Editing Small interfering RNA (section)

== Intracellular delivery ==
{{main|Intracellular delivery}}
There is great potential for [[RNA interference]] (RNAi) to be used therapeutically to reversibly silence any gene. For RNAi to realize its therapeutic potential, small interfering RNA (siRNA) must be delivered to the site of action in the cells of target tissues. But finding safe and efficient delivery mechanisms is a major obstacle to achieving the full potential of siRNA-based therapies.  Unmodified siRNA is unstable in the bloodstream, has the potential to cause [[immunogenicity]], and has difficulty readily navigating cell membranes.<ref name="Delivery materials for siRNA therap">{{cite journal |last1=Rosemary |first1=Kanasty |title=Delivery materials for siRNA therapeutics |journal=Nat Mater |year=2013 |volume=12 |issue=11 |pages=967–977 |doi=10.1038/nmat3765 |pmid=24150415|bibcode=2013NatMa..12..967K }}</ref> As a result, chemical alterations and/or delivery tools are needed to safely transfer siRNA to its site of action.<ref name="Delivery materials for siRNA therap"/>
There are three main techniques of delivery for siRNA that differ on efficiency and toxicity.

=== Transfection ===
In this technique siRNA first must be designed against the target gene. Once the siRNA is configured against the gene it has to be effectively delivered through a transfection protocol. Delivery is usually done by [[cationic liposome]]s, polymer nanoparticles, and lipid conjugation.<ref>{{cite web|title=Transfection: ''In Vitro'' Transfection|first1=Alex|last1=Fanelli| name-list-style = vanc |url=http://transfection.ws/|date=2016|access-date=5 December 2017}}</ref> This method is advantageous because it can deliver siRNA to most types of cells, has high efficiency and reproducibility, and is offered commercially. The most common commercial reagents for [[transfection]] of siRNA are [[Lipofectamine]] and Neon Transfection. However, it is not compatible with all cell types and has low in vivo efficiency.<ref>{{cite journal | vauthors = Jensen K, Anderson JA, Glass EJ | title = Comparison of small interfering RNA (siRNA) delivery into bovine monocyte-derived macrophages by transfection and electroporation | journal = Veterinary Immunology and Immunopathology | volume = 158 | issue = 3–4 | pages = 224–32 | date = April 2014 | pmid = 24598124 | pmc = 3988888 | doi = 10.1016/j.vetimm.2014.02.002 }}</ref><ref>{{cite book|title=Textbook of Medical Biochemistry| vauthors = Chatterjea MN |location=New Delhi |publisher=Jaypee Brothers Medical Publishers|year=2012|pages=304|edition=8th}}</ref>

=== Electroporation ===
{{Main|Electroporation}}
Electrical pulses are also used to intracellularly deliver siRNA into cells. The cell membrane is made of phospholipids which makes it susceptible to an electric field. When quick but powerful electrical pulses are initiated the lipid molecules reorient themselves, while undergoing thermal phase transitions because of heating. This results in the making of hydrophilic pores and localized perturbations in the lipid bilayer cell membrane also causing a temporary loss of semipermeability. This allows for the escape of many intracellular contents, such as ions and metabolites as well as the simultaneous uptake of drugs, molecular probes, and nucleic acids. For cells that are difficult to transfect electroporation is advantageous however cell death is more probable under this technique.<ref>{{cite web|url=http://www.sabosciences.com|title=siRNA Delivery Methods into Mammalian Cells |date=2016-10-13}}</ref>

This method has been used to deliver siRNA targeting VEGF into the xenografted tumors in nude mice, which resulted in a significant suppression of tumor growth.<ref>{{cite book | vauthors = Takei Y | chapter = Electroporation-Mediated siRNA Delivery into Tumors | title = Electroporation Protocols | series = Methods in Molecular Biology | volume = 1121 | pages = 131–8 | year = 2014 | pmid = 24510818 | doi = 10.1007/978-1-4614-9632-8_11 | isbn = 978-1-4614-9631-1 }}</ref>

=== Viral-mediated delivery ===
The gene silencing effects of transfected designed siRNA are generally transient, but this difficulty can be overcome through an RNAi approach. Delivering this siRNA from DNA templates can be done through several recombinant viral vectors based on retrovirus, adeno-associated virus, [[adenovirus]], and [[lentivirus]].<ref>{{cite book | vauthors = Talwar GP, Hasnain S, Sarin SK | date = January 2016 | title = Textbook of Biochemistry, Biotechnology, Allied and Molecular Medicine | edition = 4th | publisher = PHI Learning Private Limited | page = 873 | isbn = 978-81-203-5125-7 }}</ref> The latter is the most efficient virus that stably delivers siRNA to target cells as it can transduce nondividing cells as well as directly target the nucleus.<ref name=pmid16397511>{{cite journal | vauthors = Morris KV, Rossi JJ | title = Lentiviral-mediated delivery of siRNAs for antiviral therapy | journal = Gene Therapy | volume = 13 | issue = 6 | pages = 553–8 | date = March 2006 | pmid = 16397511 | pmc = 7091755 | doi = 10.1038/sj.gt.3302688 }}</ref> These specific viral vectors have been synthesized to effectively facilitate siRNA that is not viable for transfection into cells. Another aspect is that in some cases synthetic viral vectors can integrate siRNA into the cell genome which allows for stable expression of siRNA and long-term gene knockdown. This technique is advantageous because it is in vivo and effective for difficult to transfect cell. However problems arise because it can trigger antiviral responses in some cell types leading to mutagenic and immunogenic effects.

This method has potential use in gene silencing of the central nervous system for the treatment of [[Huntington's disease]].<ref>{{cite book | vauthors = Cambon K, Déglon N | title = Trinucleotide Repeat Protocols | chapter = Lentiviral-Mediated Gene Transfer of siRNAs for the Treatment of Huntington's Disease | series = Methods in Molecular Biology | volume = 1010 | pages = 95–109 | year = 2013 | pmid = 23754221 | doi = 10.1007/978-1-62703-411-1_7 | isbn = 978-1-62703-410-4 }}</ref>