Researchers have reversed disease in mice using a CRISPR-Cas9 technique that alters gene activity rather than the underlying sequence. The discovery is of particular significance because it sidesteps the issue of mutations that result from inserting or removing genes.
How the new technique works
The new technique, which was developed by a team led by Juan Carlos Izpisua Belmonte of the Salk Institute for Biological Studies, has successfully used two adeno-associated viruses (AAVs) as the mechanism to introduce genetic manipulation machinery into cells in post-natal mice.
The researchers inserted the gene for the Cas9 enzyme into one AAV virus. They used another AAV virus to introduce a short single guide RNA (sgRNA), which specified the precise location in the mouse genome where Cas9 would bind, and a transcriptional activator. The shorter sgRNA was only 14 or 15 nucleotides, compared with the standard 20 nucleotides used in most CRISPR-Cas9 techniques, and this prevented Cas9 from cutting the DNA.
In essence, the modified guide RNA was used to bring a transcriptional activator to work together with the Cas9 and delivered that complex to the region of the genome the researchers were targeting.
The complex sits in the region of DNA of interest and promotes expression of a gene of interest. Similar techniques could be used to activate virtually any gene or genetic pathway without the risk of introducing potentially harmful mutations.
Disease reversed without cutting DNA
The new technique reversed acute kidney disease and restored normal kidney function in a mouse model. It also induced some liver cells to differentiate into pancreatic-like cells to produce insulin. This partially reversed a mouse model of type-1 diabetes.
The team also showed that they could recover muscle growth and function in mouse models of muscular dystrophy, a disease with a known gene mutation. According to the researchers, their strategy was not to correct the mutated gene but to increase the expression of genes in the same pathway as the mutated gene, thus overriding the effect of the mutation.
Further study will look at safety, practicality, and efficiency before moving it to a clinical environment, but preliminary data points toward safety and freedom from undesired genetic mutations.
To learn more about this advance, see the researchers' published findings in Cell.