N-acetylaspartate supports the energetic demands of developmental myelination via oligodendroglial aspartoacylase.

Breakdown of neuro-glial N-acetyl-aspartate (NAA) metabolism results in the failure of developmental myelination, manifest in the congenital pediatric leukodystrophy Canavan disease caused by mutations to the sole NAA catabolizing enzyme aspartoacylase. Canavan disease is a major point of focus for efforts to define NAA function, with available evidence suggesting NAA serves as an acetyl donor for fatty acid synthesis during myelination. Elevated NAA is a diagnostic hallmark of Canavan disease, which contrasts with a broad spectrum of alternative neurodegenerative contexts in which levels of NAA are inversely proportional to pathological progression. Recently generated data in the nur7 mouse model of Canavan disease suggests loss of aspartoacylase function results in compromised energetic integrity prior to oligodendrocyte death, abnormalities in myelin content, spongiform degeneration, and motor deficit. The present study utilized a next-generation "oligotropic" adeno-associated virus vector (AAV-Olig001) to quantitatively assess the impact of aspartoacylase reconstitution on developmental myelination. AAV-Olig001-aspartoacylase promoted normalization of NAA, increased bioavailable acetyl-CoA, and restored energetic balance within a window of postnatal development preceding gross histopathology and deteriorating motor function. Long-term effects included increased oligodendrocyte numbers, a global increase in myelination, reversal of vacuolation, and rescue of motor function. Effects on brain energy observed following AAV-Olig001-aspartoacylase gene therapy are shown to be consistent with a metabolic profile observed in mild cases of Canavan disease, implicating NAA in the maintenance of energetic integrity during myelination via oligodendroglial aspartoacylase.

Comparison of Endovascular and Intraventricular Gene Therapy With Adeno-Associated Virus-α-L-Iduronidase for Hurler Disease.

BACKGROUND: Hurler disease (mucopolysaccharidosis type I [MPS-I]) is an inherited metabolic disorder characterized by deficiency of the lysosomal enzyme α-L-iduronidase (IDUA). Currently, the only therapies for MPS-I, enzyme replacement and hematopoietic stem cell transplantation, are generally ineffective for central nervous system manifestations. OBJECTIVE: To test whether brain-targeted gene therapy with recombinant adeno-associated virus (rAAV5)-IDUA vectors in an MPS-I transgenic mouse model would reverse the pathological hallmarks. METHODS: Gene therapy approaches were compared using intraventricular or endovascular delivery with a marker (rAAV5-green fluorescent protein) or therapeutic (rAAV5-IDUA) vector. To improve the efficiency of brain delivery, we tested different applications of hyperosmolar mannitol to disrupt the blood-brain barrier or ependymal-brain interface. RESULTS: Intraventricular delivery of 1 × 10 viral particles of rAAV5-IDUA with systemic 5 g/kg mannitol co-administration resulted in IDUA expression throughout the brain, with global enzyme activity >200% of the baseline level in age-matched, wild-type mice. Endovascular delivery of 1 × 10 viral particles of rAAV5-IDUA to the carotid artery with 29.1% mannitol blood-brain barrier disruption resulted in mainly ipsilateral brain IDUA expression and ipsilateral brain enzyme activity 42% of that in wild-type mice. Quantitative assays for glycosaminoglycans showed a significant decrease in both hemispheres after intraventricular delivery and in the ipsilateral hemisphere after endovascular delivery compared with untreated MPS-I mice. Immunohistochemistry for ganglioside GM3, another disease marker, showed reversal of neuronal inclusions in areas with IDUA co-expression in both delivery methods. CONCLUSION: Physiologically relevant biochemical correction is possible with neurosurgical or endovascular gene therapy approaches for MPS-I. Intraventricular or endovascular delivery of rAAV5-IDUA was effective in reversing brain pathology, but in the latter method, effects were limited to the ipsilateral hemisphere.

Improvement in clinical outcomes following optimal targeting of brain ventricular catheters with intraoperative imaging.

