Gene electrotransfer is a versatile biotechnology technique that enables the transfer of genetic material into prokaryotic or eukaryotic cells. It is based on a physical method named electroporation, where a transient increase in the permeability of cell membrane is achieved when submitted to short and intense electric pulses, thus enabling the transport of large molecules (naked plasmid DNA, antisense oligonucleotides, siRNA) into cells that otherwise cannot permeate through the cell membrane. Gene electrotransfer was first described in the 1980s and since then due to its ease of application and efficiency become a routine method for introducing foreign genes into bacterial, yeast, plant, and animal cells in vitro and into different tissues, including muscle, tumors, liver, and skin in vivo.
Contents
Physical principle
Application of electric pulses of sufficient strength to the cell causes an increase in the trans-membrane potential difference, which provokes the membrane destabilization. Cell membrane permeability is increased and otherwise nonpermeant molecules enter the cell. Although the mechanisms of gene electrotransfer are not yet fully understood, it was shown that the introduction of DNA only occurs in the part of the membrane facing the cathode and that several steps are needed for successful transfection: electrophoretic migration of DNA towards the cell, DNA insertion into the membrane, translocation across the membrane, migration of DNA towards the nucleus, transfer of DNA across the nuclear envelope and finally gene expression. There are a number of factors that can influence the efficiency of gene electrotransfer, such as: temperature, parameters of electric pulses, DNA concentration, electroporation buffer used, cell size and the ability of cells to express transfected genes. In in vivo gene electrotransfer also DNA diffusion through extracellular matrix, properties of tissue and overall tissue conductivity are crucial.
Biotechnological applications
Bacterial transformation is generally the easiest way to make large amounts of a particular protein needed for biotechnology purposes or in medicine. Since gene electrotransfer is very simple, rapid and highly effective technique it first became very convenient replacement for other transformation procedures.
Medical applications
First medical application of electroporation was used for introducing poorly permeant anticancer drugs into tumor nodules. Soon also gene electrotransfer became of special interest because of its low cost, easiness of realization and safety. Namely, viral vectors can have serious limitations in terms of immunogenicity and pathogenicity when used for DNA transfer.
In vivo gene electrotransfer was first described in 1991 and today there are many preclinical studies of gene electrotransfer. The method is used to deliver large variety of therapeutic genes for potential treatment of several diseases, such as: disorders in immune system, tumors, metabolic disorders, monogenetic diseases, cardiovascular diseases, analgesia….
Also first phase I clinical trial of gene electrotransfer in patients with metastatic melanoma was reported. Electroporation mediated delivery of a plasmid coding gene for interleukin-12 (pIL-12) was performed and safety, tolerability and therapeutic effect were monitored. Study concluded, that gene electrotransfer with pIL-12 is safe and well tolerated. In addition partial or complete response was observed also in distant non treated metastases, suggesting the systemic treatment effect. Based on these results they are already planning to move to Phase II clinical study. There are currently several ongoing clinical studies of gene electrotransfer, where safety, tolerability and effectiveness of immunization with DNA vaccine, which is administered by the electric pulses is monitored.
Although the method is not systemic, but strictly local one, it is still the most efficient non-viral strategy for gene delivery.