Using CRISPR/Cas9 to Edit Disease Out of the Genome

By Kendall Morgan

CRISPR cured the mouse of cataractsThere can be no doubt that CRISPR/Cas9 technology has been a breakthrough for the genome-editing field. Now two studies reported in Cell Stem Cell last month show that this tool - already so useful in the laboratory - might also find its way to the clinic.

A team led by Jinsong Li from the Chinese Academy of Sciences found that mice with a dominant mutation in a gene that causes cataracts could be rescued by coinjection into zygotes of Cas9 mRNA and a single-guide RNA targeting the mutant allele. An independent team led by Hans Clevers at Hubrecht Institute in The Netherlands used the CRISPR/Cas9 genome editing system to correct the cystic fibrosis transmembrane conductor receptor (CFTR) by homologous recombination in cultured intestinal stem cells of patients with cystic fibrosis.

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CRISPR to the rescue

“I was really excited when I saw these two papers,” said the University of California Berkeley’s Jennifer Doudna, an Addgene depositor and CRISPR expert who was not involved in either study. “They are the first direct demonstration of use of genome editing to correct point mutations in two diseases with real phenotypic effect.”

The difference with CRISPR/Cas9 in comparison to other genome editing techniques is its ease of use. “It’s a very simple technology,” Doudna explained, giving credit to Addgene for their role in making the technology so readily available. “That’s why we’re seeing adoption of the technology in different labs for different applications.”

Li’s team noted that CRISPR/Cas9 has been used to generate mutations in a wide range of organisms, but its potential to efficiently correct disease hadn’t yet been realized. The researchers chose to explore its potential in mice with a dominant cataract disorder caused by a single copy of a gene known as Crygc. When all was said and done, the researchers had cured 24 mice of their disease.

Meanwhile, Hans Clevers and colleagues applied CRISPR to disease correction in adult stem cells isolated from two patients with cystic fibrosis. They were able to demonstrate functional correction of the gene in clonally expanded organoids.

“CRISPR in adult stem cells has huge potential, because the genomes of adult stem cells in organoid cultures are stable,” said Gerald Schwank, first author of the cystic fibrosis report. “This is a big advantage over induced pluripotent stem cells, which need to go through a phase of crisis where they 'collect' a number of mutations.”

Cystic fibrosis may not be the most likely candidate for gene therapy, however, because the disease affects multiple organ systems. Nevertheless, the findings are proof-of-principle for use of CRISPR to correct single-gene conditions.

Remaining hurdles in CRISPR biology

“I think it is absolutely possible to use CRISPR to cure genetic disease in the near future,” Li says, though there are some things to sort out first.

The potential for off-target effects is a major limitation. The procedure will also need to be perfected until the efficiency of repair approaches 100 percent.

“We need to be sure it's accurate enough and efficient enough for repair,” Doudna said. “And it can't be too hard to apply. These papers suggest it might work well enough to consider as a therapeutic.”

There are many fundamental questions about CRISPR/Cas biology still left to answer as well, she says, and those details will continue to be a primary preoccupation of her California lab and many others. “Those off-target effects are important,” she said. “We also really need to get a handle on how cleavage sites are repaired.” Those insights will be critical for scientists’ ability to influence editing efficiency.

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1. Yuxuan Wu et al. "Correction of a Genetic Disease in Mouse Via Use of CRISPR/Cas9." Cell Stem Cell. 13, 659-662 (5 December 2013).

2. Gerald Schwank et al. "Functional Repair of CFTR by CRISPR/Cas9 in Intestinal Stem Cell Organoids of Cystic Fibrosis Patients." Cell Stem Cell. 13, 653-658 (5 December 2013).


Topics: CRISPR, CRISPR Therapeutic Applications

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