It’s Prime Time for Gene Editing

Imagine yourself as a string of “A”s, “C”s, “G”s and “T”s. Now imagine pasting that in a Google Doc and being able to go anywhere in the code of your body to edit anything you want. You could cure genetic diseases, fix any deformities, do anything, yes, anything you want to your body, all by editing your genetic code. This may soon become a reality.

Prime editing is a new gene-editing technique capable of making a wide variety of edits, including insertions, deletions and DNA letter substitutions.

Biology 101:

In case youve forgotten all about your high school bio classes and had no idea what I was just talking about, here’s a quick refresher on the need-to-knows

That is deoxyribonucleic acid, more commonly known as DNA. It is the instruction manual for our cells; it stores the information you need to function and tells our cells how to produce proteins. DNA is in the structure of a double helix, which looks sort of like a twisted ladder (look up!), with 2 winding strands connected by rungs. These rungs are made of the chemical building blocks adenine (A), cytosine (C), guanine (G), and thymine (T), or the “A”s, “C”s, “G”s and “T”s, of DNA code.

A gene is a sequence of this code which contains the instructions on how to make a protein. This is only 1% of the DNA sequence. The rest determines when, how, and how much of these proteins are made.

So whats a protein? They are complex molecules that do most of the work in our cells, and thus our body. This includes the structure, operation, and maintenance of our tissues and organs.

RNA looks like DNA, but cut in half. It has many roles, including helping in the protein creation process.

Of course, our bodies make mistakes too. Gene mutations are alterations in a DNA sequence so that it differs from most people. These can range from a single letter to a large segment which can affect multiple genes. These can be either inherited from a parent, caused by DNA being copied erroneously, or by environmental factors, including radiation.

Now that you have a grasp on the basics, let’s talk about prime editing.

How does it work?

Prime editing uses a combination of 3 major components:

A prime editor complex consists of the Cas9 protein and reverse transcriptase. An engineered pegRNA sends the editor to the target, where the Cas9 snips one strand of the DNA. The editor then needs to replace the old DNA sequence with the edited sequence. This is where the reverse transcriptase comes in. It reads the desired edit from the pegRNA and rewrites the corresponding “A”s, “C”s, “G”s and “T”s to the end of the nicked DNA strand. Once the new letters are sealed into the genome, there is a mismatch between the original sequence on one strand and the edited sequence on the other strand of DNA. To resolve this, a different guide RNA sends the editor to the unedited strand to make a cut, which prompts the cell to remake the nicked strand, by copying the edited strand and completing the edit.

Confused?

Don’t worry, I was too the first time.

Just think of the DNA double helix as a ladder, with strands/rails AA. Prime editing cuts off one of the strands, or the side rail of the ladder, using the Cas9, allowing for the edited sequence, or a new rail on the side of the ladder, to be inserted. Now, the strands/rails are AB, with A being the original sequence and B being the edited sequence. The editor then snips the other, unedited strand/rail. The cell, detecting DNA damage, then fixes that strand/rail to match the other, edited strand/rail, so that they now match with strands/rails BB.

Wait, but don’t we already have CRISPR?

Yes, we do, but CRISPR, unlike prime editing, breaks both strands of DNA, or both rails of the ladder, and relies on the cell’s repair system to fix the damage and make the necessary edits. Because CRISPR does not have any control over the cell’s repair system, this can result in the insertion or deletion of DNA letters where the genome was cut, which may have unwanted — and sometimes dangerous — consequences, including activating cancer-causing genes. Prime editing takes the cell’s repair system out of the equation, preventing any potential errors associated with it.

Second, it isn’t exactly versatile. It was previously thought that different CRISPR tools would need to be developed for a specific type of edit: to delete a gene, insert new DNA code, or perform DNA letter substitutions (changing a letter). Prime editing can do all 3.

Finally, CRISPR is not capable of the single letter type, precise substitution edits that prime editing can do. For that, a different technique, called base editing, is turned to. Base editing changes individual letters of DNA into another through performing chemical reactions. The problem with base editing is that it can’t help with multi-letter mutations such as Tay-Sachs disease, which is an often fatal disease caused by 4 extra DNA letters in the HEXA gene. Thus, it is very limited to the problems it can solve.

The 4 letter mutation in the HEXA gene, causing Tay-Sachs disease, which affects approximately 1 in 250–300 people. This mutation cannot be corrected using base editing, however, prime editing is capable in removing the 4 extra letters.

Think of it this way: imagine the target DNA as a misspelt word. CRISPR works by deleting that word and relying on autocorrect to fix it. We all know that autocorrect can surprise you in how wrong it can be. Meanwhile, base editing can change a single letter in that misspelt word. Sure, it would work great if there was only 1 wrong letter. But what if there were 2? 3? Extra letters? A missing letter? In these cases, base editing would be useless. Prime editing allows for more control over the edit than CRISPR while maintaining the precision that base editing has. It’s the equivalent of deleting that misspelt word, and then copy-pasting the correct spelling from elsewhere.

That’s cool and all, but why should we care?

Well, for one, it could potentially correct up to 89% of all known disease-causing genetic variants, affecting millions of people worldwide. This includes people suffering from sickle cell anemia, cystic fibrosis, and Tay-Sachs disease. According to the WHO, approximately 1/100 people are diagnosed at birth with a monogenic disease, or diseases resulting from modifications in a single gene in all cells throughout the body.

Because prime editing is so versatile in the types of edits it can do, it is capable of correcting a wider variety of genetic disorders than current gene-editing techniques. Diseases previously thought incurable wouldn’t be with prime editing. Fewer parents would have to dread to see if they had passed their genetic disorder on to their children and many more people would be given the chance to live long, normal, healthy lives. Genetic diseases will be a thing of the past.

It could also be applied in agriculture. Although existing gene-editing techniques, such as CRISPR, are capable of this already, it is extremely inefficient. Prime editing boasts versatility, enabling it to perform a large variety of edits, which makes it appealing for the development of superior crops. This includes increasing yield, improving resistance to environmental stresses (ie. drought, disease and heat), and improving the quality of the crop, among many others. This can contribute significantly to food security and sustainable development, benefitting countless communities where food shortages are prevalent. Read more about it here:

https://spj.sciencemag.org/bdr/2020/9350905/

Well if it’s so awesome, why haven’t I heard about it?

At this point, the technique is still very new to the game, showing up in October of 2019. As such, it will still have to go through rigorous testing before it can be accepted. However, initial experiments with prime editing seem promising. Using prime editing, less than one-tenth of edited cells had unwanted changes, compared to up to 90% for first-gen CRISPR systems.

Being a new technology, there is still work to be done before scientists can even think about using it on humans. Researchers have to figure out how to deliver the tool into the body and to the cells safely and effectively. Viral and nonviral methods have been studied, and animal testing is planned to happen.

Furthermore, there is the possibility that the introduction of a reverse transcriptase could result in the coping of other RNA in the cell to insert into the genome, resulting in possible, unpredictable consequences.

Experiments and studies will continue to arise to assess how well the system functions in different cells and organisms. However, if all goes well, we may very well expect to see prime editing be the way that disease-causing genetic mutations are repaired. It will be a major step towards a day where humans can make any DNA change at any position of any organism.

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