Rescue of Skeletal Muscles of γ-Sarcoglycan- Deficient Mice with Adeno-Associated Virus-Mediated Gene Transfer

Abstract

In humans, a subset of cases of Limb-girdle muscular dystrophy (LGMD) arise from mutations in the genes encoding one of the sarcoglycan (alpha, beta, gamma, or delta) subunits of the dystrophin-glycoprotein complex. While adeno-associated virus (AAV) is a potential gene therapy vector for these dystrophies, it is unclear if AAV can be used if a diseased muscle is undergoing rapid degeneration and necrosis. The skeletal muscles of mice lacking gamma-sarcoglycan (gsg-/- mice) differ from the animal models that have been evaluated to date in that the severity of the skeletal muscle pathology is much greater and more representative of that of humans with muscular dystrophy. Following direct muscle injection of a recombinant AAV [in which human gamma-sarcoglycan expression is driven by a truncated muscle creatine kinase (MCK) promoter/enhancer], we observed significant numbers of muscle fibers expressing gamma-sarcoglycan and an overall improvement of the histologic pattern of dystrophy. However, these results could be achieved only if injections into the muscle were prior to the development of significant fibrosis in the muscle. The results presented in this report show promise for AAV gene therapy for LGMD, but underscore the need for intervention early in the time course of the disease process.

Noninvasive monitoring of therapeutic gene transfer in animal models of muscular dystrophies

“…The coding sequence of the human a-sarcoglycan is inserted between the AAV2 inverted terminal repeats (ITR) and is placed under the control of an MCK promoter fragment which contains the region À2155 to +7 base pairs from the transcription initiation site. 23 The pAAV-U7-SD23-BP22 carrying antisense sequences against splicing sequences of the murine dystrophin exon 23 has been previously described. 21 …”

“…For dystrophin, this has been carried out in the form of micro-dystrophin or of a modified U7 snRNA carrying an antisense sequence aimed at restoring the translational frame by inducing exon skipping and, for sarcoglycans, in the form of a normal version of the corresponding cDNA. [20][21][22][23][24][25] Before a therapeutic strategy can be implemented in a clinical trial, it is necessary to obtain objective indicators of therapeutic efficacy, especially since the approaches tested may have different outcomes and may correct some but not necessarily all the pathophysiological parameters. In the case of dystrophin or sarcoglycan deficiencies, a therapeutical benefit can be characterized by the restoration of the DGC at the sarcolemma, a correction of the dystrophic muscle histopathology or recovery of membrane integrity, and eventually by an increase in specific force and a decreased susceptibility to contraction-induced injury.…”

Adeno-associated virus vector-mediated gene transfer into dystrophin-deficient skeletal muscles evokes enhanced immune response against the transgene product

“…Several groups have previously demonstrated effective AAV vector-mediated gene transfer into skeletal muscles of mdx mice, 28 ␦-sarcoglycandeficient hamsters (Bio 14.6) 21,22 and ␥-sarcoglycandeficient mice. 23 On the other hand, Cordier et al 24 have shown that an AAV vector driven by CMV promoter elicits strong cellular and humoral immune responses to the transgene product after intramuscular injection into ␥-sarcoglycan-null mice.…”

“…[16][17][18]20 Thus, AAV vectors may be applicable to treatment of inherited neuromuscular disorders such as DMD. In fact, the AAV vector has been successfully introduced into ␦-sarcoglycan-deficient hamsters (Bio 14.6) 21,22 and ␥-sarcoglycan-deficient mice, 23 animal models of limb girdle muscular dystrophies (LGMD). A recent report, however, showed lower levels of transgene expression together with substantial immune response to the therapeutic transgene product in ␥-sarcoglycan-null mice.…”

Efficient and long-term intracardiac gene transfer in δ-sarcoglycan-deficiency hamster by adeno-associated virus-2 vectors

“…11,12 Currently, AAV vectors are the most promising gene delivery system in muscle tissues due to its efficient, widespread and persistent gene transfer. Although extensive studies have been carried out using AAV vectors in skeletal muscles for both muscular dystrophies [13][14][15][16][17][18] and for metabolic diseases, [19][20][21][22] few studies involved cardiac muscles.…”