Correcting Neuromuscular Deficits With Gene Therapy in Pompe Disease

Abstract

Objective We have recently reported on the pathology of the neuromuscular junction (NMJ) in Pompe disease, reflecting disruption of neuronal and muscle homeostasis as a result of glycogen accumulation. The aim of this study was to examine how the alteration of NMJ physiology contributes to Pompe disease pathology; we performed molecular, physiological, and histochemical analyses of NMJ-related measures of the tibialis anterior muscles of young-, mid-, and late-stage alpha-glucosidase (GAA)-deficient mice. Methods We performed intramuscular injection of an adeno-associated virus (AAV)9 vector expressing GAA (AAV9-hGAA) into the tibialis anterior muscle of Gaa–/– mice at early, mid, and severe pathological time points. We analyzed expression of NMJ-related genes, in situ muscle force production, and clearance of glycogen in conjunction with histological assessment of the NMJ. Results Our data demonstrate that AAV9-hGAA is able to replace GAA to the affected tissue and modify AChR mRNA expression, muscle force production, motor endplate area, and innervation status. Importantly, the degree of restoration for these outcomes is limited by severity of disease. Early restoration of GAA activity was most effective, whereas late correction of GAA expression was not effective in modifying parameters reflecting NMJ structure and function nor in force restoration despite resolution of glycogen storage in muscle. Interpretation Our data provide new mechanistic insight into the pathology of Pompe disease and suggest that early systemic correction to both neural and muscle tissues may be essential for successful correction of neuromuscular function in Pompe disease.

Copackaged AAV9 Vectors Promote Simultaneous Immune Tolerance and Phenotypic Correction of Pompe Disease

“…[37][38][39] Early restoration of enzyme activity to both neural and skeletal muscle tissue is likely to have the greatest effect on physiological outcomes related to treatment strategies as our studies and those of others have demonstrated time-dependent arrests in functional decline after gene therapy. 11,74,75 Reliance upon liver-derived GAA to target neural tissues is dependent upon crosscorrection of neighboring cells through secretion and reuptake of GAA, yet the protein is too large to cross the blood-brain barrier and therefore does not address the neuropathology related to Pompe disease. 6,31 For these reasons, ERT or exclusively hepatic gene transfer would not address glycogen accumulation particularly in the motor neurons involved with ambulation or respiration as evidenced by patients continuing to be wheelchairand ventilator-dependent after long-term treatment of ERT.…”

“…11,[42][43][44][45][46][47] In addition, we and others have described the capabilities of AAV-mediated expression of GAA to elicit detrimental immune responses against the transgene. 1,50,55 To address this complication for improved systemic outcomes, we pursued the use of a liver-specific promoter (apolipoprotein Ehepatocyte locus control region-human alpha-1 antitrypsin promoter [LSP]) to drive expression of human codon-optimized GAA (coGAA) for induction of GAA-specific T regs capable of dampening a systemic immune response against GAA.…”

“…11,37 Under anesthesia the skin and fascia surrounding the distal hind limb were surgically removed exposing the TA. The extensors digitorum and peroneus longus tendons near the ankle were cut to eliminate the influence of additional muscles.…”

“…51 With the hypothesis that the DES:LSP ratio would confer adequate cardiac and neuromuscular expression based on previous publications, we copackaged these constructs at that ratio to produce a single gene therapy product (copackaged DES:LSP). 11,46,47 We hypothesized that this would have the same benefits as producing these vectors separately and combining them but would provide a quicker and more costeffective method of production.…”

Profile of alglucosidase alfa in the treatment of Pompe disease: safety, efficacy, and patient acceptability

“…Therefore, an early therapy appears to be the most effective. 79 First clinical trials of gene transfer treatment in five ventilator-dependent Pompe patients demonstrated acceptable safety outcomes and longer periods of unassisted breathing. 80,81 Genome editing via CRISPR/Cas9 82 is another revolutionary approach that might tremendously change the therapeutic options for Pompe patients by modulating disease-causing alleles.…”