Effective gene transfer to solid tumors using different nonviral gene delivery techniques: Electroporation, liposomes, and integrin-targeted vector

Abstract

In this study, we measured transfection efficiency in vitro and in vivo using the following nonviral approaches of gene delivery: injection of plasmid DNA, electroporation -assisted, liposome -enhanced, and integrin -targeted gene delivery, as well as the combination of these methods. Four histologically different tumor models were transfected with a plasmid encoding the green fluorescent protein ( GFP ) ( B16 mouse melanoma, P22 rat carcinosarcoma, SaF mouse sarcoma, and T24 human bladder carcinoma ) using adherent cells, dense cell suspensions, and solid tumors. Emphasis was placed on different electroporation conditions to optimise the duration and amplitude of the electric pulses, as well as on different DNA concentrations for effective gene delivery. In addition, transfection efficiency was correlated with cell density of the tumors. The major in vivo findings were: ( a ) electroporationassisted gene delivery with plasmid DNA, employing long electric pulses with low amplitude, yielded significantly better GFP expression than short electric pulses with high amplitude; ( b ) electroporation combined with liposome -DNA complexes yielded the highest percentage of transfected tumor area in B16F1 tumor ( 6% ); ( c ) transfection efficiency of electroporation -assisted plasmid DNA delivery was dependent on tumor type; ( d ) integrin -targeted vector, alone or combined with electroporation, was largely ineffective. In conclusion, our results demonstrate that some nonviral methods of gene delivery are feasible and efficient in transfecting solid tumors. Therefore, this makes nonviral methods attractive for further development.

In vivo characterization of horseradish peroxidase with indole-3-acetic acid and 5-bromoindole-3-acetic acid for gene therapy of cancer

“…12 Assessment of vascular density Frozen tumor sections were stained for CD34 to detect blood vessels following manufacturer's instructions (rat anti-mouse CD34, 1:200, MCA1825, Serotec, Kidlington, UK). Vessel density was quantified using a Chalkley eyepiece graticule (Graticules Ltd, Tonbridge, Kent, UK) essentially as described earlier.…”

MicroRNAs targeting mutant K-ras by electrotransfer inhibit human colorectal adenocarcinoma cell growth in vitro and in vivo

“…In electrotransfection protocol, electric pulses were applied 10 min after intratumoral injection of plasmid DNA. 27 Assessment of in vivo electrogene therapy using miRNA-K-ras Electrogene therapy of LoVo tumors with pmiRNA-K-ras was assessed by comparing the effects of electrotransfection with pmiRNA-K-ras to other treatment procedures. Quantitative RT-PCR: on day 6 after treatment, two tumors per group were excised and frozen in liquid nitrogen.…”

Control by pulse parameters of DNA electrotransfer into solid tumors in mice

“…[6][7][8] However, most studies were carried out in muscle, where high transfection efficiencies can be obtained. So far, the transfection efficiencies obtained in tumors following electropulsation have been smaller compared to other tissues, [10][11][12] but a significant therapeutic effect can nevertheless be obtained by electrotransfer of therapeutic genes into tumors. 7,8 Classically, electrically mediated gene delivery is obtained by submitting the target to a train of pulses lasting a few milliseconds or 100 microseconds at a 1 Hz frequency, and adjusting the voltage to the electrode width.…”

“…7,8 Classically, electrically mediated gene delivery is obtained by submitting the target to a train of pulses lasting a few milliseconds or 100 microseconds at a 1 Hz frequency, and adjusting the voltage to the electrode width. [10][11][12][13][14][15][16] A new protocol with high transfection efficacy in muscle and skin has recently been described, consisting of a combination of one high-field (HV, B1000 V cm À1 ) and one (or several) low-field (LV, B100 V cm À1 ) pulse. [17][18][19][20][21][22][23] For simplicity, we called all along the paper 'field' what indeed is the voltage to electrode width ratio, that is, an experimentally controlled parameter.…”