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Dystrophin restoration in skeletal, heart and skin arrector pili smooth muscle of mdx mice by ZM2 NP–AON complexes

Gene Therapy volume 17, pages 432–438 (2010)Cite this article

Abstract

Potentially viable therapeutic approaches for Duchenne muscular dystrophy (DMD) are now within reach. Indeed, clinical trials are currently under way. Two crucial aspects still need to be addressed: maximizing therapeutic efficacy and identifying appropriate and sensible outcome measures. Nevertheless, the end point of these trials remains painful muscle biopsy to show and quantify protein restoration in treated boys. In this study we show that PMMA/N-isopropil-acrylamide+ (NIPAM) nanoparticles (ZM2) bind and convey antisense oligoribonucleotides (AONs) very efficiently. Systemic injection of the ZM2–AON complex restored dystrophin protein synthesis in both skeletal and cardiac muscles of mdx mice, allowing protein localization in up to 40% of muscle fibers. The mdx exon 23 skipping level was up to 20%, as measured by the RealTime assay, and dystrophin restoration was confirmed by both reverse transcription-PCR and western blotting. Furthermore, we verified that dystrophin restoration also occurs in the smooth muscle cells of the dorsal skin arrector pili, an easily accessible histological structure, in ZM2–AON-treated mdx mice, with respect to untreated animals. This finding reveals arrector pili smooth muscle to be an appealing biomarker candidate and a novel low-invasive treatment end point. Furthermore, this marker would also be suitable for subsequent monitoring of the therapeutic effects in DMD patients. In addition, we demonstrate herein the expression of other sarcolemma proteins such as α-, β-, γ- and δ-sarcoglycans in the human skin arrector pili smooth muscle, thereby showing the potential of this muscle as a biomarker for other muscular dystrophies currently or soon to be the object of clinical trials.

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Acknowledgements

The Telethon Italy grants GGP05115, GUP07011 and GGP09093 (to AF) are acknowledged. Thanks are also due to Prof A Medici (Department of Chemistry, University of Ferrara) to the Industria Chimica Emiliana (ICE Reggio Emilia) Grant (to AF), and to TREAT-NMD Network of Excellence of EU FP7 n. 036825 (to LM and Telethon-Italy). We are also grateful to the ISS National AIDS Program grants, in support of the Nanoparticle technology platform, awarded to AC, LT and ML. The ZM2–AON compound and its effects have been patented at the University of Ferrara, Industrial Liaison Office, patent IP number TO2009A000782.

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Authors and Affiliations

  1. Department of Experimental and Diagnostic Medicine, Section of Medical Genetics, University of Ferrara, Ferrara, Italy

    A Ferlini, P Sabatelli, M Fabris, E Bassi, S Falzarano, F Gualandi, L Merlini & P Rimessi

  2. IGM-CNR, Unit of Bologna c/o IOR, Bologna, Italy

    P Sabatelli

  3. Department of Neurological Sciences and Vision, Section of Clinical Neurology, University of Verona, Verona, Italy

    G Vattemi

  4. Department of Biology and Evolution, University of Ferrara, Ferrara, Italy

    D Perrone

  5. Department of Human Anatomical Sciences, University of Bologna and Laboratory of Cell Biology, IOR, Bologna, Italy

    N M Maraldi

  6. Department of Environmental and Life Sciences INSTM, University of Eastern Piedmont, Alessandria, Italy

    K Sparnacci & M Laus

  7. Department of Histology, Microbiology, and Medical Biotechnology, University of Padua, Padua, Italy

    A Caputo, P Bonaldo & P Braghetta

Authors
  1. A Ferlini
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  2. P Sabatelli
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  3. M Fabris
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  5. S Falzarano
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  7. D Perrone
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  8. F Gualandi
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  9. N M Maraldi
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  10. L Merlini
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  12. M Laus
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  13. A Caputo
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  14. P Bonaldo
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  15. P Braghetta
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  16. P Rimessi
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Corresponding author

Correspondence to A Ferlini.

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Supplementary Information accompanies the paper on Gene Therapy website (http://www.nature.com/gt)

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Ferlini, A., Sabatelli, P., Fabris, M. et al. Dystrophin restoration in skeletal, heart and skin arrector pili smooth muscle of mdx mice by ZM2 NP–AON complexes. Gene Ther 17, 432–438 (2010). https://doi.org/10.1038/gt.2009.145

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