Today it is already used as a programmable tool to cleave practically any double-stranded DNA sequence with precision that eases the introduction of chosen DNA sequences (or genes) into a target genome via "donor" DNA. Parallel to extensive amount of information derived from mapping various genomes, this tool opens the door to editing specific areas in cell’s genome.
In basic biology, the method is used to obtain insights into cell behavior, as well as to engineer specific cell lines and model organisms. In biomedicine it has a potential to result in new therapies for human diseases, but also to make revolutionary and controversial changes in the genome of practically any living cell – including human embryos and stem cells.
Improved delivery systems and the use of nanocarriers significantly improved the efficacy and specificity of CRISPR/Cas technology. There is a dramatic increase in the number of studies and publications that employed this technique to edit the genome of human somatic cells and induced pluripotent stem cells. Furthermore, there are companies that already offer commercial kits and services to assist researchers from various scientific fields that are interested in this method.
CRISPR/Cas9 system can be employed to create exceptionally customizable genetic adjustments such as chromosomal alterations. It represents an expeditious and comprehensible model for the initiation and progression of various tumors, as well as for drug sensitivity and resistance predictions in cancer.
Such meticulous genome editing features enable easier and faster modeling of malignant diseases, which holds a promise of accelerating translational research. One possibility this technology offers is the formation of personalized platforms, i.e. creation of specific models that will mimic distinct presentation of malignant diseases and test drug susceptibility.
Another approach is the use of CRISPR/Cas9 system in immunotherapy for immune cells reprogramming outside of a human body. Likewise, the system can be used to knock out certain genes in order to increase LDL receptor levels and decrease cholesterol levels, showing a therapeutic potential for preventing cardiovascular diseases.
In infectious disease realm, CRISPR/Cas9 can be employed to selectively exhaust certain bacterial community of an exceptionally harmful species (which will be more prominent as we continue to describe human microbiome). It also shows promise in the treatment of latent and persistent viral infections.
The inherent plasticity and modularity of the CRISPR/Cas9 technology has resulted in the rise of many genome engineering applications, the majority of which have been successfully executed in cell culture systems. A significant proportion of these are currently being adapted for their use in vivo.
Such efficiency of genome editing and ease of use have resulted in the development of CRISPR knockout libraries that cover whole genome and hold the potential to knock out every gene in the genome (with already conducted proof-of-principle experiments on both human and mouse cells).
Naturally, editing of the genome via CRISPR-based technology is not without concerns and disadvantages. Two major challenges at the moment are off-target effects (which is perhaps the biggest concern) and certain obstacles in delivery (one disadvantage is the size of the Cas9 protein).
In order minimize aforementioned off-target effects of CRISPR/Cas9 system, a modified system that employs truncated guide RNAs (abbreviated as tru-gRNAs) was developed and validated. Its usage resulted in a decrease of unwanted off-target effects, without compromising their target efficiency.
Thakore PI, Gersbach CA. Genome Engineering for Therapeutic Applications. In: Laurence J, Franklin M, editors. Translating Gene Therapy to the Clinic: Techniques and Approaches. Academic Press, Elsevier, 2015; pp. 27-44.