AAV-mediated intramuscular delivery of myotubularin corrects the myotubular myopathy phenotype in targeted murine muscle and suggests a function in plasma membrane homeostasis

Myotubular myopathy (XLMTM, OMIM 310400) is a severe congenital muscular disease due to mutations in the myotubularin gene (MTM1) and characterized by the presence of small myofibers with frequent occurrence of central nuclei. Myotubularin is a ubiquitously expressed phosphoinositide phosphatase with a muscle-specific role in man and mouse that is poorly understood. No specific treatment exists to date for patients with myotubular myopathy. We have constructed an adeno-associated virus (AAV) vector expressing myotubularin in order to test its therapeutic potential in a XLMTM mouse model. We show that a single intramuscular injection of this vector in symptomatic Mtm1-deficient mice ameliorates the pathological phenotype in the targeted muscle. Myotubularin replacement in mice largely corrects nuclei and mitochondria positioning in myofibers and leads to a strong increase in muscle volume and recovery of the contractile force. In addition, we used this AAV vector to overexpress myotubularin in wild-type skeletal muscle and get insight into its localization and function. We show that a substantial proportion of myotubularin associates with the sarcolemma and I band, including triads. Myotubularin overexpression in muscle induces the accumulation of packed membrane saccules and presence of vacuoles that contain markers of sarcolemma and T-tubules, suggesting that myotubularin is involved in plasma membrane homeostasis of myofibers. This study provides a proof-of-principle that local delivery of an AAV vector expressing myotubularin can improve the motor capacities of XLMTM muscle and represents a novel approach to study myotubularin function in skeletal muscle

Thérapie génique des maladies du motoneurone à l'aide de vecteurs dérivés des AAV

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Major Subsets of Human Dendritic Cells Are Efficiently Transduced by Self-Complementary Adeno-Associated Virus Vectors 1 and 2▿

Dendritic cells (DC) are antigen-presenting cells pivotal for inducing immunity or tolerance. Gene transfer into DC is an important strategy for developing immunotherapeutic approaches against infectious pathogens and cancers. One of the vectors previously described for the transduction of human monocytes or DC is the recombinant adeno-associated virus (rAAV), with a genome conventionally packaged as a single-stranded (ss) molecule. Nevertheless, its use is limited by the poor and variable transduction efficiency of DC. In this study, AAV type 1 (AAV1) and AAV2 vectors, which expressed the enhanced green fluorescent protein and were packaged as ss or self-complementary (sc) duplex strands, were used to transduce different DC subsets generated ex vivo and the immunophenotypes, states of differentiation, and functions of the subsets were carefully examined. We show here for the first time that a single exposure of monocytes (Mo) or CD34+ progenitors (CD34) to sc rAAV1 or sc rAAV2 leads to high transduction levels (5 to 59%) of differentiated Mo-DC, Mo-Langerhans cells (LC), CD34-LC, or CD34-plasmacytoid DC (pDC), with no impact on their phenotypes and functional maturation of these cells, compared to those of exposure to ss rAAV. Moreover, we show that all these DC subpopulations can also be efficiently transduced after commitment to their differentiation pathways. Furthermore, these DC subsets transduced with sc rAAV1 expressing a tumor antigen were potent activators of a CD8+-T-cell clone. Altogether, these results show the high potential of sc AAV1 and sc AAV2 vectors to transduce ex vivo conventional DC, LC, or pDC or to directly target them in vivo for the design of new DC-based immunotherapies

Intravenous Administration of Self-complementary AAV9 Enables Transgene Delivery to Adult Motor Neurons

Therapeutic gene delivery to the whole spinal cord is a major challenge for the treatment of motor neuron (MN) diseases. Systemic administration of viral gene vectors would provide an optimal means for the long-term delivery of therapeutic molecules from blood to the spinal cord but this approach is hindered by the presence of the blood–brain barrier (BBB). Here, we describe the first successful study of MN transduction in adult animals following intravenous (i.v.) delivery of self-complementary (sc) AAV9 vectors (up to 28% in mice). Intravenous MN transduction was achieved in adults without pharmacological disruption of the BBB and transgene expression lasted at least 5 months. Importantly, this finding was successfully translated to large animals, with the demonstration of an efficient systemic scAAV9 gene delivery to the neonate and adult cat spinal cord. This new and noninvasive procedure raises the hope of whole spinal cord correction of MN diseases and may lead to the development of new gene therapy protocols in patients