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Limb Girdle Muscular Dystrophies type 2I (LGMD2I), a recessive autosomal muscular dystrophy, is caused by mutations in the Fukutin Related Protein (FKRP) gene. It has been proposed that FKRP, a ribitol-5-phosphate transferase, is a participant in α-dystroglycan (αDG) glycosylation, which is important to ensure the cell/matrix anchor of muscle fibers. A LGMD2I knock-in mouse model was generated to express the most frequent mutation (L276I) encountered in patients. The expression of FKRP was not altered neither at transcriptional nor at translational levels, but its function was impacted since abnormal glycosylation of αDG was observed. Skeletal muscles were functionally impaired from 2 months of age and a moderate dystrophic pattern was evident starting from 6 months of age. Gene transfer with a rAAV2/9 vector expressing Fkrp restored biochemical defects, corrected the histological abnormalities and improved the resistance to eccentric stress in the mouse model. However, injection of high doses of the vector induced a decrease of αDG glycosylation and laminin binding, even in WT animals. Finally, intravenous injection of the rAAV-Fkrp vector into a dystroglycanopathy mouse model due to Fukutin (Fktn) knock-out indicated a dose-dependent toxicity. These data suggest requirement for a control of FKRP expression in muscles., (© The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Deficiency of the dysferlin protein presents as two major clinical phenotypes: limb-girdle muscular dystrophy type 2B and Miyoshi myopathy. Dysferlin is known to participate in membrane repair, providing a potential hypothesis to the underlying pathophysiology of these diseases. The size of the dysferlin cDNA prevents its direct incorporation into an adeno-associated virus (AAV) vector for therapeutic gene transfer into muscle. To bypass this limitation, we split the dysferlin cDNA at the exon 28/29 junction and cloned it into two independent AAV vectors carrying the appropriate splicing sequences. Intramuscular injection of the corresponding vectors into a dysferlin-deficient mouse model led to the expression of full-length dysferlin for at least 1 year. Importantly, systemic injection in the tail vein of the two vectors led to a widespread although weak expression of the full-length protein. Injections were associated with an improvement of the histological aspect of the muscle, a reduction in the number of necrotic fibers, restoration of membrane repair capacity and a global improvement in locomotor activity. Altogether, these data support the use of such a strategy for the treatment of dysferlin deficiency.

Limb-girdle muscular dystrophy (LGMD) is a hereditary myopathy presenting clinical and genetic heterogeneity. In 1991, a recessive form (LGMD2A) was linked to chromosome 15q in a genetic isolate from the Isle of La Réunion. Confirmation of this localization was subsequently reported in Brazilian and northern Indiana Amish pedigrees. Here we report the exclusion of the LGMD2A locus in six Amish kindreds from southern Indiana that are related by multiple consanguineous links to the same northern Indiana families in which the involvement of the chromosome 15 locus was previously demonstrated. These findings indicate unexpected genetic heterogeneity of LGMD in an Indiana Amish isolate. Furthermore, genetic analyses also ruled out the possible involvement of the chromosome 2 locus recently described (LGMD2B), thus demonstrating that a mutation within at least one additional locus leads to this condition. Several candidate genes putatively involved in neuromuscular disorders were also excluded.

A gene responsible for an autosomal recessive form of limb girdle muscular dystrophy (LGMD2, MIM number 253600) has been localized on chromosome 15. After genotyping additional markers of this chromosome, two were found to flank the disease locus within an interval that was assessed as 7 centiMorgans. The screening of the CEPH YAC libraries with the corresponding probes allowed the isolation of YACs which were used in fluorescence in situ hybridization to define the LGMD2 cytogenetic interval as 15q15.1-15q21.1. Four different approaches were pursued for the establishment of the physical map of this area which allowed the assembly of an uninterrupted YAC contig spanning an estimated 10-12 megabases, with an average STS resolution of 140 kb or for the 25 polymorphic microsatellites on this map, of 400 kb. Twelve genes and 25 genetic markers were positioned in this contig, which is constituted of a minimum of 10 clones.