Rescue of Pompe disease in mice by AAV-mediated liver delivery of secretable acid α-glucosidase

Abstract

Glycogen storage disease type II or Pompe disease is a severe neuromuscular disorder caused by mutations in the lysosomal enzyme, acid α-glucosidase (GAA), which result in pathological accumulation of glycogen throughout the body. Enzyme replacement therapy is available for Pompe disease; however, it has limited efficacy, has high immunogenicity, and fails to correct pathological glycogen accumulation in nervous tissue and skeletal muscle. Using bioinformatics analysis and protein engineering, we developed transgenes encoding GAA that could be expressed and secreted by hepatocytes. Then, we used adeno-associated virus (AAV) vectors optimized for hepatic expression to deliver the GAA transgenes to Gaa knockout (Gaa−/−) mice, a model of Pompe disease. Therapeutic gene transfer to the liver rescued glycogen accumulation in muscle and the central nervous system, and ameliorated cardiac hypertrophy as well as muscle and respiratory dysfunction in the Gaa−/− mice; mouse survival was also increased. Secretable GAA showed improved therapeutic efficacy and lower immunogenicity compared to nonengineered GAA. Scale-up to nonhuman primates, and modeling of GAA expression in primary human hepatocytes using hepatotropic AAV vectors, demonstrated the therapeutic potential of AAV vector–mediated liver expression of secretable GAA for treating pathological glycogen accumulation in multiple tissues in Pompe disease.

Pompe Disease: From Basic Science to Therapy

“…The authors showed in further in vivo experiments that these transgenes, delivered by liver-tropic rAAV, resulted in high levels of secreted GAA, low immunogenicity, and metabolic cross-correction in muscle, central nervous system, and spinal cord. Experiments in nonhuman primates demonstrated the safety of this approach and confirmed that the liver-secreted GAA is efficiently taken up by peripheral tissues [154].…”

“…A more recent preclinical study sought to optimize the liver-directed strategy by testing genetically engineered GAA transgenes (that were codon optimized and contained small deletions within the progene and modified secretion signals) for their ability to be expressed by hepatocytes and produce the highly secretable GAA protein [154]. The authors showed in further in vivo experiments that these transgenes, delivered by liver-tropic rAAV, resulted in high levels of secreted GAA, low immunogenicity, and metabolic cross-correction in muscle, central nervous system, and spinal cord.…”

Targeted delivery of acid alpha-glucosidase corrects skeletal muscle phenotypes in Pompe disease mice

“…Studies normalizing glycogen levels solely in the brain and CNS in PD mice (using AAV gene therapy) have shown correction of some neuromuscular phenotypes even in the absence of improvement in cardiac or skeletal muscle fibers, demonstrating that a full reversion of clinical phenotypes in PD ERT will require multisystem delivery 30,31 . While we and others have seen a trend or partial reduction in CNS glycogen storage (Supplementary Figure S9) 27 , it remains unknown whether this reduction will be clinically significant in PD patients. To maximize enzyme delivery to affected tissue types, an effector like CD63, which -in addition to cardiac and skeletal musclesis also expressed in the CNS, would likely be more appropriate than more tissue-restricted internalizers like ITGA7.…”

“…α-hCD63huSC:GAA has higher glycogen removal at equivalent serum levels compared to GAA To determine whether the superior efficacy of α-hCD63SC:GAA is due to both improved targeting as well as improved secretion of α-hCD63SC:GAA compared to GAA (Figures 2A, 2B, and S8), or to primarily only one of these factors, we used a GAA construct that has several modifications to improve expression and secretion. In this construct, AAV-LSP2 GAA 27 , the most notable change is a replacement of the endogenous signal peptide with a chymotrysinogen B2 signal peptide. Additionally, in our α-hCD63SC:GAA construct we changed the scFv to fully human α-hCD63 variable domains (α-hCD63huSC:GAA), as a fully human protein is desirable for possible translation to human trials.…”

Exploiting epigenetics for the treatment of inborn errors of metabolism

“…The ultimate goal of such viral gene therapy is to restore production of the functional protein for sustained periods. For those IEMs in which the corrected proteins should be either secreted by or expressed in easily transduced cells (eg, hepatocytes), gene therapy would indeed provide a realistic cure if the transgene insertion could be targeted to harmless sites in the genome. However, when using conventional viruses, the lack of integration control may cause insertional mutagenesis and consequently severe side effects such as cancer.…”