AAVrh.10 delivery of novel APOE2-Christchurch variant suppresses amyloid and Tau pathology in Alzheimer's disease mice.

Gene therapy to treat hereditary disorders conventionally delivers the normal allele to compensate for loss-of-function mutations. More effective gene therapy may be achieved using a gain-of-function variant. We tested the hypothesis that AAVrh.

Expression and processing of mature human frataxin after gene therapy in mice.

Friedreich's ataxia is a degenerative and progressive multisystem disorder caused by mutations in the highly conserved frataxin (FXN) gene that results in FXN protein deficiency and mitochondrial dysfunction. While gene therapy approaches are promising, consistent induction of therapeutic FXN protein expression that is sub-toxic has proven challenging, and numerous therapeutic approaches are being tested in animal models. FXN (hFXN in humans, mFXN in mice) is proteolytically modified in mitochondria to produce mature FXN.

Validation of Anti-Adeno Associated Virus Serotype rh10 (AAVrh.10) Total and Neutralizing Antibody Immunogenicity Assays.

Immunogenicity assessment of Adeno-Associated Virus (AAV) vectors is a critical part of gene therapy drug development. Whether the assays are used for inclusion/exclusion criteria or to monitor the safety and efficacy of the gene therapy, they are critical bioanalytical assessments. While total anti-AAV assays are perceived as easier to develop and implement than neutralizing anti-AAV assays, the gene therapy field is still nascent, and it is not yet clear which of the assays should be implemented at what stage of drug development.

Assessment of Safety and Biodistribution of AAVrh.10hCLN2 Following Intracisternal Administration in Nonhuman Primates for the Treatment of CLN2 Batten Disease.

CLN2 disease is a fatal, childhood autosomal recessive disorder caused by mutations in ceroid lipofuscinosis type 2 (CLN2) gene, encoding tripeptidyl peptidase 1 (TPP-1). Loss of TPP-1 activity leads to accumulation of storage material in lysosomes and resultant neuronal cell death with neurodegeneration. Genotype/phenotype comparisons suggest that the phenotype should be ameliorated with increase of TPP-1 levels to 5-10% of normal with wide central nervous system (CNS) distribution.

Identification of Safe and Effective Intravenous Dose of AAVrh.10hFXN to Treat the Cardiac Manifestations of Friedreich's Ataxia.

Friedreich's ataxia (FA) is a life-threatening autosomal recessive disorder characterized by neurological and cardiac dysfunction. Arrhythmias and heart failure are the main cause of premature death. From prior studies in murine models of FA, adeno-associated virus encoding the normal human frataxin gene (AAVrh.

Migalastat Tissue Distribution: Extrapolation From Mice to Humans Using Pharmacokinetic Modeling and Comparison With Agalsidase Beta Tissue Distribution in Mice.

Approved therapies for Fabry disease (FD) include migalastat, an oral pharmacological chaperone, and agalsidase beta and agalsidase alfa, 2 forms of enzyme replacement therapy. Broad tissue distribution may be beneficial for clinical efficacy in FD, which has severe manifestations in multiple organs. Here, migalastat and agalsidase beta biodistribution were assessed in mice and modeled using physiologically based pharmacokinetic (PBPK) analysis, and migalastat biodistribution was subsequently extrapolated to humans.

Challenges in treating Pompe disease: an industry perspective.

Pompe disease is a rare inherited metabolic disorder of defective lysosomal glycogen catabolism due to a deficiency in acid alpha-glucosidase (GAA). Alglucosidase alfa enzyme replacement therapy (ERT) using recombinant human GAA (rhGAA ERT) is the only approved treatment for Pompe disease. Alglucosidase alfa has provided irrefutable clinical benefits, but has not been an optimal treatment primarily due to poor drug targeting of ERT to skeletal muscles.

Improved efficacy of a next-generation ERT in murine Pompe disease.

Pompe disease is a rare inherited disorder of lysosomal glycogen metabolism due to acid α-glucosidase (GAA) deficiency. Enzyme replacement therapy (ERT) using alglucosidase alfa, a recombinant human GAA (rhGAA), is the only approved treatment for Pompe disease. Although alglucosidase alfa has provided clinical benefits, its poor targeting to key disease-relevant skeletal muscles results in suboptimal efficacy.

Glucosylceramide and Glucosylsphingosine Quantitation by Liquid Chromatography-Tandem Mass Spectrometry to Enable In Vivo Preclinical Studies of Neuronopathic Gaucher Disease.

