Now that the entire human genome, or set of genes, has been mapped and is at our disposal, the promise of gene therapy lies with being able to “edit” genes as they vary from individual to individual in ways that either cause or might be used to prevent a disease.
Recently, a Johns Hopkins research team headed by BrightFocus Grantee Don Zack, MD, PhD, announced they’ve improved upon the state-of-the-art tool for manipulating the genome in research settings. Lead author, Vinod Ranganathan, PhD, Don Zack, and colleagues at the Wilmer Eye Institute, have expanded CRISPR gene editing technology, which is essentially a way to use microscopic “scissors” to edit the spelling of DNA.
The group’s results were published online in the prestigious journal Nature Communications on August 8, with Ranganathan as first author. The modified CRISPR technique is expected to improve the speed and efficiency of gene function studies, aid in the development of new cellular models of diseases, and eventually help treat genetic conditions.
Zack, who’s a member of the ophthalmology faculty at Wilmer and also part of its Center for Genetic Engineering and Molecular Ophthalmology, has held several BrightFocus grants to study macular degeneration and glaucoma. He became focused on improving the CRISPR technology as part of his 2011-13 BrightFocus grant into factors that might protect retinal pigmented epithelium cells in macular degeneration.
Up ahead, “we plan to apply the [CRISPR] methodology to retina-related diseases,” he said in recent correspondence. “We thankfully acknowledged BrightFocus support in the manuscript.”
This recent discovery has the potential to help genetic research not only in retinal disease, but across a wide range of diseases. Most changes to genes in the research world are being done these days using the CRISPR technology. The Wilmer group could be improving the access of these “molecular scissors” across the entire genetic research world. Scientists may adopt this technique or something similar going forward.
“It’s gratifying to know that BrightFocus-funded research can have positive repercussions not only for the specific disease under examination, but in improving the tools for thousands of scientists worldwide,” said Guy Eakin, PhD, Bright Focus Vice President for Scientific Affairs.
How CRISPR Works To Slice Genes
First used as a gene editing tool in 2013, CRISPR technology permits precise modification of pieces of DNA from chromosomes. To do so, it relies on an enzyme that cuts DNA at a desired site after being directed there by molecules that are part of an immune function. Bacteria sent to battle viruses are programmed to recognize the DNA sequence where the slice is desired.
However, even though it accomplished DNA editing in weeks or months, compared with the months to years it used to take, the earlier version of CRISPR still had limitations. Rather than being able to cut the genome at any site — and using these cuts to disable genes or as open sites to insert new ones —it could only gain access to a limited subset of sites on the genome.
By reprogramming the coding CRISPR uses to identify its targets (incorporating the nucleotides adenine (A) or guanine (G), rather than just guanine) the Hopkins team more than doubled the number of accessible sites. Also, since further analysis revealed that target sites in the genome beginning with A are more often found near disease genes, that made the modified CRISPR technique even more useful for targeting areas of DNA that are of most interest for research and ultimately disease modification.
The team tested the new, improved CRISPR by inserting a gene in various cell lines that made the cells glow green. Then they expanded the experiment and used CRISPRs to mutate a gene responsible for a blinding eye disease known as retinitis pigmentosa (RP). After checking their results, they concluded that the improved technology may be useful for studying or eventually treating RP and similar macular degeneration genetic disorders affecting the eye.
“This new method gives us a lot more flexibility for genetic engineering,” explained Zack in a Hopkins news release.
“The old technique is like an express train that can only make stops every few miles along DNA. This new technique is like a local train — we can generate mutations more efficiently than we ever could before.”
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