Gene therapy for Canavan's disease takes a step forward

Canavan's disease (CD) is a rare but devastating pediatric leukodystrophy that causes progressive spongy neurodegeneration and is invariably fatal in congenital form.1 The disease is associated with >54 loss-of-function mutations2,3,4 in the enzyme aspartoacylase (ASPA), leads to accumulation of the substrate N-acetyl aspartic acid (NAA) in the brain, and is diagnosed via the presence of NAA aciduria.1 CD is characterized by dysmyelination, intramyelinic edema (leading to hydrocephalus), and extensive vacuolation of the central nervous system (CNS) white matter.5 Currently there is no established therapy that affects progression of the disease, and survival is based primarily on improved general medical care. A previous gene therapy attempt using liposome-encapsulated plasmid DNA6 had shown encouraging although transient decreases in local NAA concentrations in the treated brains, which prompted a gene therapy clinical protocol using recombinant AAV serotype 2 (rAAV2) in the hope of better dissemination of the vector and more sustainable NAA reductions.7 In a recent issue of Science Translational Medicine, Leone et al.8 report long-term follow-up of 13 of the 28 patients enrolled in this trial, who received intracranial injections of first-generation rAAV vectors-based on serotype 2 nearly a decade ago. The study evaluates the long-term safety, dosing parameters, and efficacy of the treatment. The findings suggest that widespread ASPA gene transfer throughout the entire CNS might be necessary for alleviating the extensive neuropathology and maximizing outcomes in patients with CD. Additionally, the findings underline the importance of early intervention, because improvements in younger patients appeared to be more pronounced than those in older patients. The study suggests that rAAV-mediated gene therapy is the most promising therapeutic modality for CD to date; it seems likely that gene therapy for CD and other inherited neurological diseases will advance rapidly in the near future as less invasive delivery methods and more efficient vectors for pan-CNS transductions are developed.9,10,11 The authors of the study had to deal with several challenges in addition to the rarity of the disease. Enrollment of age-matched children who showed similar trends in disease progression was difficult because of the sheer variety of mutations that cause CD. Patients with similar phenotypes and complete lack of ASPA activity were enrolled and grouped into cohorts based on age. Although the levels of NAA were not fully normalized by the gene therapy, there was an encouraging trend toward reductions in NAA in the treated brains. In-depth assessment of individual brain regions showed a statistically significant improvement over untreated subjects in one of the four regions assessed. Unfortunately, the varied rates of disease progression and the small number of patients confounded interpretation of the results. Indeed, only when the authors had removed the oldest cohorts from some analyses were they able to identify statistically significant changes. Nevertheless, the trial is a step forward in rAAV gene therapy attempts, showing encouraging but marginal improvements in the most characteristic feature of CD: NAA accumulation in the brain. The authors studied atrophy of brain mass by serial magnetic resonance imaging followed by digital image processing to estimate enlargement of ventricles, changes in morphology of anatomical landmarks, or decrease of white matter mass by cross-sectional measurements of regional thickness in brain slices. The gene therapy helped to stabilize atrophy and even slowed progression of the disease in some patients, but there was a lack of a uniform response across cohorts. Improvements of gross motor functions relative to age-matched controls are an important measure of alleviation of the CD phenotype. To this end, the authors conducted a battery of standardized tests following gene therapy and found that, although there were significant improvements in the patient groups (when the oldest cohort was removed from the analyses), the raw scores nevertheless fell within the range of spastic quadriplegic patients. However, these results should not be considered discouraging, because a previous preclinical study of intracranial rAAV2-AspA delivery to a naturally occurring CD disease model in rats did not result in motor function improvement.12 Moreover, neurological examinations showed statistically significant improvements in motor functions in the younger cohorts of treated CD patients, indicating the possible advantage of early therapeutic intervention. This result also underlines the importance of defining a therapeutic window for the patients that will help improve the quality of life in addition to prolonging it. The intensity and frequency of seizures in treated patients did not entirely reflect the effects of gene therapy; the patients remained on antiepileptic medication during the study. To assess improvements resulting from the gene therapy, the authors followed changes in the frequency and dosage of the anti­epileptic medications, and they found statistically significant reductions in the intensity and frequency of seizure episodes that were correlated with reduced dosages. The two documented post–gene therapy adverse events observed in the study were unrelated to either the vector or the delivery procedure. The absence of any major adverse event lends further support to the use of rAAVs as clinical vectors for CD gene therapy. Furthermore, this study is a harbinger of next-generation gene therapy for CD and other currently untreatable CNS disorders. The first-generation single-stranded rAAV2 vector has been useful for generating clinical safety data on the brain, eyes, lung, and liver. This study was initiated at a time when the more potent, self-complementary vector genome design was not yet available, and our expectations must be tempered accordingly. Nevertheless, the study is an important landmark characterized by several firsts: the first rAAV trial in the brain, the first academic production of clinical-grade rAAV for human use (by the Vector Core of the University of North Carolina at Chapel Hill), and treatment of the youngest patient (a 3-month-old infant) by gene transfer. The rapid advances in AAV vectorology suggest that next-generation AAV vectors—characterized by higher transduction efficiency in the CNS and the ability to cross the blood–brain barrier for pan-CNS distribution via intravascular delivery—should facilitate global CNS gene therapy for CD.9,10 This principle received support in a recently published preclinical study in which intravenous administration of a novel rAAV improved motor functions and mitigated disease phenotypes in a mouse model of spinal muscular atrophy, signifying that alternative second-generation rAAVs should permit successful gene therapy of currently untreatable CNS diseases, including CD.13