The slinky sister of CRISPR that 'skips' over regions in DNA that cause disease

The technique still has one big hurdle to cross before heading towards clinical trials.

Genetic engineering witnessed a revolution after the birth of the gene editing tool named CRISPR in 2010.

Since its first application in gene editing experiments in 2013, CRISPR has been cited close to 2000 times in published research, some of which are breakthroughs of their own, earning it a whole lot of mojo as 'Breakthrough of the Year' in 2015.

The groundbreaking tool saw somewhat of a reinvention this week, with University of Illinois researchers adjusting the original mechanism to skip small regions of a gene, in contrast to making breaks at the start of a gene sequence like the original CRISPR method does.

The new spin-off, which its inventors have coined 'CRISPR-SKIP', is described in a study published in the journal Genome Biology.

Setting itself apart

In most cells, strands of DNA are sectioned into regions flagged as 'exons' that proteins are made from. These are close, but not adjacent, to each other on a given strand of DNA, and have 'intervening' regions called introns between them which don't appear to code for proteins.

When proteins are being made, DNA is read from start to finish, but only exons coded into RNA for protein synthesis. Often, these exons are stitched into long fragments with multiple individual exons back-to-back in a single RNA sequence. These ensembles of mature RNA are 'spliced' or broken apart before proteins are made from them.

The new CRISPR-SKIP technique changes a single base at the start of a target exon. The cell's machinery would then view the anomaly as an intervening, non-coding sequence and ignore or 'skip' the sequence during protein synthesis. A resulting protein would be slightly altered and is sure to cause some of the other change in how the protein functions.

Permanent changes

"While skipping exons results in proteins that are missing a few amino acids, the resulting truncated proteins often retain partial or full activity – which may be enough to restore function in some genetic diseases," said Perez-Pinera, lead author of the study.

Targeting just one or two exons in one such disease, Duchenne muscular dystrophy (DMD), reduced the number of disease-causing mutations by 77 percent, something the authors describes as an 'exciting' prospect for the use of CRISPR-SKIP in therapy.

Lasting effects

So far, techniques for targeted exon skipping have been yielded transient results, requiring periodic injections of therapeutic DNA to sustain the treatment, which is lifelong in nearly every case.

“By editing a single base in genomic DNA using CRISPR-SKIP, we can eliminate exons permanently and, therefore, achieve a long-lasting correction of the disease with a single treatment,” said Alan Luu, author of the study to University of Illinois press.

“The process is also reversible if we would need to turn an exon back on.”

University of Illinois researchers Professor Pablo Perez-Pinera, graduate student Alan Luu, professor Jun Song and graduate student Michael Gapinske,inventors of the CRISPR-SKIP technology

University of Illinois researchers Professor Pablo Perez-Pinera, graduate student Alan Luu, professor Jun Song and graduate student Michael Gapinske,inventors of the CRISPR-SKIP technology

This strategy of exon-skipping has already proven useful in early studies for cancer, rheumatoid arthritis, Huntington’s disease, Duchenne muscular dystrophy (DMD) treatments, the study reports.

Therapeutic value

However, the technique still has one big hurdle to cross before heading towards clinical trials.

The authors conceded that CRISPR-SKIP causes some mutations that are off-target — a big no-no for novel therapies aimed at use in people.

“We hope that future improvements to gene editing technologies will increase the specificity of CRISPR-SKIP so we can begin to address some of the problems that have kept gene therapy from being widely applied in the clinic,” Professor Jun Song, a lead author of the study, said to the University's press.

The researchers have tested the technology in multiple cell types, including mice and human cells in healthy and cancerous conditions.

The technique now awaits further scrutiny in tests using live animal models of diseases — a first step in the long journey that any and all therapies with promise must brave before reaching a clinic near you.

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