In keeping with this hypothesis, increasing the hydrophobicity of the X linker above a threshold value ( Figure 2B ) while maintaining charge spacing in a series of (R-X-R) 4 -PMO analogs ( Figure 2B ) had little impact on cellular uptake (Figure 4 ) but decreased significantly splicing correction efficiency ( Figure 2C ). Being too hydrophobic might conceivably lead to entrapment into membranes and as a consequence might be detrimental to endosomal release. The parent and most active (R-Ahx-R) 4 -PMO conjugate was rather resistant to proteolytic degradation in serum but was still cleaved by cellular proteases (11) . It was thus anticipated that the (r-Ahx-R) 4 -PMO in which some L-Arg residues have been replaced by their D-analog would become more protease-resistant and as a consequence more active in the splicing correction assay. Unexpectedly, (r-Ahx-R) 4 -PMO was significantly less active than (R-Ahx-R) 4 -PMO thus indicating that metabolic stability is not a limiting factor at least in these in vitro experiments. Whether (r-Ahx-R) 4 -PMO might be of interest for in vivo applications will have to be evaluated using transgenic murine models for splicing correction. Whether the lower biological activity of (r-Ahx-R) 4 -PMO could be due to its increased affinity for heparin is a possibility. Alternatively, earlier work from our group (11) has shown that the peptide part of (R-Ahx-R) 4 -PMO was rapidly degraded in cells thus releasing free PMOs. It is fully possible that the more stable CPP entity may decrease the rate of endosomal release of PMO. Along the same lines, linking a splice correcting PNA and a CPP (R 6 Pen in this case) by a stable linker gave rise to a lower efficiency than in the case of a reducible disulfide linker (27) .