Manuela Corti

The respiratory neuromuscular system in Pompe disease.

Pompe disease is due to mutations in the gene encoding the lysosomal enzyme acid α-glucosidase (GAA). Absence of functional GAA typically results in cardiorespiratory failure in the first year; reduced GAA activity is associated with progressive respiratory failure later in life. While skeletal muscle pathology contributes to respiratory insufficiency in Pompe disease, emerging evidence indicates that respiratory neuron dysfunction is also a significant part of dysfunction in motor units. Animal models show profound glycogen accumulation in spinal and medullary respiratory neurons and altered neural activity. Tissues from Pompe patients show central nervous system glycogen accumulation and motoneuron pathology. A neural mechanism raises considerations about the current clinical approach of enzyme replacement since the recombinant protein does not cross the blood-brain-barrier. Indeed, clinical data suggest that enzyme replacement therapy delays symptom progression, but many patients eventually require ventilatory assistance, especially during sleep. We propose that treatments which restore GAA activity to respiratory muscles, neurons and networks will be required to fully correct ventilatory insufficiency in Pompe disease.

B-cell depletion is protective against anti-AAV capsid immune response: a human subject case study

Gene therapy strategies for congenital myopathies may require repeat administration of adeno-associated viral (AAV) vectors due to aspects of the clinical application, such as: (i) administration of doses below therapeutic efficacy in patients enrolled in early phase clinical trials; (ii) progressive reduction of the therapeutic gene expression over time as a result of increasing muscle mass in patients treated at a young age; and (iii) a possibly faster depletion of pathogenic myofibers in this patient population. Immune response triggered by the first vector administration, and to subsequent doses, represents a major obstacle for successful gene transfer in young patients. Anti-capsid and anti-transgene product related humoral and cell-mediated responses have been previously observed in all preclinical models and human subjects who received gene therapy or enzyme replacement therapy (ERT) for congenital myopathies. Immune responses may result in reduced efficacy of the gene transfer over time and/or may preclude for the possibility of re-administration of the same vector. In this study, we evaluated the immune response of a Pompe patient dosed with an AAV1-GAA vector after receiving Rituximab and Sirolimus to modulate reactions against ERT. A key finding of this single subject case report is the observation that B-cell ablation with rituximab prior to AAV vector exposure results in non-responsiveness to both capsid and transgene, therefore allowing the possibility of repeat administration in the future. This observation is significant for future gene therapy studies and establishes a clinically relevant approach to blocking immune responses to AAV vectors.

Targeted approaches to induce immune tolerance for Pompe disease therapy

Enzyme and gene replacement strategies have developed into viable therapeutic approaches for the treatment of Pompe disease (acid α-glucosidase (GAA) deficiency). Unfortunately, the introduction of GAA and viral vectors encoding the enzyme can lead to detrimental immune responses that attenuate treatment benefits and can impact patient safety. Preclinical and clinical experience in addressing humoral responses toward enzyme and gene therapy for Pompe disease have provided greater understanding of the immunological consequences of the provided therapy. B- and T-cell modulation has been shown to be effective in preventing infusion-associated reactions during enzyme replacement therapy in patients and has shown similar success in the context of gene therapy. Additional techniques to induce humoral tolerance for Pompe disease have been the targeted expression or delivery of GAA to discrete cell types or tissues such as the gut-associated lymphoid tissues, red blood cells, hematopoietic stem cells, and the liver. Research into overcoming preexisting immunity through immunomodulation and gene transfer are becoming increasingly important to achieve long-term efficacy. This review highlights the advances in therapies as well as the improved understanding of the molecular mechanisms involved in the humoral immune response with emphasis on methods employed to overcome responses associated with enzyme and gene therapies for Pompe disease.

Evaluation of Readministration of a Recombinant Adeno-Associated Virus Vector Expressing Acid Alpha-Glucosidase in Pompe Disease: Preclinical to Clinical Planning

