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Adenylosuccinate lyase deficiency- a cause of seizures in children: Development of an assay for its diagnosis

Alsanousi, A; (2008) Adenylosuccinate lyase deficiency- a cause of seizures in children: Development of an assay for its diagnosis. Doctoral thesis , UCL (University College London). Green open access


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Adenylosuccinate Lyase (ADSL) Deficiency is one of the rare causes of neonatal and infantile seizures. This condition results from autosomal recessive mutations in the gene coding for the enzyme ADSL, mapped to human chromosome 22ql3.1-13.2. The ADSL enzyme plays an important role in the de novo pathways of purine metabolism and thus deficiency, and/ or decreased catalytic activity of this enzyme is manifested in form of repeated seizures in early childhood. Other neurological symptoms and signs may be present, such as psychomotor retardation, autistic features, axial and peripheral hypotonia, and muscle atrophy and secondary feeding problems leading to severe growth retardation. Although ADSL deficiency is considered a rare cause of childhood epilepsy and only 50 cases have been reported till now world wide, the magnitude of the problem can not be determined unless rapid accurate diagnostic test is introduced, especially, in communities with high rate of consanguinity. Fortunately, several reports have given a hint for Succinyladenosine or S-Ado as a new promising diagnostic marker for this disorder. This information may have enormous impact in the era of enzyme replacement therapy and thus this condition could be to some extent preventable, once proper genetic counseling and testing for carriers of the mutations in families with this condition is possible. In the present study we aimed to develop and validate a new HPLC method for the detection of S-Ado in the CSF. Material and Methods HPLC system with anion exchange Sphere Clone 5u SAX (250 x 4.6) has been developed using synthetic S-Ado in different concentrations to obtain the linear calibration curve. A total of 23 CSF samples were obtained from 23 patients, 12 females (52.2%), 11 male (47.8%), age ranging between 1-16 years, are processed for measurement of S-Ado using the currently developed HPLC system. All patients were referred to the Neurometabolic unit at the National Hospital for Neurology and Neurosurgery during the year 2005 with seizures except 2 samples form known ADSL deficiency male patients received as gift from Dr. Keith Hyland (Horizon Molecular Medicine, Atlanta, GA, USA). We considered the presence of RBCs or hemolysis in the CSF as exclusion criteria. Outcome and Results A rapid, accurate, in addition sensitive and specific HPLC system with anion exchange SAX column, to detect S-Ado in CSF of patients with ADSL deficiency has been successfully developed and validated. The total time for the test < 15 minutes, and the method validation was achieved by measuring the S-Ado concentration in the CSF of 21 unknown patients presented to the Neurometabolic unit, the observed S-Ado reference range concentration for these cases was found to be varying between (0.55 to 2.36uM). None of the randomly selected disease control cases showed an abnormal S-Ado concentration, whereas the two CSF samples for known cases of ADSL deficiency have shown very high concentrations (356.8 and 357.1 iM) of S-Ado compared with non-ADSL patients. Recovery of S-Ado from spiked CSF after addition of a known concentration (10M) was achieved by with very high accuracy (range 92%-100%)i.e. greater than 98% in average .The physical properties of S-Ado were found to be stable substance in different and wide range of store temperature i.e. in (-80 C) and also when stored at room temperature (20.8 C), for more than 24 hours .Thus an age related reference range was established for S-Ado in the CSF and the statistical analysis showed no significant cumulative effect of the age or the sex on this reference range. The study was completed within total of 3 month duration, and mean intraday variation was 5% and the interday variation was < 10%. Conclusion: the current method is sensitive specific and enables rapid reliable detection of S-Ado in the CSF of patient with ADSL deficiency in < 15 min

Type: Thesis (Doctoral)
Title: Adenylosuccinate lyase deficiency- a cause of seizures in children: Development of an assay for its diagnosis
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Thesis digitised by ProQuest.
UCL classification: UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Brain Sciences > UCL Queen Square Institute of Neurology
URI: https://discovery.ucl.ac.uk/id/eprint/1566961
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