OBJECT: The accurate placement of cerebral ventricular shunt catheters in hydrocephalus is an important clinical problem. Malfunction of shunts remains their most common complication and greatest liability, and the influence of catheter position on shunt function remains poorly defined. The objectives of this study were as follows: 1) determine the accuracy of intraventricular catheter placement with respect to a historically favored target, defined as a 1-cm radius sphere at the anterior lip of the ipsilateral foramen of Monro; 2) confirm that this target represents a satisfactory site for frontal and occipital catheter placement by examining whether inaccuracy is associated with more shunt failures; and 3) determine whether catheter trajectory, use of image confirmation, or other factors are associated with either the accuracy or the longevity of shunts. METHODS: A retrospective cohort analysis was conducted on 236 patients with 426 ventricular shunts placed or revised at the University of Minnesota over a 10-year period. RESULTS: Accuracy of shunt placement was optimal in 43.9% of patients and suboptimal or poor in 56.1% of patients. Time to failure was significantly affected by the accuracy of catheter placement with respect to the ipsilateral foramen of Monro, with a 57% higher risk of failure with suboptimal placement (hazard ratio [HR] 1.57, 95% CI 1.26-1.96; p < 0.001) and a 66% higher risk with poor placement (HR 1.66, 95% CI 1.45-1.89; p < 0.001) relative to optimal placement. The odds of highly suboptimal or unacceptable placement were significantly increased by lack of any intraoperative imaging (OR 5.89, 95% CI 2.36-14.65; p < 0.001). Use of a nonfrontal posterior trajectory also showed a trend toward poor placement (OR 1.64, p = 0.138). CONCLUSIONS: The historical target for catheter tip placement within 1 cm of the foramen of Monro in the ipsilateral lateral ventricle was associated with significantly longer revision-free survival compared with other locations. This effect remained significant after adjusting for age and whether there was a prior history of shunting. The accuracy of catheter placement in both pediatric and adult patients was strongly associated with use of intraoperative fluoroscopic confirmation. In analyses comparing intraoperative fluoroscopy and no imaging, there was a non-statistically significant difference in the 3-year time to failure, but the worst-case scenario of catastrophic short-term failure was almost completely avoided with fluoroscopy. The authors conclude that accuracy of placement is critical for shunt survival, and that use of intraoperative imaging confirmation may optimize outcomes by avoiding the majority of unacceptable placements.

Long-term follow-up after gene therapy for canavan disease.

Canavan disease is a hereditary leukodystrophy caused by mutations in the aspartoacylase gene (ASPA), leading to loss of enzyme activity and increased concentrations of the substrate N-acetyl-aspartate (NAA) in the brain. Accumulation of NAA results in spongiform degeneration of white matter and severe impairment of psychomotor development. The goal of this prospective cohort study was to assess long-term safety and preliminary efficacy measures after gene therapy with an adeno-associated viral vector carrying the ASPA gene (AAV2-ASPA). Using noninvasive magnetic resonance imaging and standardized clinical rating scales, we observed Canavan disease in 28 patients, with a subset of 13 patients being treated with AAV2-ASPA. Each patient received 9 × 10(11) vector genomes via intraparenchymal delivery at six brain infusion sites. Safety data collected over a minimum 5-year follow-up period showed a lack of long-term adverse events related to the AAV2 vector. Posttreatment effects were analyzed using a generalized linear mixed model, which showed changes in predefined surrogate markers of disease progression and clinical assessment subscores. AAV2-ASPA gene therapy resulted in a decrease in elevated NAA in the brain and slowed progression of brain atrophy, with some improvement in seizure frequency and with stabilization of overall clinical status.

Validating an Ataxia Functional Composite Scale in spinocerebellar ataxia.

The Ataxia Functional Composite Scale (AFCS) may provide a sensitive and reproducible assessment of treatment responses in studies of the spinocerebellar ataxias (SCA). We previously assessed the effects of buspirone in a cohort of patients with SCA via the International Cooperative Ataxia Rating Scale (ICARS). At each assessment period, AFCS scores were also obtained. A strong correlation of AFCS with ICARS scores was demonstrated at all assessment periods. This study supports the validity of the AFCS as a useful assessment of ataxia in this population.

Treatment of spinocerebellar ataxia with buspirone.

Preliminary data suggest potential benefit of 5-HT receptor agonists in the treatment of ataxias. We studied the effects of buspirone in a cohort of twenty patients with spinocerebellar ataxia (SCA). Twenty patients were treated in this double-blind, placebo controlled, cross-over trial with either buspirone HCl 30 mg twice daily or placebo for 3 months. Buspirone was not shown to be superior to placebo in the treatment of patients with SCA.

Mild-onset presentation of Canavan's disease associated with novel G212A point mutation in aspartoacylase gene.