Gaucher disease (GD) is caused by mutations in the GBA1 gene that encodes the lysosomal enzyme acid β-glucosidase (GCase). Reduced GCase activity primarily leads to the accumulation of two substrates, glucosylceramide (GlcCer) and glucosylsphingosine (GlcSph). Current treatment options have not been shown to ameliorate the neurological pathology observed in the most severe forms of GD, clearly representing an unmet medical need.

Duvoglustat HCl Increases Systemic and Tissue Exposure of Active Acid α-Glucosidase in Pompe Patients Co-administered with Alglucosidase α.

Duvoglustat HCl (AT2220, 1-deoxynojirimycin) is an investigational pharmacological chaperone for the treatment of acid α-glucosidase (GAA) deficiency, which leads to the lysosomal storage disorder Pompe disease, which is characterized by progressive accumulation of lysosomal glycogen primarily in heart and skeletal muscles. The current standard of care is enzyme replacement therapy with recombinant human GAA (alglucosidase alfa [AA], Genzyme). Based on preclinical data, oral co-administration of duvoglustat HCl with AA increases exposure of active levels in plasma and skeletal muscles, leading to greater substrate reduction in muscle.

Coformulation of a Novel Human α-Galactosidase A With the Pharmacological Chaperone AT1001 Leads to Improved Substrate Reduction in Fabry Mice.

Fabry disease is an X-linked lysosomal storage disorder caused by mutations in the gene that encodes α-galactosidase A and is characterized by pathological accumulation of globotriaosylceramide and globotriaosylsphingosine. Earlier, the authors demonstrated that oral coadministration of the pharmacological chaperone AT1001 (migalastat HCl; 1-deoxygalactonojirimycin HCl) prior to intravenous administration of enzyme replacement therapy improved the pharmacological properties of the enzyme. In this study, the authors investigated the effects of coformulating AT1001 with a proprietary recombinant human α-galactosidase A (ATB100) into a single intravenous formulation.

A GCase chaperone improves motor function in a mouse model of synucleinopathy.

Mutation of the lysosomal hydrolase acid-β-glucosidase (GCase), which leads to reduced GCase activity, is one of the most frequent genetic risk factors for Parkinson's disease (PD) and promotes α-synuclein accumulation in the brain, a hallmark of PD and other synucleinopathies. Whether targeting GCase pharmacologically is a valid therapeutic strategy for sporadic PD in the absence of GCase mutation is unknown. We have investigated whether increasing the stability, trafficking, and activity of wild-type GCase could be beneficial in synucleinopathies by administering the pharmacological chaperone AT2101 (afegostat-tartrate, isofagomine) to mice that overexpress human wild-type α-synuclein (Thy1-aSyn mice).

The pharmacological chaperone AT2220 increases the specific activity and lysosomal delivery of mutant acid alpha-glucosidase, and promotes glycogen reduction in a transgenic mouse model of Pompe disease.

Pompe disease is an inherited lysosomal storage disorder that results from a deficiency in acid α-glucosidase (GAA) activity due to mutations in the GAA gene. Pompe disease is characterized by accumulation of lysosomal glycogen primarily in heart and skeletal muscles, which leads to progressive muscle weakness. We have shown previously that the small molecule pharmacological chaperone AT2220 (1-deoxynojirimycin hydrochloride, duvoglustat hydrochloride) binds and stabilizes wild-type as well as multiple mutant forms of GAA, and can lead to higher cellular levels of GAA.

Immune responses and hypercoagulation in ERT for Pompe disease are mutation and rhGAA dose dependent.

Enzyme replacement therapy (ERT) with recombinant human acid-α-glucosidase (rhGAA) is the only FDA approved therapy for Pompe disease. Without ERT, severely affected individuals (early onset) succumb to the disease within 2 years of life. A spectrum of disease severity and progression exists depending upon the type of mutation in the GAA gene (GAA), which in turn determines the amount of defective protein produced and its enzymatic activity.

Migalastat HCl reduces globotriaosylsphingosine (lyso-Gb3) in Fabry transgenic mice and in the plasma of Fabry patients.

Fabry disease (FD) results from mutations in the gene (GLA) that encodes the lysosomal enzyme α-galactosidase A (α-Gal A), and involves pathological accumulation of globotriaosylceramide (GL-3) and globotriaosylsphingosine (lyso-Gb3). Migalastat hydrochloride (GR181413A) is a pharmacological chaperone that selectively binds, stabilizes, and increases cellular levels of α-Gal A. Oral administration of migalastat HCl reduces tissue GL-3 in Fabry transgenic mice, and in urine and kidneys of some FD patients.

Pharmacological chaperones as therapeutics for lysosomal storage diseases.

