December 10, 2017
Targeted Genome Editing: Researchers Reversed Kidney Disease, Muscular Dystrophy In Mice

Targeted genome editing advancements could spell the end of a multitude of human diseases. A gene editing system using the functions of CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) and CRISPR-associated (Cas) genes has been widely used in the scientific community in recent years. The technology has already been used to target important genes in many animals including humans, zebra-fish, monkeys, rabbits, pigs, rats, and mice. Researchers have used this system to introduce mutations in targeted genes using gRNA. The only issue with this older CRISPR-Cas9 approach is that it actually cuts DNA molecules. The cutting apart of the double-stranded DNA molecule could cause new, unexpected mutations and uncertain side effects.

Now, researchers at the Salk Institute have made a major breakthrough with the CRISPR-Cas9 technique. They modified the gene targeting system so that it alters gene activity of disease-associated genes and does not cut the DNA. This is the first time researchers have shown that they can alter the actual phenotype of an animal with genetic editing technology and still preserve the overall integrity of the animal's DNA.

More than half of the animal models showed improved health after the new CRISPR intervention, according to the the Los Angeles Times.

The new Salk technique uses two adeno-associated viruses (AAVs) as the mechanism to introduce gene altering cells into mice. They inserted the gene for the Cas9 enzyme into one AAV virus. Then, they used another AAV virus to introduce a short single guide RNA (sgRNA) into the mice cells. The second AAV virus specified exactly where the Cas9 would bind on the mouse genome. According to Science Daily, the sgRNA has fewer nucleotides than what was used in earlier CRISPR-Cas9 techniques. The fewer nucleotides prevented the Cas9 from cutting the DNA.

"Basically, we used the modified guide RNA to bring a transcriptional activator to work together with the Cas9 and delivered that complex to the region of the genome we were interested in," co-first author Hsin-Kai Liao explained. The team claims that similar techniques could be used to activate any gene without the risk of causing unexpected genetic mutations while reversing the course of disease.

"We wanted to change the cell fate with therapeutic efficiency without a DNA cut," co-first author Fumiyuki Hatanaka said, according to Science Daily.

That's exactly what they did.

Genome editing without cutting has reversed disease in mouse models.

They managed to use this gene targeting therapy to reverse diseases in mice. They were able to use their newly modified technique to silence genes that were linked to acute kidney disease. This remarkably restored normal kidney function in the mice. Also, they caused liver cells to differentiate into cells that would produce insulin like the pancreas normally would, partially reversing type 1 diabetes in a mouse model. They even managed to recover muscle functioning in mouse models of muscular dystrophy. They didn't even try to repair the gene that was causing the disease. They just increased genetic expression that was on the same pathway as the mutated, disease-causing gene. This effect over-rode the effects of the original, damaged gene!

"We are not fixing the gene; the mutation is still there," senior author Juan Carlos Izpisua Belmonte of the Salk Institute for Biological Studies explained. "Instead, we are working on the epigenome and the mice recover the expression of other genes in the same pathway. That is enough to recover the muscle function of these mutant mice."

The researchers say that it seems as though the technique is perfectly safe and that it doesn't seem to cause undesirable, unexpected genetic mutations. They think this therapy could one day be used to treat even neurological disorders like Parkinson's disease, Alzheimer's disease, and a multitude of other diseases.