A recombinant serotype 9 adeno-associated virus (rAAV9) vector carrying a transgene that expresses codon-optimized human acid alpha-glucosidase (hGAA, or GAA) driven by a human desmin (DES) promoter (i.e., rAAV9-DES-hGAA) has been generated as a clinical candidate vector for Pompe disease. The rAAV9-DES-hGAA vector is being developed as a treatment for both early- and late-onset Pompe disease, in which patients lack sufficient lysosomal alpha-glucosidase leading to glycogen accumulation. In young patients, the therapy may need to be readministered after a period of time to maintain therapeutic levels of GAA. Administration of AAV-based gene therapies is commonly associated with the production of neutralizing antibodies that may reduce the effectiveness of the vector, especially if readministration is required. Previous studies have demonstrated the ability of rAAV9-DES-hGAA to correct cardiac and skeletal muscle pathology in Gaa(-/-) mice, an animal model of Pompe disease. This article describes the IND-enabling preclinical studies supporting the program for a phase I/II clinical trial in adult patients with Pompe. These studies were designed to evaluate the toxicology, biodistribution, and potential for readministration of rAAV9-DES-hGAA injected intramuscularly into the tibialis anterior muscle using an immune modulation strategy developed for this study. In the proposed clinical study, six adult participants with late-onset Pompe disease will be enrolled. The goal of the immune modulation strategy is to ablate B-cells before the initial exposure of the study agent in one leg and the subsequent exposure of the same vector to the contralateral leg four months after initial dosing. The dosing of the active agent is accompanied by a control injection of excipient dosing in the contralateral leg to allow for blinding and randomization of dosing, which may also strengthen the evidence generated from gene therapy studies in the future. Patients will act as their own controls. Repeated measures, at baseline and during the three months following each dosing will assess the safety, biochemical, and functional impact of the vector.

Inspiratory muscle conditioning exercise and diaphragm gene therapy in Pompe disease: Clinical evidence of respiratory plasticity.

ABSTRACT Pompe disease is an inherited disorder due to a mutation in the gene that encodes acid &agr;‐glucosidase (GAA). Children with infantile‐onset Pompe disease develop progressive hypotonic weakness and cardiopulmonary insufficiency that may eventually require mechanical ventilation (MV). Our team conducted a first in human trial of diaphragmatic gene therapy (AAV1‐CMV‐GAA) to treat respiratory neural dysfunction in infantile‐onset Pompe. Subjects (aged 2–15 years, full‐time MV: n = 5, partial/no MV: n = 4) underwent a period of preoperative inspiratory muscle conditioning exercise. The change in respiratory function after exercise alone was compared to the change in function after intramuscular delivery of AAV1‐CMV‐GAA to the diaphragm with continued exercise. Since AAV‐mediated gene therapy can reach phrenic motoneurons via retrograde transduction, we hypothesized that AAV1‐CMV‐GAA would improve dynamic respiratory motor function to a greater degree than exercise alone. Dependent measures were maximal inspiratory pressure (MIP), respiratory responses to inspiratory threshold loads (load compensation: LC), and physical evidence of diaphragm activity (descent on MRI, EMG activity). Exercise alone did not change function. After AAV1‐CMV‐GAA, MIP was unchanged. Flow and volume LC responses increased after dosing (p < 0.05 to p < 0.005), but only in the subjects with partial/no MV use. Changes in LC tended to occur on or after 180 days. At Day 180, the four subjects with MRI evidence of diaphragm descent had greater maximal voluntary ventilation (p < 0.05) and tended to be younger, stronger, and use fewer hours of daily MV. In conclusion, combined AAV1‐CMV‐GAA and exercise training conferred benefits to dynamic motor function of the diaphragm. Children with a higher baseline neuromuscular function may have greater potential for functional gains. HighlightsChildren with Pompe disease and ventilatory insufficiency received AAV1‐CMG‐GAA.Exercise alone did not change respiratory muscle function in any subjects.In less‐affected children, dynamic respiratory function improved after gene therapy.

Altered activation of the diaphragm in late-onset Pompe disease

Pompe disease is an inherited neuromuscular disorder that affects respiratory function and leads to dependence on external ventilatory support. We studied the activation of the diaphragm using bilateral phrenic magnetic stimulation and hypothesized that diaphragm compound muscle action potential (CMAP) amplitude and evoked transdiaphragmatic pressure (Twitch PDI) would correlate to disease severity. Eight patients with late onset Pompe disease (LOPD, aged 14-48 years) and four healthy control subjects completed the tests. Maximal Twitch PDI responses were progressively reduced in patients with LOPD compared to control subjects (1.4-17.1cm H2O, p<0.001) and correlated to voluntary functional tests (p<0.05). Additionally, CMAP amplitude (mA) was lower in the patients who used nighttime or fulltime ventilatory support, when compared to controls and patients who used no ventilatory support (p<0.005). However, the normalized (%peak) Twitch PDI and CMAP responses were similar between patients and controls. This suggests a loss of functional phrenic motor units in patients, with normal recruitment of remaining motor units.

