G.P.279: Efficient AAV-mediated transfer of FKRP in a new mouse model of Limb Girdle Muscular Dystrophy 2I

Abstract

Dystroglycanopathies constitute a group of genetic diseases caused by defective glycosylation of alpha-dystroglycan (aDG), a membrane glycoprotein involved in the cell/matrix anchoring of muscle fibers. The aDG glycosylation, a very complex process, requires many proteins whose functions are not fully elucidated. In particular, mutations in the FKRP gene encoding Fukutin related protein, lead to hypoglycosylation of aDG, resulting in different forms of dystroglycanopathies, among which Limb Girdle Muscular Dystrophy type 2I (LGMD2I). We generated a knock-in mouse model of LGMD2I, carrying the most frequent mutation (L276I) encountered in LGMD2I patients. Molecular characterization of this mouse model showed that the introduction of the mutation did not alter the expression of FKRP. However, the protein appears to have altered function since abnormal glycosylation of aDG and reduction of laminin binding was observed. Histologically, the muscles of this model show a dystrophic pattern starting from 6 months of age, consisting both in the presence of central nuclei and in fiber size variability. Interestingly, functional muscle impairment can be observed as early as 2 months of age by a decrease of the muscle resistance to eccentric mechanical stress. To evaluate gene transfer as a therapeutic approach, we cloned the FKRP cDNA in an AAV vector under the transcriptional control of the desmin promoter. The recombinant AAV2/9 vector was injected intramuscularly or intravenously in the mouse model. Expression of the FKRP transgene was obtained, both at RNA and protein levels. The glycosylation of aDG was restored as well as laminin binding. A histological rescue was observed by the decrease of fibers with centrally located nuclei. The AAV vector also improved the muscle function, since it conferred a better resistance to eccentric stress to the injected muscles.