Résumé :
|
Several recent advances have pointed out a variety of possible therapeutic approaches in muscular dystrophies, from pharmacological treatments to gene therapy and cell therapy with different types of newly identified stem cells. A combination of these strategies might enhance the possibility of successful therapy. Recently, a forced exclusion (skipping) of a single exon was used to restore the reading frame, giving rise to a shorter, but still functional dystrophin protein in an animal model of DMD. Such endeavour has been accomplished in vivo in the mdx mouse model by using AAV vectors expressing antisense sequences linked to short nuclear RNAs. This approach seems highly safe and efficient but some crucial problems remain as the control of the immune response against the vector itself. To circumvent these problems, we thought to combine the exon-skipping rationale with an autologous cell therapy approach. In such a view, we have intended a lentiviral vector expressing a U7 snRNA containing antisense sequences designed to skip exon 51 in a significant subset of DMD patients. Our results demonstrated the possibility to apply this method for the re-expression of a functional dystrophin in subpopulations of DMD CD133+ stem cells. These cells were isolated from patients with deletion of exons 49 and 50 that leads to destruction of the mRNA reading frame. Previous work demonstrated the potentiality of this non dystrophic subpopulation to actively participate in muscle regeneration in a dystrophic animal model (scid/mdx). In this way, we showed, for the first time, the achievement of in vivo dystrophin recovery from exon skipping treated DMD CD133+ stem cells engrafted into the scid/mdx mice muscles. The exon skipping method confers the advantage to be directed against the mutated endogenous dystrophin transcripts, preserving the natural level of the protein. Moreover, autologous transplantation allows circumventing the problem of immune response encountered during gene therapy approaches using direct in vivo viral vector administrations. 1Stem cell Laboratory, Department of Neurological Science, Fondazione IRCCS Ospedale Maggiore Policlinico, Centro Dino Ferrari, University of Milan, via F. Sforza 35, 20122, Milan, Italy; 2Department of Experimental Medicine, University of Pavia, Human Physiology unit, via Forlanini 6, 27100, Pavia, Italy; 3 UMR S 787 (INSERM & UPMC), Institut de Myologie, Faculté de Médecine Pierre et Marie Curie, 105 bd de l'Hopital, 75634 Paris Cedex 13, France; 4UNISTEM, Centro Interdipartimentale di Ricerca sulle Cellule Staminali, University of Milan, via Balzaretti 9, 20133, Milan, Italy.
|