The research, published in Cell, provides proof-of-concept for a tool that could one day be used to reverse age-related diseases such as hearing loss and macular degeneration in humans.
'Although many studies have demonstrated that CRISPR/Cas9 can be applied as a powerful tool for gene therapy, there are growing concerns regarding unwanted mutations generated by the double-strand breaks through this technology,' says Professor Juan Carlos Izpisua Belmonte at the Salk Institute in La Jolla, senior author of the paper. 'We were able to get around that concern.'
The CRISPR/Cas9 system typically snips double-strand breaks (DSBs: cutting through both strands of the double helix in one place) in targeted regions of the genome in order to edit DNA. But a DSB is the most lethal type of damage a cell can sustain, so these breaks would be risky in the DNA of living humans. The researchers' aim was to create a new version of the CRISPR/Cas9 genome editing technology that can affect the activation of genes without cutting the DNA strand at all.
To do this the scientists used CRISPR guide RNAs and a a non-cutting version of the enzyme Cas9, combining the specificity of this approach with molecular switches which are known to be effective in manipulating the epigenome but are prone to off-target effects.
As a proof-of-concept, Professor Belmonte's team used their new approach to treat several diseases, including type 1 diabetes, acute kidney disease, and muscular dystrophy in mice. In each case, they used their system to boost the expression of a specific endogenous gene which alleviated disease symptoms.
'We are not fixing the gene, the mutation is still there,' said Professor Belmonte. 'Instead ... 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 team is now working to improve the specificity of their system and to apply it to more cell types to treat a wider range of human diseases, as well as to reverse age-related conditions. More safety tests will be needed before human trials.