Targeted gene repair -- in the arena

Abstract

The development of targeted gene repair is under way and, despite some setbacks, shows promise as an alternative form of gene therapy. This approach uses synthetic DNA molecules to activate and direct the cell's inherent DNA repair systems to correct inborn errors. The progress of this technique and its therapeutic potential are discussed in relation to the treatment of genetic diseases.

Figure 1

Proposed mechanism of targeted gene repair directed by chimeric RNA/DNA oligonucleotides. The RNA/DNA chimera (inset) is a single-stranded oligonucleotide consisting of RNA and DNA residues. The molecule acts by annealing at the site in the target DNA, with the RNA section (yellow) hybridizing with perfect complementarity to one strand and the DNA stretch (blue) hybridizing to the other strand. A T/G base pair mismatch is formed and is acted upon by the cell’s DNA repair systems. In this case, the G residue is excised and replaced by an A to base pair with the T base provided by the chimera. The chimera dissociates, leaving behind a C/A mismatch, which is presumably corrected by the mismatch-repair system to generate a T•A base pair. Here, the action of the chimera directs the exchange of a C•G base pair with a T•A base pair. Reproduced with permission from Science’s STKE (46).

Figure 2

Targeted gene repair with modified single-stranded oligonucleotides. The mutant base is recognized by the single-stranded oligonucleotide, which is designed to hybridize to a region immediately surrounding the targeted base. At one specific site, a mismatch is created (C/T here) with the displacement of the complementary strand, which contains the G residue (G). DNA repair activity catalyzed by the cellular machinery inserts an A on the hybridized strand to pair with the T residue of the template. A second repair event completes the cycle by removing the G residue and replacing it with a T. In this case, a conversion of the G•C base pair to an A•T base pair results from the gene repair reaction. Reproduced with permission from Current Opinion in Molecular Therapeutics (47).