Changing the Very Foundation of Everything Around Us: CRISPR-Cas9 Gene Editing

To accompany my last blog about gene editing, I want to give you all a bit of background on one of the up and coming methods of genome editing.

CRISPR-Cas9 gene editing was discovered in 2012 by Jennifer Doudna, Emmanuelle Charpentier, and colleagues, and was refined by Feng Zhang and colleagues. This method of gene editing includes three steps: recognition, cleavage, and repair. First, the sgRNA (single guide ribonucleic acid) recognizes a target sequence in the gene of interest using complementary base pairs. Then, the Cas-9 nuclease breaks the gene, cleaving it from the rest of the DNA (deoxyribonucleic acid). Finally, the double-stranded break is repaired by joining non-homologous ends or by performing homology-directed repair cellular mechanisms. 

In the early stages of the discovery of genome editing, CRISPR (Clustered Regularly Interspaced Palindromic Repeats) was found to be a key element in the adaptive immune system of prokaryotes, the bacteria in our body, that protects them from repeated viral attacks. In fact, the first CRISPR was cloned from E. coli accidentally in 1987. The Cas gene, which codes for nuclease protein, is normally found adjacent to CRISPR so it works well with it to edit genes as its original function is to help cleave viral nucleic acid. Since its discovery, the scope of this method has been expanded from a strictly-bacterial impact to its use on eukaryotes and multicellular organisms.

This method is carried out by delivering Cas enzymes and gRNAs directly into cells by means of viral and non-viral vectors. Viral vectors use the shell of a virus, called the capsid, to deliver a desired copy of a gene into the cell. Non-viral vectors are Naked DNA that is delivered by direct administration into a cell. Some of the most common viruses used by viral vectors for gene therapy are adenovirus, adeno-associated virus, and lentivirus. During administration, these viruses are inactive to prevent infection but despite this, the immune system may react to the virus and cause inflammation. Many non-viral vectors used in gene therapy fall under the larger category of cationic polymers because the negative charge on plasmid DNA becomes neutralized in the presence of these polymers which results in the formation of polymer-DNA complexes (polyplexes).  

 As of late, there are many new human CRISPR-Cas9 clinical trials that aim to edit the genes that cause certain genetic diseases such as diabetes, immune disorders, blood disorders, cardiovascular disorders, etc. As this technology is improved and the failures/side effects of some methods of CRISPR-Cas9 administration are solved, hopefully we will be able to cure many genetic disorders and possibly eradicate them entirely. 

Note: Sorry it has been so long since my last post, I was in a bit of a mental rut and was struggling to find time to write about what I had been reading.

Bibliography:

Asmamaw, Misganaw, and Belay Zawdie. “Mechanism and Applications of CRISPR/Cas-9-Mediated Genome Editing.” Biologics : targets & therapy vol. 15 353-361. 21 Aug. 2021, doi:10.2147/BTT.S326422

Barua, Sutapa et al. “Discovery of cationic polymers for non-viral gene delivery using combinatorial approaches.” Combinatorial chemistry & high throughput screening vol. 14,10 (2011): 908-24. doi:10.2174/138620711797537076

Xu, Xiaojie et al. “Delivery of CRISPR/Cas9 for therapeutic genome editing.” The journal of gene medicine vol. 21,7 (2019): e3107. doi:10.1002/jgm.3107

Zhang, Dangquan et al. “CRISPR/Cas: A powerful tool for gene function study and crop improvement.” Journal of advanced research vol. 29 207-221. 21 Oct. 2020, doi:10.1016/j.jare.2020.10.003