Lysosomal enzymes are responsible for the degradation of a wide variety of glycolipids, oligosaccharides, proteins, and glycoproteins. Inherited mutations in the genes that encode these proteins can lead to reduced stability of newly synthesized lysosomal enzymes. While often catalytically competent, the mutated enzymes are unable to efficiently pass the quality control mechanisms of the endoplasmic reticulum, resulting in reduced lysosomal trafficking, substrate accumulation, and cellular dysfunction.

The pharmacological chaperone AT2220 increases recombinant human acid α-glucosidase uptake and glycogen reduction in a mouse model of Pompe disease.

Pompe disease is an inherited lysosomal storage disease that results from a deficiency in the enzyme acid α-glucosidase (GAA), and is characterized by progressive accumulation of lysosomal glycogen primarily in heart and skeletal muscles. Recombinant human GAA (rhGAA) is the only approved enzyme replacement therapy (ERT) available for the treatment of Pompe disease. Although rhGAA has been shown to slow disease progression and improve some of the pathophysiogical manifestations, the infused enzyme tends to be unstable at neutral pH and body temperature, shows low uptake into some key target tissues, and may elicit immune responses that adversely affect tolerability and efficacy.

Lysosomal dysfunction in a mouse model of Sandhoff disease leads to accumulation of ganglioside-bound amyloid-β peptide.

Alterations in the lipid composition of endosomal-lysosomal membranes may constitute an early event in Alzheimer's disease (AD) pathogenesis. In this study, we investigated the possibility that GM2 ganglioside accumulation in a mouse model of Sandhoff disease might be associated with the accumulation of intraneuronal and extracellular proteins commonly observed in AD. Our results show intraneuronal accumulation of amyloid-β peptide (Aβ)-like, α-synuclein-like, and phospho-tau-like immunoreactivity in the brains of β-hexosaminidase knock-out (HEXB KO) mice.

Co-administration with the pharmacological chaperone AT1001 increases recombinant human α-galactosidase A tissue uptake and improves substrate reduction in Fabry mice.

Fabry disease is an X-linked lysosomal storage disorder (LSD) caused by mutations in the gene (GLA) that encodes the lysosomal hydrolase α-galactosidase A (α-Gal A), and is characterized by pathological accumulation of the substrate, globotriaosylceramide (GL-3). Regular infusion of recombinant human α-Gal A (rhα-Gal A), termed enzyme replacement therapy (ERT), is the primary treatment for Fabry disease. However, rhα-Gal A has low physical stability, a short circulating half-life, and variable uptake into different disease-relevant tissues.

Identification and characterization of pharmacological chaperones to correct enzyme deficiencies in lysosomal storage disorders.

Many human diseases result from mutations in specific genes. Once translated, the resulting aberrant proteins may be functionally competent and produced at near-normal levels. However, because of the mutations, the proteins are recognized by the quality control system of the endoplasmic reticulum and are not processed or trafficked correctly, ultimately leading to cellular dysfunction and disease.

The pharmacological chaperone isofagomine increases the activity of the Gaucher disease L444P mutant form of beta-glucosidase.

Gaucher disease is caused by mutations in the gene that encodes the lysosomal enzyme acid beta-glucosidase (GCase). We have shown previously that the small molecule pharmacological chaperone isofagomine (IFG) binds and stabilizes N370S GCase, resulting in increased lysosomal trafficking and cellular activity. In this study, we investigated the effect of IFG on L444P GCase.

The pharmacological chaperone 1-deoxygalactonojirimycin reduces tissue globotriaosylceramide levels in a mouse model of Fabry disease.

Fabry disease is an X-linked lysosomal storage disorder caused by a deficiency in alpha-galactosidase A (alpha-Gal A) activity and subsequent accumulation of the substrate globotriaosylceramide (GL-3), which contributes to disease pathology. The pharmacological chaperone (PC) DGJ (1-deoxygalactonojirimycin) binds and stabilizes alpha-Gal A, increasing enzyme levels in cultured cells and in vivo. The ability of DGJ to reduce GL-3 in vivo was investigated using transgenic (Tg) mice that express a mutant form of human alpha-Gal A (R301Q) on a knockout background (Tg/KO), which leads to GL-3 accumulation in disease-relevant tissues.

Structure of acid beta-glucosidase with pharmacological chaperone provides insight into Gaucher disease.

Gaucher disease results from mutations in the lysosomal enzyme acid beta-glucosidase (GCase). Although enzyme replacement therapy has improved the health of some affected individuals, such as those with the prevalent N370S mutation, oral treatment with pharmacological chaperones may be therapeutic in a wider range of tissue compartments by restoring sufficient activity of endogenous mutant GCase. Here we demonstrate that isofagomine (IFG, 1) binds to the GCase active site, and both increases GCase activity in cell lysates and restores lysosomal trafficking in cells containing N370S mutant GCase.