Potential of Prime Editing over CRISPR in Healthcare



Everyone up to date on biotechnological advances has heard of the CRISPR system for editing genomes. But it is also true that there is a lot of fuss about this technology, and not for nothing. The original function of this bacterial system is one of the most ingenious ways for simple organisms to acquire immunity to repeated contact with bacterial viruses.


Clustered regularly interspaced short palindromic repeats (CRISPR) are families of DNA fragments separated by recognizable spacers that bacteria hydrolyze from viral genomes and store in their own genome. They thus form a kind of "necklace" containing all the isoforms of viral regions with which the bacteria have come into contact. The beads of this "necklace" will serve as a mold in a new viral infection when there is a new exposure to the viral genome, and a group of endonucleases called Cas will be the enzymes in charge of inactivating this pathogenic sequence, in what is known as the crRNA complex. There are three main types of CRISPR response, with more or less significant differences in the process. The most used for genome editing is type II because Cas9 multifunctional protein is needed here, which grants at least some efficiency and specificity.


The fundamental problem of DNA editing is to repair the double strand break without losing information and preserving specificity. A design of CRISPR system could solve it in several ways. This discovery of the bacterial immune system was quickly recognized as the system with the greatest potential in gene editing, both for its flexibility and precision compared to its competition (other systems with endonucleases, such as zinc fingers). However, some studies claimed the occurrence of spontaneous mutations using this genomic editing platform. Although other studies have doubted the veracity of these accusations, the fact is that empirically there were off-target changes that made in vivo editing unfeasible at an early stage.


Undoubtedly, the number of citations and start-ups that have been born in the shadow of this innovation of tremendous potential has been exponential in this last decade, year after year. Now, there are so many CRISPR systems that it is difficult to refer to just one. The most promising one, however, is the so-called Prime Editing. It consists of three major elements: a protein domain with a modified Cas9 to cut a single strand (nickase action) and a reverse transcriptase bound to Cas9, a single guide RNA (sgRNA), and an RNA fragment called Prime Editing Guide. This pegRNA has two primary functions: to increase specificity with the genomic target and also serves as a template for the reverse transcriptase.


Once the reverse transcriptase action has finished replacing the strand, other endonucleases degrade the original unbound fragment. Now, the modification is only on one of the strands, so a guide RNA must be used again for the Cas9 enzyme to cut on the "healthy" strand. This is sufficient since the cellular mechanisms themselves will repair the DNA strand using the modified strand that we had introduced in the first place as a template.


As we have seen, the fundamental problem of in vivo gene editing can be solved with Prime Editing. However, the efficiency of this system is still likely to improve in the coming years, making this technology more and more accessible with fewer off-targets. Some limitations of this technology are the size of the inserts and the number of cells to be modified. That said, the applications are almost limitless, not only for Prime Editing but for the compendium of variations of the present and future CRISPR system. It is estimated that 90% of genetic diseases could be eradicated with current knowledge alone. Moreover, they are not only restricted to direct solutions; there are also indirect ones, by editing the genome of pathogens.


One start-up clearly focused on this strategy is Locus Biosciences, which uses the CRISPR-Cas3 system to provide a pathogen-specific bactericidal solution in a complex microbiome. Bioinformatics and machine learning tools are used to design the viral platforms where these CRISPR genes are implanted.


Another very interesting application in health is not palliation or cure, but diagnosis using CRISPR. This is what they are doing at Caspr Biotech with a very well thought-out multidisciplinary approach. Affinity is its greatest strength, as it can detect any RNA or DNA sequence. It is also fast, reliable, and inexpensive.


The Future


Whenever I am asked why it is necessary to edit the genome of organisms, I answer patiently and try to reason with the opposing positions. If we weigh the pros and cons, we will understand that the benefit for humanity is magnificent, and on the other hand, the harm is 100% avoidable if these modifications are carried out with the proper safety protocols. In this context, CRISPR has yet to improve in large-scale genome editing. Together with de novo synthesis of genomes, these are the two approaches with the greatest potential to solve the world's problems. Because not only health but also pollution, the greenhouse effect, and biodiversity loss depend directly on how genomic techniques advance and how they are applied. Of course, CRISPR remains the reigning tool in vivo editing.


Ethical barriers are important in a society, but they tend to blur over time. Many companies are tapping into this niche eagerly. It is in our power to train and demand responsible measures from our leaders to encourage these proposals.



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