TY  - JOUR
A1  - Lehto, Taavi
A1  - Alvarez, Alejandra Castillo
A1  - Gauck, Sarah
A1  - Gait, Michael J.
A1  - Coursindel, Thibault
A1  - Wood, Matthew J. A.
A1  - Lebleu, Bernard
A1  - Boisguerin, Prisca
T1  - Cellular trafficking determines the exon skipping activity of Pip6a-PMO in mdx skeletal and cardiac muscle cells
JF  - Nucleic acids research
N2  - Cell-penetrating peptide-mediated delivery of phosphorodiamidate morpholino oligomers (PMOs) has shown great promise for exon-skipping therapy of Duchenne Muscular Dystrophy (DMD). Pip6a-PMO, a recently developed conjugate, is particularly efficient in a murine DMD model, although mechanisms responsible for its increased biological activity have not been studied. Here, we evaluate the cellular trafficking and the biological activity of Pip6a-PMO in skeletal muscle cells and primary cardiomyocytes. Our results indicate that Pip6a-PMO is taken up in the skeletal muscle cells by an energy-and caveolae-mediated endocytosis. Interestingly, its cellular distribution is different in undifferentiated and differentiated skeletal muscle cells (vesicular versus nuclear). Likewise, Pip6a-PMO mainly accumulates in cytoplasmic vesicles in primary cardiomyocytes, in which clathrin-mediated endocytosis seems to be the predominant uptake pathway. These differences in cellular trafficking correspond well with the exon-skipping data, with higher activity in myotubes than in myoblasts or cardiomyocytes. These differences in cellular trafficking thus provide a possible mechanistic explanation for the variations in exon-skipping activity and restoration of dystrophin protein in heart muscle compared with skeletal muscle tissues in DMD models. Overall, Pip6a-PMO appears as the most efficient conjugate to date (low nanomolar EC50), even if limitations remain from endosomal escape.
Y1  - 2014
U6  - https://doi.org/10.1093/nar/gkt1220
SN  - 0305-1048
SN  - 1362-4962
VL  - 42
IS  - 5
SP  - 3207
EP  - 3217
PB  - Oxford Univ. Press
CY  - Oxford
ER  -