Physical Delivery
An Overview
Various physical manipulations have been used to enhance the efficiency (rate and extent) of gene delivery to the desired defective cells, especially in the in vivo conditions. These physical methods for gene transfer have also demonstrated the ability to circumvent various extra- and intracellular barriers, which significantly compromise the efficiency of gene delivery, including massive dilution of DNA upon injection, accessibility of the target site, and entry into the cell and the nucleus, thus enhancing gene expression in the desired cells for the desired duration.
Electroporation (EP)
EP has proved to be a successful method for drug delivery across the skin in vitro and in vivo and has also been used as an effective in vitro gene delivery system in prokaryotic and eukaryotic cells. It utilizes physical force to import therapeutic drugs and macromolecules, such as DNA and proteins, from extracellular compartments into cells. EP transiently increases cell membrane permeability caused by short, pulsed electric field application, temporarily disrupting the structural integrity of cell membranes. Electric pulses of different intensity and duration are applied according to the target cell, tissue or organ. This procedure has been well received for its use in vivo in the clinic due to its simplicity and low adverse effects.
Figure 1. Electroporation. (Mehta, 2015)
Hydrodynamic Intravascular Injection
The hydrodynamic gene delivery via tail vein injection is a highly efficient method to deliver nucleic acids to the liver in small animals. This way of gene delivery using a rapid injection of a relatively large volume of DNA solution has opened a new avenue for gene therapy studies in vivo. This approach combines naked DNA and hydrodynamic pressure generated by rapid injection of a large volume of fluid into a blood vessel, to deliver genetic materials into parenchyma cells. The hydrodynamic injection is widely applicable for the delivery of transgene in important organs such as the liver, kidney, skeleton muscle, heart, lung, and spleen. All these organs show marked transgene expression, but especially among the liver, which has the highest transgene expression.
Figure 2. Hydrofection. (Herrero, 2009)
Sonoporation
In terms of sonoporation, it is a non-viral gene delivery system for increasing plasmid DNA transfer across the biological cell membranes. It works by transient permeabilization of the cell membrane and by transferring therapeutic DNA effectively across tissue and into cells by application of ultrasound energy. The ultrasound technique has been routinely used clinically for both therapeutic and diagnostic purposes, and it covers a broad range of frequencies and waveforms. Similar to electric pulses, high-intensity ultrasounds can permeabilize cell membranes to deliver cargo molecules by pore formation.
Figure 3. Sonofection. (Tomizawa, 2013)
Particle Bombardment (Gene Gun)
Gene gun is a procedure in which particles bearing nucleic acids are propelled at high velocity to impact on a biological target in order to penetrate deeply and transfect cells by delivery of cargo molecules. There are different gene gun devices commercially available (Powderject, PowderMed, Iaculor Injection) that can be used successfully for most of the purposes. The most recommended material for preparing the particles employed with the gene gun procedure is gold. Gold can ionize to have a positive charge, which will interact well with DNA or RNA thanks to the negative charge on every phosphate group in the polynucleotide.
Figure 4. Gene gun.
Jet Injection
Jet injection technology has developed as an applicable alternative to viral or liposomal gene delivery systems. This is used for the direct gene transfer of naked DNA into cells. Jet injection allows gene transfer into different tissues with deeper penetration of the desired DNA into the targeted tissue at comparable transfer efficiencies achieved by particle bombardment. The jet injection has been used in a wide variety of clinical applications such as antigen immunization (vaccines), hormone delivery and local anesthesia. It has also been employed for many gene therapy strategies, like antitumor therapy via jet injection of DNA encoding suicide genes (cytosine deaminase) or genetic vaccines.
Figure 5. Jet injection. (Ishikawa, 2017)
Overview of Different Physical Methods for Gene Delivery
| Method | Advantages | Disadvantages |
| Electroporation | Simplicity; low cost; wide electrode variety; widely employed for in vitro, in vivo and clinic | Short-term pain, erythema, discomfort; tissue damage |
| Sonoporation | Noninvasive process, efficacy (lower than electroporation); combined with bubbles or nanocarriers increases the efficiency | Low precision; low reproducibility; cell damage by shear forces and increase temperature; tissue damage (but less than electroporation) |
| Jet injection | Minimal or no immune response; low toxicity | Delicate procedure; requires training; high time consuming |
| Particle Bombardment | Gene gun device | Requires particles; the type and size define the toxicity and tissue penetration; cell damage |
| Hydrodynamic injection | Single injection in small animals; Catheter-guide in large animals | Hemodynamic changes; transitory increase in liver enzymes in plasma |
References
- Mehta, S. (2015). Electroporation.
- Tomizawa, M.; et al. (2013). Sonoporation: Gene transfer using ultrasound. World J Methodol. 3(4): 39-44.
- Herrero, M.J.; Alino, S.F. (2009). Naked DNA Liver delivery by hydrodynamic injection. GeneTherRev.
- Ishikawa, K. Cardiac Gene Therapy Methods and Protocols (1st ed. 2017, Methods in Molecular Biology. 1521). New York, NY: Springer New York: Imprint: Humana.
