A NICER approach to genome editing developed

The gene editing technique CRISPR/Cas9 has allowed researchers to make precise and impactful changes to an organism’s DNA to fix mutations that cause genetic disease; however, this method can also result in unintended DNA mutations that may have negative effects. Japanese researchers have now developed a new gene editing technique, based on the creation of multiple small cuts in single DNA strands by an enzyme called a nickase, that is just as effective as CRISPR/Cas9 while significantly reducing these unintended mutations.

Traditional CRISPR/Cas9 editing uses small pieces of genetic code called guide RNAs and an enzyme called Cas9. The guide RNAs target a specific section of the DNA and the Cas9 enzyme initiates a break in the double-stranded DNA structure at this location. This double-strand break is key for initiating changes to the DNA.

However, cellular repair of double-strand breaks can lead to unintended DNA mutations, as well as the integration of exogenous DNA to the human genome, which raises safety concerns for clinical applications of CRISPR/Cas9 technology. To minimise these unintended mutations, the Osaka University-led research team investigated the use of Cas9 nickase, which creates single-strand breaks or ‘nicks’ in DNA that are typically repaired without causing mutations. Their work has been described in the journal Nature Communications.

“Each chromosome in the genome has a ‘homologous’ copy,” said lead author Akiko Tomita. “Using the NICER technique, heterozygous mutations — in which a mutation appears in one chromosome but not its homologous copy — are repaired using the unmutated homologous chromosome as a template.”

For their initial experiments, the research team used human lymphoblast cells with a known heterozygous mutation in a gene called TK1. When these cells were treated with nickase to induce a single cut in the TK1 region, TK1 activity was recovered at a low rate. However, when the nickase induced multiple nicks in this region on both homologous chromosomes, gene correction efficiency was enhanced approximately 17-fold via activation of a cellular repair mechanism.

“Further genomic analysis showed that the NICER technique rarely induced off-target mutations,” said senior author Shinichiro Nakada. “We were also pleased to find that NICER was able to restore the expression of disease-causing genes in cells derived from genetic diseases involving compound heterozygous mutations.”

Because the NICER method does not involve DNA double-strand breaks or the use of exogenous DNA, it appears to be a safe alternative to conventional CRISPR/Cas9 methods. NICER may thus represent a novel approach for the treatment of genetic diseases caused by heterozygous mutations.

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