Abstracts for the 34th HGSA Annual Scientific Meeting, Melbourne, Australia November 16-18, 2010

2010 Twin Research and Human Genetics  
X-linked adrenoleukodystrophy (X-ALD) is characterized by two main phenotypes. Childhood cerebral ALD with onset of cerebral demyelination between 5-12 years results in a devastating neurologic degradation, and often death before adolescence. Adrenomyeloneuropathy (AMN) that affects adults males and heterozygous women is characterized by a pure myelopathy. However, about 35% of AMN males develop also cerebral demyelination with the same poor prognosis as in boys. The X-ALD (ABCD1) gene codes
more » ... a peroxisomal transporter (ALD protein) are involved in the metabolism of verylong-chain fatty acids. Cerebral demyelination of X-ALD can be arrested in children and adults by allogeneic HSC transplantation (HCT) within 12-18 months, provided the procedure is performed at an early stage of the disease. The long-term beneficial effects of HCT in X-ALD are likely due to the progressive turn-over of brain microglia that are derived from myeloid progenitors in the bonemarrow. Despite the increased availability of cord blood, not all boys or adults with cerebral and who are candidate for HCT have a suitable HLA-matched donor. In addition, allogeneic HCT remains associated with significant mortality risk, particularly in adults. In two boys with cerebral ALD but without HLA-matched donor, we recently reported that HSC gene therapy with lentiviral vector has comparable effects to those seen after allogeneic HCT in arresting cerebral demyelination. Data on ALD protein expression as well as the identification of identical lentiviral insertion sites in myeloid and lymphoid lineages strongly suggested that HSCs were transduced in the patients. Hematopoiesis remained polyclonal without evidence of clonal skewing or dominance. Data will be presented on longer follow-up of these two treated patients, as well as on a third patient who has been treated more recently by HSC gene therapy. Technologies developed in the research laboratory, especially those for analysis of many genes at a time, are being adapted to the clinical laboratory setting, and bringing with them issues related to sample preparation, turn-around time, sensitivity, data acquisition and interpretation, governmental regulation and oversight, and ethical complications. Abstracts for the 34th HGSA Annual Scientific Meeting, The neuronal ceroid-lipofuscinoses (NCLs) are a group of severe recessively inherited neurodegenerative disorders that are characterized by retinal pigmentary changes and vision loss, seizures, loss of motor and cognitive function and early death. The pathogenesis of neuronal loss has not been established. My laboratory has been defining the pathophysiology of Juvenile NCL (CLN3, Batten's disease). Many hypotheses have been suggested regarding the function of the CLN3 protein (Battenin) including neuronal anti-apoptosis via the ceramide pathway, synaptic neurotransmission, vacuolar pH regulation, an autoimmune reaction to neuronal proteins, and endosomal/lysosomal trafficking. We have shown Battenin to function in a novel palmitoylprotein delta-9 desaturase pathway. We have developed a siRNA knock down model in cultured neuroblastoma cells to study pathways that are perturbed in Battenin deficiency. This model was shown to have increased intracellular calcium which we hypothesized would be a trigger for neuronal apoptosis. We have studied the effects of a panel of 42 calcium channel modulators on this abnormal calcium content to determine if any of these drugs can modulate the effect and thus be candidates for therapeutic intervention. 7 were shown to normalize the abnormal calcium content and some of these drugs are available for different indications. Although the pathogenesis of CLN3 remains incomplete, it is now clear that at least part of the neuronal cell death is triggered by increased intracellular calcium content and the apoptosis pathway. This pathway can be modulated in vitro by the use of calcium channel antagonists that are already on the market for other indications and for which safety profiles are established. At the moment, it is not known if data from CLN3 studies can be applied to the other NCL's or other causes of neuronal death but if a common pathway is established then this may open up a significant therapeutic potential. Introduction: The congenital disorders of protein glycosylation are a family of enzyme defects in assembly (Type I), transfer, and remodeling (Type II) of glycan structures to proteins. The features have wide variation and, while certain common presentations are known, the correlation between genotype and phenotype is not strong. Screening for these disorders in any unexplained syndrome is recommended. In protein glycosylation, the glycan structure is assembled in the endoplasmic reticulum and transferred for attachment to either an Asparagine residue with an N-linkage or to Serine/Threonine residues with an O-linkage. The current method of screening for these disorders is IEF, which discriminates charge differences in Transferrin or Apolipoprotein C-III due to failure to attach sialic acid residues to an incomplete glycan. Charge based screens cannot differentiate individual enzyme defects, gives false positive results with protein variants, and reportedly cannot identify 11/44 known disorders. We report the development and evaluation of MALDO-TOF/TOF methods for the characterisation of both N and O-glycan disorders. Methods: Briefly, the Apo C-III method uses acetonitrile for removal of albumin followed by TFA acidification and then MALDI ionisation on a DHB matrix. The N-glycan method uses affinity chromatography to isolate IgG and Transferrin, a yeast enolase internal standard, sialidase and PNGase-F to release free glycans which are extracted on a carbon column, and then MALDI ionisation on a DHB matrix. Results: The Apo C-III method discriminated and quantitated all forms of the mucin Type 1 O-glycan. Comparison against patient samples identified COG7, ATP6VOA2, and two novel disorders of ApoC-III -1 and Apo C-III -2. The N-glycan method can identify and quantitate 15 glycan structures, and has confirmed primary abnormalities of PMM2, MPI, COG7 and also Galactosemia. Normal structures were apparent in protein variant samples with abnormal IEF patterns. Conclusion: MALDI-TOF mass analysis of glycans provides a more informative investigation of possible CDG patients.
doi:10.1375/twin.13.6.620 fatcat:waz3wo2snrgdjigvmen5y2qtaq