was performed to evaluate DNA damage. In addition, clonogenic assay was done to evaluate cell viability. Obtained results showed bigger bleomycin electrotransfer after usage of combination of bleomycin, pDNA and electroporation. In conditions of 1HV electroporation intensity, 200 µg/ml pDNA and 20 ng/ml BLM concentrations, DNA cleavage reaches up to 10% [...]Aplinkos tyrimų centrasGamtos mokslų fakultetasVytauto Didžiojo universiteta