Canavan Disease

Author: iris chiara salaroglio
Date: 01/08/2012



Canavan disease (CD), also called Canavan-Van Bogaert-Bertrand disease, aspartoacylase deficiency or aminoacylase 2 deficiency, is a neurodegenerative childhood disorder characterized by spongy degeneration of the white matter of the brain. It belongs to a group of genetic disorders called leukodystrophies, in which is predominant the degeneration of myelin.
There are three phenotypic variants of CD depending upon the type of mutation :
1-congenital, with the most severe symptoms at birth;
2-infantile, the most prevalent form, in which disease symptoms appear between 3 and 6 months;
3-juvenile with late onset of disease, normally after 5 years of age.
The disease progression in infants evolves rapidly, few patients survived up to 10 years of age or even longer.

Initially it was recognized by Globus and Strauss as spongy brain disease in 1928, then was described by Myrtelle May Canavan in 1931 as a spongy degeneration of the cerebral and cerebellar areas of the brain. Van Bogaert and Bertrand provided an overview of the clinical features with a detailed description in three Ashkenazi infants [1949], recognizing as an autosomal recessive disease. Hagenfeldt and colleagues identified this disease as “N-acetylaspartic aciduria” due to a deficiency of ASPA activity, finding reduced ASPA activity in skin fibroblasts from a child with severe leukodystrophy. They proposed that the observed dysmyelination in the CNS was due to a failure of NAA to serve as an acetate carrier of acetyl groups from mitochondria to cytoplasm for lipogenesis. In 1988 Matalon and colleagues were the first to connect N-acetylaspartic aciduria and ASPA deficiency specifically to Canavan disease analyzing three children with Canavan disease. Once enzyme identified, they cloned the gene in 1993; following they established the specific mutations and carrier frequency rate screening 4,000 Ashkenazi Jewish individuals.

The Diseases DatabaseURL

Epidemiology and Inheritance

Canavan disease is pan-ethnic, inherited in an autosomal recessive pattern.

It is more prevalent among Ashkenazi Jews of Eastern European extraction.
Interestingly, only two mutations were found to be the basis for Canavan disease among 98% of individuals of Ashkenazi Jewish ancestry. This limited number of mutations allows screening programmes to identify carriers on this population.
The predominant mutation among Jewish individuals is a missense mutation (Glu285Ala) with substitution of glutamic acid to alanine and it was found 83.6% mutations in 104 alleles from 52 unrelated Ashkenazi Jewish patients (Kaul et al., 1996; Sistermans et al., 2000).
The second predominant mutation is a nonsense mutation where the codon 231 for tyrosine is substituted by a termination codon (Tyr231X) and it was found in 13.4% of the alleles from Jewish patients (Kaul et al., 1996; Sistermans et al., 2000). Screening of healthy Jews reveals that l/37-1/40 is a carrier of one of these two mutations. The incidence of the disease in this population is estimated at l/6000.

In non-Jewish patients (individuals of European, Middle Eastern, Turkish, Gypsies and African American ancestry) the mutations are different and more diverse.
The most common Canavan mutation (about 40-48%) in non-Jewish patients of European ancestry is Ala305Glu which is a missense mutation in exon VI, substituting alanine to glutamic acid; it was observed in 35.7% of the 70 alleles from 35 unrelated non-Jewish patients (Kaul et al., 1994a,b). Many of the other mutations often occur only in one family or in few patients at low frequency with milder symptoms of CD.

MutationResidual Activity (%)TypeEthnic group
Ala305Glu0.0missenseNn Jewish

N-Acetylaspartate in the CNS: From neurodiagnostics to neurobiology

A mutation of aspartoacylase gene in a Turkish patient with Canavan disease


Infants with Canavan disease appear normal at birth, but usually show signs of delayed development and decreased muscle tone (hypotonia), including head lag between 2 and 6 months of age. By one year, macrocephaly is often evident, and motor development is severely impaired. Infants later develop optic atrophy, and hypotonicity converts to limb stiffness and spasticity. The affected children become increasingly debilitated with age, often including seizures and an inability to move voluntarily or swallow.

Possible Complications :Blindness, Inability to walk, Mental retardation, Exaggerated reflexes (hyperreflexia), Joint stiffness, Loss of tissue in the optic nerve of the eye (optic atrophy)


Canavan disease is genetic disorder due to deficiency of enzyme Aspartoacylase (ASPA), leading to a progressive fatal leukodystrophy in affected infants.

