Peptide nucleic acid delivery to human mitochondria

Peptide nucleic acids (PNAs) are synthetic polynucleobase molecules, which bind to DNA and RNA with high affinity and specificity. Although PNAs have enormous potential as anti-sense agents, the success of PNA-mediated gene t herapy will require efficient cellular uptake and sub-cellular trafficking. At present these mechanisms are poorly understood To address this, we have studied the uptake of biotinylated PNAs into cultured cell lines using flu orescence confocal microscopy. In human myoblasts, initial punctate stainin g was followed by the release of PNAs into the cytosol and subsequent local isation and concentration in the nucleus. To determine whether PNAs could a lso be used as therapeutic agents for mtDNA disease, we attempted to locali se PNAs to the mitochondrial matrix. When attached to the presequence pepti de of the nuclear-encoded human cytochrome c oxidase (COX) subunit VIII the biotinylated PNA was successfully imported into isolated organelles in vit ro. Furthermore, delivery of the biotinylated peptide-PNA to mitochondria i n intact cells was confirmed by confocal microscopy. These studies demonstr ate that biotinylated PNAs can be directed across cell membranes and to a s pecific sub-cellular compartment within human cells highlighting the import ance of these novel molecules for human gene therapy.