We describe two sisters with a mild-onset variant of Canavan's disease who presented at age 50 and 19 months with developmental delay but without macrocephaly, hypotonia, spasticity, or seizures. Remarkably, both patients had age-appropriate head control, gross motor development, and muscle tone. There were very mild deficits in fine motor skills, coordination, and gait. Both sisters had a history of strabismus, but otherwise vision was normal. The older child showed evidence of mild cognitive and social impairment, whereas language and behavior were normal for age in the infant. Both patients were found to be compound heterozygotes for C914A (A305E) and G212A (R71H) mutations in ASPA. Like all other known ASPA mutations, this previously unknown G212A mutation appears to have low absolute enzyme activity. Nevertheless, it is associated in these patients with an extremely benign phenotype that is highly atypical of Canavan's disease. Biochemical and clinical data were evaluated using a generalized linear mixed model generated from 25 other subjects with Canavan's disease. There were statistically significant differences in brain chemistry and clinical evaluations, supporting a distinct variant of Canavan's disease. Future studies of ASPA enzyme structure and gene regulation in these subjects could lead to a better understanding of Canavan's pathophysiology and improvements in ASPA gene therapy.

Lithium citrate for Canavan disease.

Current evidence suggests that the effects of lithium on metabolic and signaling pathways in the brain may vary depending on the specific clinical condition or disease model. For example, lithium increases levels of cerebral N-acetyl aspartate in patients with bipolar disorder but does not appear to affect N-acetyl aspartate levels in normal human subjects. Conversely, lithium significantly decreases whole-brain levels of N-acetyl aspartate in a rat genetic model of Canavan disease in which cerebral N-acetyl aspartate is chronically elevated. While N-acetyl aspartate is a commonly used surrogate marker for neuronal density and correlates with neuronal viability, grossly elevated whole-brain levels of N-acetyl aspartate in Canavan disease are associated with dysmyelination and mental retardation. This report describes the first clinical application of lithium in a human subject with Canavan disease. Spectroscopic and clinical changes were observed over the time period in which lithium was administered, which reversed during a 2-week wash-out period after withdrawal of lithium. This investigation reports decreased N-acetyl aspartate levels in the brain regions tested and magnetic resonance spectroscopic values that are more characteristic of normal development and myelination, suggesting that a larger, controlled trial of lithium may be warranted as supportive therapy for Canavan disease by decreasing abnormally elevated N-acetyl aspartate.

Simultaneous high performance liquid chromatographic separation of purines, pyrimidines, N-acetylated amino acids, and dicarboxylic acids for the chemical diagnosis of inborn errors of metabolism.

OBJECTIVES: To set up a novel simple, sensitive, and reliable ion-pairing HPLC method for the synchronous separation of several purines, pyrimidines, N-acetylated amino acids, and dicarboxylic acids for the chemical diagnosis and screening of inborn errors of metabolism (IEM). DESIGN AND METHODS: The separation was set up using a Hypersil C-18, 5-microm particle size, 250 x 4.6 mm column, and a step gradient using two buffers and tetrabutylammonium hydroxide as the pairing reagent. A highly sensitive diode array UV detector was set up at a wavelength between 200 and 300 nm that revealed purines and pyrimidines at 260 nm and other compounds at 206 nm. RESULTS: Compounds were determined in the plasma of 15 healthy adults, in the urine of 50 healthy subjects (1-3 years, 4-6 years, 8-10 years, 12-18 years, 25-35 years), and in 10 non-pathological amniotic fluid samples. To assess the validity of the chemical diagnosis of IEM, plasma and urine samples were analyzed in patients affected by Canavan disease (n = 10; mean age 4.6 +/- 2.3). Low plasma levels of N-acetylaspartate (16.96 +/- 19.57 micromol/L plasma; not detectable in healthy adults) and dramatically high urinary N-acetylaspartate concentrations (1872.03 +/- 631.86 micromol/mmol creatinine; 450 times higher than that which was observed in age-matched controls) were recorded. Neither N-acetylglutamate nor N-acetylaspartylglutamate could be detected in the plasma or urine of controls or patients with Canavan disease. CONCLUSIONS: The results demonstrate the suitability of the present ion-pairing HPLC separation with UV detection of cytosine, cytidine, creatinine, uracil, uridine, beta-pseudouridine, adenine, 3-methyladenine, hypoxanthine, xanthine, xanthosine, inosine, guanosine, ascorbic acid, thymine, thymidine, uric acid, 1-methyluric acid, orotic acid, N-acetylaspartate, N-acetylglutamate, N-acetylaspartylglutamate, malonic acid, methylmalonic acid, GSH, and GSSG as a reliable method for the prenatal and neonatal chemical diagnosis and screening of IEM using biological fluids.