Effective delivery of antisense peptide nucleic acid oligomers into cells by anthrax protective antigen

Abstract Peptide nucleic acid ( PNA ) is highly stable and binds to complementary RNA and DNA with high affinity, but it resists cellular uptake, thereby limiting its bioavailability. We investigated whether protective antigen ( PA , a non-toxic component of anthrax toxin) could transport antisense PNA oligomers into reporter cells that contain luciferase transgenes with mutant β-globin IVS2 intronic inserts, which permit aberrant pre-mRNA splicing and impair luciferase expression. PNA oligomers antisense to mutant splice sites in these IVS2 inserts induced luciferase expression when effectively delivered into the cells. PNA 18-mers with C-terminal poly-lysine tails [ PNA (Lys) 8 ] demonstrated modest sequence-specific antisense activity by themselves at micromolar concentrations in luc - IVS2 reporter cell cultures. However, this activity was greatly amplified by PA . Antisense PNA (Lys) 8 with but not without PA also corrected the IVS2-654 β-globin splice defect in cultured erythroid precursor cells from a patient with β-thalassemia [genotype, IVS2-654(β 0 /β E ) ], providing further evidence that anthrax PA can effectively transport antisense PNA oligomers into cells.