A DNA cutting tool that can manipulate human genomic DNA could ultimately find applications in gene therapy, say Japanese scientists.

Makoto Komiyama, Narumi Shigi and colleagues at the University of Tokyo recently made the DNA cutter - ARCUT - and used it to cut bacterial DNA at one target site. Now they have shown that it can be tuned to cut human genomic DNA selectively and also to repair it.

ARCUT consists of a cerium(IV) complex which cuts the DNA and a recognition system of peptide nucleic acids, which resemble natural nucleic acids but with a peptide rather than a sugar backbone. The cutter's target site is pre-determined by base pairing between the DNA being cut and the bases in the cutter, which can be easily designed to bind a specific scission site. This is currently not possible with naturally occurring DNA cutting enzymes, which cut the genome repeatedly every 4⁴ or 4⁶ base pairs (approximately 700000 times in human DNA). ARCUT's selectivity meant that the Tokyo team was able to use the cutter to target one site in human genomic DNA - a system 1000 times larger than the Escherichia coli DNA they tested previously.

Site-specific cleavage of double-stranded DNA by ARCUT

The researchers have also shown that ARCUT can promote homologous recombination, a process in which nucleic acid fragments are exchanged between lengths of DNA. Whilst several technologies already exist that do this by introducing double strand breaks in DNA, their design and preparation is often difficult and slow. This is because they use protein based nucleic acid cleaving enzymes which recognise the DNA target site through complex protein-DNA interactions. 'We have been working on chemistry based artificial cutting enzymes believing that they could be the solution to these problems,' says Komiyama. 

Peter Nielsen works with peptide nucleic acids at the University of Copenhagen, Denmark, and is an expert in DNA recognition and gene targeting. He comments that, although further work is needed to test ARCUT in vivo, it is an exciting breakthrough. 'The DNA cleavage system is very interesting, being a truly chemical system with good efficiency,' he says. 'It is also interesting in connection with regimes for gene repair.' 

Indeed, the team has already used ARCUT to repair damaged DNA in human cells. The next step for the group is to apply the system in vivo so that it can be used for gene therapy and preliminary experiments have been encouraging. Komiyama's team has demonstrated that blue fluorescent protein incorporated into the human genome can be converted to its green analogue using ARCUT-promoted homologous recombination. 
 
Philippa Ross

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