Peptide-conjugated oligonucleotides evoke long-lasting myotonic dystrophy correction in patient-derived cells and mice
Arnaud F. Klein, … , Denis Furling, Matthew J.A. Wood
Arnaud F. Klein, … , Denis Furling, Matthew J.A. Wood
Published November 1, 2019; First published September 3, 2019
Citation Information: J Clin Invest. 2019;129(11):4739-4744. https://doi.org/10.1172/JCI128205.
View: Text | PDF
Categories: Concise Communication Therapeutics

Peptide-conjugated oligonucleotides evoke long-lasting myotonic dystrophy correction in patient-derived cells and mice

Abstract

Antisense oligonucleotides (ASOs) targeting pathologic RNAs have shown promising therapeutic corrections for many genetic diseases including myotonic dystrophy (DM1). Thus, ASO strategies for DM1 can abolish the toxic RNA gain-of-function mechanism caused by nucleus-retained mutant DMPK (DM1 protein kinase) transcripts containing CUG expansions (CUGexps). However, systemic use of ASOs for this muscular disease remains challenging due to poor drug distribution to skeletal muscle. To overcome this limitation, we test an arginine-rich Pip6a cell-penetrating peptide and show that Pip6a-conjugated morpholino phosphorodiamidate oligomer (PMO) dramatically enhanced ASO delivery into striated muscles of DM1 mice following systemic administration in comparison with unconjugated PMO and other ASO strategies. Thus, low-dose treatment with Pip6a-PMO-CAG targeting pathologic expansions is sufficient to reverse both splicing defects and myotonia in DM1 mice and normalizes the overall disease transcriptome. Moreover, treated DM1 patient–derived muscle cells showed that Pip6a-PMO-CAG specifically targets mutant CUGexp-DMPK transcripts to abrogate the detrimental sequestration of MBNL1 splicing factor by nuclear RNA foci and consequently MBNL1 functional loss, responsible for splicing defects and muscle dysfunction. Our results demonstrate that Pip6a-PMO-CAG induces long-lasting correction with high efficacy of DM1-associated phenotypes at both molecular and functional levels, and strongly support the use of advanced peptide conjugates for systemic corrective therapy in DM1.

Authors

Arnaud F. Klein, Miguel A. Varela, Ludovic Arandel, Ashling Holland, Naira Naouar, Andrey Arzumanov, David Seoane, Lucile Revillod, Guillaume Bassez, Arnaud Ferry, Dominic Jauvin, Genevieve Gourdon, Jack Puymirat, Michael J. Gait, Denis Furling, Matthew J.A. Wood

×

Figure 1

Pip6a-PMO corrects molecular and functional defects in HSA-LR mice.

Options: View larger image (or click on image) Download as PowerPoint
Pip6a-PMO corrects molecular and functional defects in HSA-LR mice. HSA-...
HSA-LR mice were injected with single or multiple doses of naked PMO-CAG7 (PMO) or Pip6a-PMO-CAG7 (Pip6a-PMO) by intravenous injection and analyzed 2 weeks after treatment. (A) Quantification of splicing correction induced by a single 12.5 mg/kg dose of Pip6a-PMO, or 12.5 or 200 mg/kg dose of PMO in gastrocnemius of treated mice (WT n = 4; HSA-LR n = 10; PMO n = 4; Pip6a-PMO n = 6). (B) Representative RT-PCR results and quantification of alternative splicing profiles in gastrocnemius (gast.) and quadriceps (quad.) of HSA-LR mice treated with 3 injections of PMO at 200 mg/kg or Pip6a-PMO at 12.5 mg/kg (WT n = 7; HSA-LR n = 16; PMO n = 4; Pip6a-PMO n = 8). (C) Representative maximal force/time curves obtained by in situ force measurements of Pip6a-PMO–treated mice (3 times 12.5 mg/kg) compared with WT and HSA-LR mice. (D) Correction of myotonia (measured as the area under the force/time curve during relaxation after maximal muscle contraction) in Pip6a-PMO–injected HSA-LR mice (WT n = 6; HSA-LR n = 7; Pip6a-PMO n = 8). Data are expressed as mean ± SEM, except for D (boxes, 25th–75th percentile; whiskers, min to max; black line, the mean). ***P < 0.001; ****P < 0.0001 by 1-way ANOVA with Newman-Keuls post hoc test. NS, not significant.