Disease research has a lot of safeguards built in, requirements that must be reached before proper human treatment can be started. While it is true that sometimes these conditions are far more onerous than is fair or essential, they do in a general sense help pave the road for effective human medicine. That is why we use human-like model species that can be an effective benchmark for what the same treatment will do when tested in humans, so as to make things safer for that first group of trial participants.

One of the most prevalent and useful models used in science is obviously one of those closely related to humanity in general, the macaque monkey. Since it is so similar to us in design, we can be moderately confident that any negative side effects will be expressed equally in macaques as they would in humans. Though there are often biases in the types of models that actually end up being used in general research, usually because of ease of access, a requirement of numbers, or time restrictions.

Thanks to some combination of those, gene editing science has largely stuck only with mice models. But, as more and more experiments have begun to show, mice might not be the best model for comparative human studies. They differ from us in some critical ways and so the impact of a treatment may not be applicable. Meanwhile, macaques have seen significant use when it comes to the related field of gene therapy research, but this hasn’t been replicated for gene editing.

Silencing Bad Cholesterol

Now, scientists from the University of Pennsylvania wish to change that with their experiment attempting to treat hypercholesterolemia, otherwise known as high cholesterol. Their goal is to edit and silence the gene PCSK9, responsible for the protein of the same name that antagonizes the low-density lipoprotein receptor. LDL is what is commonly referred to as “bad cholesterol”, while HDL is the preferred alternative. This is because LDL can become oxidized after exposure to free radicals or cellular debris, making it bind to other molecules and build up in the arteries leading to heart disease and other conditions.

The researchers chose to go with a more unique gene editing option known as meganucleases. These are a specially engineered form of endonucleases that naturally cut DNA before or after particular sequences. These modified versions can be set to target the sequences that the scientists want to cut or isolate in the genome. They took the endonuclease they wanted to use from the algal species Chlamydomonas reinhardtii due to the extensive prior research into its gene editing abilities.

It should be noted, however, that meganucleases remain more difficult to use than more conventional gene editing technologies like TALENs and CRISPR, which is why they aren’t as common in scientific research. They were only selected for this experiment due to past research in macaques with certain vectors like adeno-associated virus (AAV). Since it is known that AAV pairs well with meganuclease complexes.

The sequence that was chosen to be targeted in the PCSK9 gene was one that exists in both the macaque and human versions of the gene, therefore heightening the comparisons between the results. It is also a highly conserved and unique part of the sequence that is found only in primate species. The macaques were then injected with the AAV vectors containing the meganucleases, with no side effects or negative reactions noted from the infusion of the viral vector.

Extending The Effects

Serum levels of PCSK9 were observed to drop based on the level of dosage of the vector given. In the low dosage group, the protein level dropped by 3.2%, in the mid dosage group 45%, and in the high dosage group 84%. This was reflected with a significant drop in serum LDL levels for the mid and high groups of 48% and 60%, respectively. Tissue samples from the liver showed that the incorporation of the cleavage mechanism shutting down PCSK9 reached 70-80% of the cells at the high dose end and only 15-35% on the lower dosages, thereby showing that the inactivation response is directly dose dependent based on how much of the vector, and thereby the meganucleases, is applied.

These higher levels are far more than enough to deal with medical conditions like high cholesterol issues and therefore the treatment has therapeutic potential for use in humans. It should be noted that, in comparison to gene therapy that aims to have stable long-term expression of transgenes, this form of gene editing treatment works as only transient expression or silencing of targeted gene sequences. If the cells targeted are non-dividing, then the effects can be more longer-lasting. Otherwise, when it comes to cells like those in the liver, division eventually overtakes any gene alteration.

The researchers are not entirely sure why expression drops after around 4-6 months, but they suspect that the AAV vectors or the meganuclease transgene complexes are inactivated themselves by the host cells through some unknown process. Also, it should be noted that there was some eventual immune response, which was unsurprising due to the use of a Chlamydomonas enzyme that would be viewed as foreign material. The overall bodily impact was measured as minimal however, though the scientists suggested that future experiments test using immune system modulators, like glucocorticoids, during vector administration to see if that reduces the toxic immune response even further.

A Medical Opportunity

While there are certainly further improvements that are possible in the future, the amount of success achieved in this experiment provides enough safety to allow for human trials to begin sooner rather than later and the researchers suggest that such trials are done based on the individual severity of the health needs of any target patient population. Therefore, the ones who need it the most and whose lives are at risk are best suited for this treatment even without expanded improvements to it.

But those improvements as well will eventually come, allowing for expanded usage of the treatment as a medical option for those with high cholesterol issues. Yet another condition that can be managed thanks to the capabilities of modern science and the power of gene editing.

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Photo CCs: Macca mulata from Wikimedia Commons

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