Meet The Technology: CRISPR/Cas9

Posted on at October 6, 2017
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Dr. Xiao-Jiang Li, Distinguished Professor of Human Genetics at the Emory University School of Medicine, discusses the use of the gene-editing technique CRISPR/Cas9 to remove from the genome the mutation that causes Huntington’s disease. Li’s own research CRISPR/Cas9 shows great promise in mouse models of the disease.

Dr. Xiao-Jiang Li

Distinguished Professor of Human Genetics,
Emory University School of Medicine

HD INSIGHTS: Xiao-Jiang Li and his colleagues wrote an outstanding recent brief report for The Journal of Clinical Investigation, titled “CRISPR/Cas9 Mediated Gene Editing Ameliorates Neurotoxicity in Mouse Model of Huntington’s Disease.” Professor Li, lots of people are interested in the gene-editing technique CRISPR/Cas9. Can you briefly describe this technique for our readers?

LI: This is a new technology, which can basically use a tool to edit genomes and to eliminate mutations in genes. We know that a CAG repeat expansion is the cause of Huntington’s disease. So, you can design the CRISPR/Cas9 specifically to target the expanding CAG repeat and remove it from the endogenous gene. The Huntington mouse model carries this expanding repeat in its genome, so you use the CRISPR/Cas9 to make a correction of the mutation in the mouse genome. You can eliminate some kinds of phenotypes associated with the CAG repeat.

HD INSIGHTS: You introduced the human mutant huntingtin gene into mice and then used an adenovirus to deliver CRISPR/Cas9 to see if that could inactivate the mutant huntingtin gene in adult mice. Is that correct?

LI: Yes, because the mouse model was generated by inserting human CAG repeat into the mouse gene, the mutant protein is expressed at the endogenous level, but it carries the much larger repeat, just like in a human patient. To correct the gene mutation in the neuronal cells in the brain, we have to use an adenoviral vector to deliver the CRISPR/Cas9.

HD INSIGHTS: How do you deliver the adenovirus into the mouse?

LI: You have to do a stereotactic brain injection. You can specifically deliver the adenoviral vector into the striatum because we know the striatum is the most affected area in the brain in Huntington patients. And in that particular Huntington mouse model, people had already found that the expression of mutant protein in the striatum can cause mice to acquire a deficit in motor function. So once we do the injection we can look at the motor function to see if there is any improvement of phenotype.

HD INSIGHTS: What did you find?

LI: We found the improvement of motor function. This is a very good sign to indicate that removing mutant protein expression is beneficial for phenotype improvement.

HD INSIGHTS: You rightly point out in your paper that there’s concern that this non-allele-specific approach will also decrease normal wild-type expression of huntingtin. Did you find any side effects in your study?

LI: Early studies showed that if you deplete huntingtin expression in mice, the mice cannot develop and will die at the embryological stage. So, Huntington is very critical for neuronal survive. Our study has found that huntingtin is important for early development of central nervous system, but when the animals become adults, you can just delete huntingtin. You don’t see the obvious neurodegeneration. We found that when you inactivate normal or wild-type alleles, you don’t see obvious neurodegeneration. We don’t see any side effects, at least for a few months. We can use CRISPR/Cas9 to inactivate huntingtin expression and achieve therapeutical benefit.

HD INSIGHTS: Given the promising results of your study, what are the next steps?

LI: The immediate issue we have to address is in the long-term effect of CRISPR/Cas9 and the biosafety of this technology. Studies using the Huntington animal model just look at the animals for several months. I think we need to do long-term observation to see if there are any side effects of removing huntingtin expression in adult brain tissue. We also need to identify an efficient way to deliver CRISPR/Cas9 into the brain tissues, because that’s a challenge.

Also, we have to use viral vectors, and people are concerned about the toxicity of viral vector expression. So the challenge is, can we find another safe way to deliver non-viral CRISPR/Cas9 into the brain tissue?

In addition, the human brain is much larger than the mouse brain. For mice, we only inject the brain with a small amount of viral vector. In terms of human brains, maybe we’ll have to do more injections, but will that bring some kind of issue of toxicity of viral vectors? These are the kinds of biosafety issues we will have to address in the future.

Finally, we should test on non-human primates before we test patients, clinically.

HD INSIGHTS: As you know, clinical trials are underway for gene silencing techniques using antisense oligonucleotides and RNA silencing techniques. Where does CRISPR/Cas9 genetic therapy stand in relationship to those interventions?

LI: They’re all different approaches, but the CRISP/Cas9 has certain advantages. It can permanently remove or correct the mutation in the genome. You don’t have to continuously inhibit the expression of a mutant gene. Other methods would probably require continuously administration of drugs or other chemicals.

HD INSIGHTS: Can you predict or guess as to when we might expect to see CRISPR/Cas9 gene editing therapies in clinical trials in humans?

LI: I don’t know how many years it will be. I think there is great promise to apply this technology to treat the genetic disorders in humans. The field is moving so fast. I think we will see more and more publications using this new technology to treat different type of diseases.

HD INSIGHTS: Professor Li, when you’re not investigating gene-editing techniques aimed at Huntington’s disease, how do you spend your time outside the lab?

LI: I spend most of my time in my lab or work. I work on different projects. Huntington’s disease is just one. I also work in China on trying to use the CRISPR/Cas9 and to generate some non-human primate models.

HD INSIGHTS: Professor Li, congratulations on the great finding and many thanks for your contributions to the field.

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