ASPA is also called aminoacylase II and hydrolyzes acetate from aspartic acid only, specifically from N-acetyl-L-aspartic acid (NAA). It differs from aminoacylase I that is not specific and hydrolyses acetate from all amino acids with the exception of aspartic acid.
It is a cytosolic protein, also capable of nuclear localization.
It’s encoded by ASPA gene, located on chromosome 17 (17p13ter).

Aspartoacylase is a homodimeric metalloenzyme of the carboxypeptidase family and each monomer binds one zinc ion. Its size is approximately 74 kDa and it has 313 amino acids, seven of which (Arg63, Asn70, Arg71, Tyr164, Arg168, Glu178 and Tyr288) form the catalytic site of the enzyme. Arg168 and Tyr288 stabilize NAA binding to the active site. Moreover human ASPA enzyme sequence is 86% identical to that of murine and the catalytic domains are 100% identical.
It’s located in various tissues, predominantly in kidney, liver and white matter of the brain, specifically in oligodendrocytes. It’s not present in neurons or astrocytes.
This enzyme is very important in Canavan desease which is determined by alteration or deficiency of ASPA gene.
This is demonstrated by two animal models of CD: the so-called Tremor rat and the ASPA knockout mouse.
The first is a stable line developed from a naturally occurring mutant with a genomic deletion on chromosome 11 spanning 4 genes, including the aspartoacylase gene, olfactory receptor gene, vanilloid receptor subtype I gene, and the calcium/ calmodulin-dependent protein kinase IV gene. It shows no ASPA activity in brain, and greatly reduced activity in kidney, and also exhibits increased brain NAA level and muscular tremors starting at about 2 weeks of age.
Similarly, the second model homozygous ASPA / knockout mice exhibits subcortical and white matter vacuolation with white matter spongiform degeneration and hypomyelination and all characteristics of CD.

The target of enzyme is NAA metabolite, the acetylated form of the amino acid aspartate, transported from neurons to the cytoplasm of oligodendrocytes. It’s normally hydrolyzed into acetate and aspartic acid and then the free acetate is converted to acetyl CoA by acetyl CoA synthetase, which is used in myelin lipid synthesis.
ASPA expression profile in the CNS follows the time course of postnatal myelination during early infancy. In fact infants born with Canavan disease are normal at birth because ASPA activity is not critical until myelin synthesis is dramatically increased shortly after birth.
Thus in brain CD patients lipid synthesis is reduced, due to a reduced supply of NAA-derived acetate. This is observed in the brains of ASPA / mice at the time of peak myelination with an approximately 80% reduction in free acetate levels. For these reasons, a dietary acetate supplementation, for example with glyceryl triacetate (GTA), may be a possible treatment for Canavan disease.

Lack of ASPA leads to a toxic NAA concentration in the brain, possibly causing osmotic dysregulation and intramyelinic water accumulation. The resulting chemical imbalance caused by high level of NAA interferes with the formation of myelin and nervous system developing. A buildup of NAA also leads to the progressive destruction of existing myelin around nerve cells. Nerve fibers without this protective covering malfunction and die, damaging the brain and causing the serious signs and symptoms of Canavan disease.
In 2000 Baslow said: ‘‘The demyelination in CD is probably not owing to the inability of oligodendrocytes to produce myelin, but to the continuous destruction of myelinating oligodendrocytes themselves by the NAA osmotic pressure generated in the sealed paranodal and the internodal regions’’.

High NAA levels are evident in the brain (and also in urine) through magnetic resonance spectroscopy (MRS), detecting detrimental effects in the CNS.

In this way it has been correlated with neuronal health or integrity.
Despite of CD, all other neurological disorders (Alzheimer disease, epilepsy, amyotrophic lateral sclerosis, schizophrenia, multiple sclerosis, AIDS, traumatic brain injury, stroke and non-neuronal brain tumors such as glioma) have reductions NAA levels resulting in neuronal loss or dysfunction.


is one of the most abundant metabolites in the central nervous system (CNS) and it is involved in all three major cell types in the CNS: neurons, astrocytes and oligodendrocytes (OL), mainly in progenitors and immature OL, while mature OL have undetectable levels of NAA.
It is considered as a marker for functional neurons in adult brain and it’s localized in neuronal mitochondria despite of ASPA which is expressed predominantly in oligodendrocytes.

