Current Progress in Electrotransfection as a Nonviral Method for Gene Delivery

Abstract

Electrotransfection (ET) is a nonviral method for delivery of various types of molecules into cells both in vitro and in vivo. Close to 90 clinical trials that involve the use of ET have been performed, and approximately half of them are related to cancer treatment. Particularly, ET is an attractive technique for cancer immunogene therapy because treatment of cells with electric pulses alone can induce immune responses to solid tumors, and the responses can be further enhanced by ET of plasmid DNA (pDNA) encoding therapeutic genes. Compared to other gene delivery methods, ET has several unique advantages. It is relatively inexpensive, flexible, and safe in clinical applications, and introduces only naked pDNA into cells without the use of additional chemicals or viruses. However, the efficiency of ET is still low, partly because biological mechanisms of ET in cells remain elusive. In previous studies, it was believed that pDNA entered the cells through transient pores created by electric pulses. As a result, the technique is commonly referred to as electroporation. However, recent discoveries have suggested that endocytosis plays an important role in cellular uptake and intracellular transport of electrotransfected pDNA. This review will discuss current progresses in the study of biological mechanisms underlying ET and future directions of research in this area. Understanding the mechanisms of pDNA transport in cells is critical for the development of new strategies for improving the efficiency of gene delivery in tumors.

Keywords: electrogene transfer; electroporation; electrotransfection; gene electroinjection; nonviral gene delivery.

Figure 1.

Physiological barriers to transport electrotransfected pDNA into the nucleus. The main barriers are the plasma membrane, cytoplasmic structures, and nuclear envelope. NPC: nuclear pore complex.

Figure 2.

Schematic of pore theory. Transient pores are induced in the plasma membrane by a pulsed electric field (from left to right). Extracellular pDNA enters the cell, denoted by the red circle, through the pores from the side facing the cathode. The pores are resealed after the pulse application. The symbols “+” and “−” represent cations and anions, respectively.

Figure 3.

Effects of DNA degradation on intracellular gene delivery. Polymer forms a complex with pDNA (polyplex) that protects pDNA from degradation by endonucleases. Naked pDNA, however, is vulnerable to degradation by nucleases in the cytosol.

Figure 4.

Proposed mechanism of ET. When the cell is exposed to a pulsed electric field, electrophoretic force will push pDNA toward the cell surface and form a complex with the plasma membrane. Then, pDNA in the complex will be internalized via endocytic pathways and move toward the perinuclear region via vesicular trafficking. Thereafter, pDNA will escape from endosomes to enter the cytosol and eventually enter the nucleus for transgene expression.

Figure 5.

Schematic of possible pathways for intracellular trafficking of naked pDNA introduced into the cell via ET. During ET, extracellular pDNA is internalized via endocytic pathways. The internalized pDNA may travel from early endosomes to lysosomes, recycle back to the cell surface, or escape from endosomes. In the cytosol, the naked pDNA may enter the nucleus for transgene expression.