Altered activation of the tibialis anterior in individuals with Pompe disease: Implications for motor unit dysfunction

Pompe disease is a progressive disease that affects skeletal muscles and leads to loss of ambulation. We investigated the activation of the tibialis anterior (TA) in late‐onset Pompe disease (LOPD) individuals during maximal voluntary contraction (MVC) and evoked involuntary responses.

Adeno-Associated Virus–Mediated Gene Therapy for Metabolic Myopathy

Metabolic myopathies are a diverse group of rare diseases in which impaired breakdown of stored energy leads to profound muscle dysfunction ranging from exercise intolerance to severe muscle wasting. Metabolic myopathies are largely caused by functional deficiency of a single gene and are generally subcategorized into three major types of metabolic disease: mitochondrial, lipid, or glycogen. Treatment varies greatly depending on the biochemical nature of the disease, and unfortunately no definitive treatments exist for metabolic myopathy. Since this group of diseases is inherited, gene therapy is being explored as an approach to personalized medical treatment. Adeno-associated virus-based vectors in particular have shown to be promising in the treatment of several forms of metabolic myopathy. This review will discuss the most recent advances in gene therapy efforts for the treatment of metabolic myopathies.

Respiratory motor function in individuals with centronuclear myopathies.

Introduction: Individuals with X‐linked myotubular myopathy (XLMTM) and other centronuclear myopathies (CNMs) frequently have profound respiratory insufficiency that requires support early in life. Still, few quantitative data exist to characterize respiratory motor function in CNM. Methods: We evaluated the reliance upon mechanical ventilation (MV), ventilatory kinematics, unassisted tidal volumes, and maximal respiratory pressures in 14 individuals with CNMs, including 10 boys with XLMTM. Results: Thirteen participants required full‐time, invasive MV. Maximal inspiratory pressures were higher in subjects who breathed unsupported at least 1 hour/day as compared with 24‐hour MV users [33.7 (11.9–42.3) vs. 8.4 (6.0–10.9) cm H2O, P < 0.05]. Years of MV dependence correlated significantly with MEP (r = −0.715, P < 0.01). Conclusions: Respiratory function in CNMs may be related to deconditioning from prolonged MV and/or differences in residual respiratory muscle strength. Results from this study may assist in evaluating severe respiratory insufficiency in neuromuscular clinical care and research. Muscle Nerve 53: 214–221, 2016

472. Evaluation of Re-Administration of a Recombinant Adeno-Associated Vector Expressing Acid-Alpha-Glucosidase (rAAV9-DES-hGAA) in Pompe Disease: Preclinical to Clinical Planning

A recombinant serotype 9 adeno-associated virus (rAAV9) vector carrying a transgene that expresses codon optimized human acid alpha-glucosidase (hGAA, or GAA) driven by a human desmin (DES) promoter (i.e. rAAV9-DES-hGAA) has been generated as a clinical candidate vector for Pompe disease. The rAAV9-DES-hGAA vector is being developed as a treatment for both early and late onset Pompe disease, in which patients lack sufficient lysosomal alpha-glucosidase leading to glycogen accumulation. In young patients, the therapy may need to be re-administered after a period of time to maintain therapeutic levels of GAA. Administration of AAV-based gene therapies is commonly associated with the production of neutralizing antibodies (NAb) that may reduce the effectiveness of the vector, especially if re-administration is required. Previous studies have demonstrated the ability of rAAV9-DES-hGAA to correct cardiac and skeletal muscle pathology in Gaa−/− mice, an animal model of Pompe disease. We describe the IND-enabling pre-clinical studies supporting the program for a phase I/II clinical trial in adult patients with Pompe. These studies were designed to evaluate the toxicology, biodistribution, and potential for re-administration of rAAV9-DES-hGAA injected intramuscularly into the tibialis anterior (TA) muscle using an immune modulation strategy developed for this study. In the proposed clinical study, six adult participants with Late-Onset Pompe Disease (LOPD) will be enrolled. The goal of the immune modulation strategy is to ablate B-cells prior to the initial exposure of the study agent in one leg and the subsequent exposure of the same vector to the contralateral leg four months after initial dosing. The dosing of active agent is accompanied by a control injection of excipient dosing in the contralateral leg to allow for blinding and randomization of dosing, which may also strengthen the approach to gene therapy studies in the future. Patients will act as their own controls. Repeated measures, at baseline and during the 3 months following each injection, will assess the safety, biochemical, and functional impact of the vector.View Large Image | Download PowerPoint Slide