Its metabolism is cyclic: NAA is metabolized by the enzyme glutamate carboxypeptidase to form a neurotransmitter, N-acetylaspartyl glutamate (NAAG), by combining with L-glutamate. NAAG is converted back into NAA and L-glutamate by glutamate carboxypeptidase II, localized to the astrocyte plasma membrane. Released NAA is an osmoregulator and is transferred transaxonally to OL within the WM of the brain, where is hydrolyzed into acetate and aspartic acid by the enzyme aspartoacylase (ASPA). The acetate and aspartic acid products, in the presence of N-acetyltransferase (NAT), can also synthesize NAA in the neuron mitochondria.

NAA has different roles in the nervous system.
First of all, it provides a critical source of acetate for myelin lipid synthesis in oligodendrocytes. NAA, as mentioned before, is essential for lipid synthesis and myelination in the CNS, especially during the peak of postnatal myelination development.
Following, NAA is the precursor for the enzyme-mediated biosynthesis of the important neuronal dipeptide N-acetylaspartylglutamate (NAAG); it is synthesized enzymatically from NAA and glutamate. NAAG lowers cAMP levels, decreases voltage dependent calcium conductance, regulates GABA receptor subunit expression, and inhibits synaptic release of GABA, confirming its regulatory role in release of glutamate. In CD NAAG, as NAA, increases causing a significant reduction (40%) in glutamate and suggesting an alteration in homeostasis of glutaminergic neurons and excitatory neurotransmission.
Other evidences show NAA acting as an organic osmolyte (counter the ‘‘anion deficit’’ in neurons) or as a cotransport substrate for a proposed ‘‘molecular water pump’’ that removes metabolic water from neurons. But NAA can’t be considered a real water pump because it doesn’t meet all of criteria for molecular water pump status: no cotransport protein has been identified and there is only scant evidence on possible gating mechanisms that specifically regulate NAA release from neurons; it’s not released under hyper-osmotic conditions, but it is released in a calcium-dependent manner upon stimulation of neuronal NMDA receptors.
So NAA seems being only a minor contributor to change in osmolarity.
Finally NAA is involved in facilitating energy metabolism in neuronal mitochondria via the aspartate aminotransferase reaction. NAA biosynthetic enzyme (Asp-NAT) acts to remove excess aspartate from the matrix via acetylation. This promots α-ketoglutarate formation from glutamate and energy production via the citric acid cycle. Thus, the aspartate aminotransferase pathway in neuronal mitochondria aids in the production of ATP by facilitating the oxidation of glutamate .

N-Acetylaspartate in the CNS: From neurodiagnostics to neurobiology

Canavan disease and the role of N-acetylaspartate in myelin synthesis

Canavan disease: a white matter disorder

Diagnosis and Prevention

The early diagnosis of CD is typically characterized by physical symptoms, which include leukodystrophy , megaloencephaly, mental retardation, optic nerve atrophy, motor function retardation, seizures, hypotonia that leads to spasticity and death. With few exceptions, these children are not usually born with the symptoms; the illness becomes evident beginning after a few months of postnatal life, or later, depending upon the type of mutation.
Following step is Biochemical test: Blood chemistry and CSF chemistry; NAA levels using gas chromatography-mass spectrometry (GC-MS) in urine (or amniotic fluid); ASPA enzyme activity assaying in cultured skin fibroblasts of patients, but it may not be reliable because the activity varies with culture conditions.
Genetic tests look for four common mutations in the aspartoacylase gene, two of which are mainly found in the Ashkenazi Jewish population.
NAA Based Cranial Imaging (Head CT scan, Head MRI scan) is very important for the Detection of CD and CNS Disorders .

Prenatal diagnosis is possible by quantifying NAA in amniotic liquid. However, there are some fluctuations in NAA levels during gestation that need to be confirmed by amniocentesis, which involves DNA analysis of amniotic cells for more precise gene mutation and early diagnosis of CD. The early screening for Jewish and prevalent form of non-Jewish mutations is helpful for early diagnosis.
Genetic counseling is recommended for people who want to have children and have a family history of Canavan disease. Testing should be considered if both parents are of Ashkenazi Jewish descents or carriers.

Canavan disease: a white matter disorder



The prognosis for Canavan disease is poor: this is often a fatal disorder. With Canavan disease the central nervous system breaks down. Patients are likely to become disabled. Death often occurs before 18 months of age, although some children with milder forms may survive into their teens and twenties .


There is no effective treatment or cure for this condition at this time. Treatment is only symptomatic and supportive. It provided help maximize nutrition, fight infection and protect breathing, but do not slow or reverse the progression of the disease.

