Targeted Gene Modification Using Triplex-Forming Oligonucleotides

In recent years, triplex-forming oligonucleotides (TFOs) have emerged as powerful tools for site-specific gene modification. Their sequence specificity, binding affinity, and ability to provoke repair and recombination make them promising reagents for altering gene expression. This chapter highlights the binding requirements for triplex formation, identifies a number of chemical modifications that have been used with some success, and discusses studies using TFOs for inhibiting transcription. It also reviews work done using TFOs and related molecules to direct site-specific DNA damage, inducing mutagenesis or sensitizing a site to recombination. TFOs were initially used as positioning devices for nonspecific mutagens but were later discovered to have mutagenic properties of their own in cells with functional nucleotide excision repair (NER) and transcription-coupled repair (TCR) pathways. In subsequent studies triplex formation was able to induce both intramolecular and intermolecular homologous recombination, revealing its potential application for gene therapy. Recent reports demonstrate the ability of these molecules to locate and modify their cognate sites in chromosomal DNA in both cell culture and live animals, laying the foundation for triplex technology in vivo.