The gene for Canavan disease has been located and now researchers are studying the potential benefits of gene therapy and stem cell therapy for people with Canavan disease, testing these potential therapeutic strategies in animal model. The main strategies, currently under investigation, are gene transfer to the brain in order to replace the mutated ASPA gene, metabolic therapy to provide the crucial missing metabolite (acetate) and enzyme therapy where the enzyme aspartoacylase is engineered to be able to enter the brain.

Modification of aspartoacylase for potential use in enzyme replacement therapy for the treatment of Canavan disease. 2011

Adeno-associated virus-mediated aspartoacylase gene transfer to the brain of knockout mouse for canavan disease. 2003

Restoration of aspartoacylase activity in CNS neurons does not ameliorate motor deficits and demyelination in a model of Canavan disease. 2005

Mouse neural progenitor cells differentiate into oligodendrocytes in the brain of a knockout mouse model of Canavan disease.2004

Canavan disease: studies on the knockout mouse.2006

Clinical Therapheutic Approaches

1. Acetate deficiency in OL is responsible of CD, so acetate supplementation has been proposed as a therapeutic approach to this fatal disease. Studies using mice at varying stages of development show glyceryl triacetate (GTA) to be significantly more effective as an acetate source than calcium acetate.
GTA is an acetate precursor, made of short-chain triglyceride with three acetate moieties on a glycerol backbone. Intragastric administration of GTA to tremor mice results in greatly increased brain acetate levels, and improves motor functions, without any new overt pathology in the mice. GTA given to infants with CD at low dose (up to 0.25 g/kg/d) resulted in no improvement in their clinical status, but also no detectable toxicity. At higher doses (4.5 g/kg/d), the treatment resultes well tolerated. The lack of clinical improvement might be explained mainly by the late onset of treatment, when significant brain damage is already present. Further larger studies of CD patients below age 3 months are required in order to test the long-term efficacy of this drug.

A safety trial of high dose glyceryl triacetate for Canavan disease. 2011

Progress toward acetate supplementation therapy for Canavan disease: glyceryl triacetate administration increases acetate, but not N-acetylaspartate, levels in brain. 2005

2. Lithium citrate has also been added for clinical application in human subjects with CD. The clinical significance of use of lithium citrate is not known. After one year of treatment, NAA levels decrease by approximately 20% in the brain region, urinary NAA levels show a reduction of 80% and measurements recorded on MRI studies suggeste a mild improvement in myelination in the frontal white matter. Patient improve alertness and visual tracking but continue with no heat support, axial hypotonia, and spastic diplegia. Given the absence of adverse effects and limited treatment options, lithium citrate may be a good alternative to stop the progression of the disease and improve the quality of life of patients.

Lithium citrate as treatment of Canavan disease. 2012

Lithium citrate reduces excessive intra-cerebral N-acetyl aspartate in Canavan disease. 2010

3. Gene transfer therapy is used to replace the mutated ASPA gene into the brain.

The very first gene therapy approved used recombinant ASPA cDNA in adeno-associated viral vector (AAV) ASPA for liposome-based delivery of ASPA gene in somatic cells and was administered to two affected children [Leone et al.,2000 ],and subsequently delivered to a cohort of 21 CD children [Janson et al.,2001 ]. Pre- and post-delivery assessments included a battery of noninvasive biochemical, radiological, and neurological tests. For better effciency, different AAV-serotype has been tested. In all subjects, treated with Adeno-associated virus serotype 2 (AAV2), there aren’t evidence of AVV2 neutralizing antibodies and there are minimal systemic signs of inflammation or immune stimulation. rAAV2 vector administration to the human CNS appears well tolerated, but no children improved.

Clinical protocol. Gene therapy of Canavan disease: AAV-2 vector for neurosurgical delivery of aspartoacylase gene (ASPA) to the humanbrain. 2000

Immune responses to AAV in a phase I study for Canavan disease. 2006

Effects of AAV-2-mediated aspartoacylase gene transfer in the tremor rat model of Canavan disease.

It has also been transferred ASPA gene to the brains of two children with Canavan disease using a non-viral vector. It was developed a polycation-condensed delivery system (LPD) linked to an adeno-associated virus (AAV)-based plasmids containing recombinant aspartoacylase (ASPA). The gene delivery system was tested in healthy rodents and primates, before proceeding to preliminary studies in 2 children with Canavan disease. LPD/pAAVaspa is well tolerated in human subjects and is associated with biochemical, radiological and clinical changes. However the children continued to have untreated Canavan disease

Aspartoacylase gene transfer to the mammalian central nervous system with therapeutic implications for Canavan disease. 2000

Sonia Costa e Iris Chiara Salaroglio

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