X-linked adrenoleukodystrophy (ald-X): functional characterization of new changes in abcd1 gene in argentine patients

AMOROSI C; DVORÁKOVÁ L; MYSKOVA H; MONTI MR; KEMP S; CISMONDI IA; GUELBERT N; DODELSON DE KREMER R; OLLER DE RAMÍREZ A.

Cusco

Congreso; VIII Congreso latinoamericano de errores innatos del metabolismo y pesquisa neonatal; 2011

Society of Inherited Metabolic Disease.

Background: X-ALD is a neurodegenerative disorder characterized by an increase in serum of very long-chain fatty acids. Over 1000 mutations have been reported in ABCD1 gene. Objectives: Analyze genomic changes in the ABCD1 gene in 10 unrelated probands. Validate new missense and intronic changes by different methods: a) bioinformatics tools b) functional analysis. Methods: Bioinformatics tools: (PolyPhen http://genetics.bwh.harvard.edu/pph, SIFThttp://blocks.fhcrc.org/sift/SIFT.html,ME(http://gener.mit.edu/bungelab/maxent/Xmaxentscan_scoreseg_acc.html) and NN (http://www.fruitfly.org/seq_tools/splice.html) Functional analysis: It was accomplished PCR/sequencing of all exons, directed mutagenesis, protein analysis by western blot and b-oxidation. Results: We identified 7 novel and 1 known changes allowing the molecular characterization of 9/10 patients. New mutations identified: 3 frameshift [an insertion (p.Ser284fs), a deletion (p.Thr254ArgfsX82) and a duplication (p.Glu380ArgfsX21)]; a deletion that causes a loss of 4 amino acids (p.Ser572_Asp575del), a splicing substitution (c.1081+5G>C) and 2 missense changes.NN, ME, SIFT and PolyPhen programs to c.1081+5G>C, p.Tyr547Cys and p.Ala341Asp predict they could be mutations.In one patient, 2 missense changes were detected, p.Ala19Ser in exon 1 and p.His669Arg in exon 10. Studies in a heterologous expression system confirmed that p.Ala19Ser change is a polymorphism, due to normal levels of protein and b-oxidation, while p.His669Arg is the causative mutation. The absence of protein and b-oxidation  confirmed that the change p.Tyr547Cys is a mutation. Conclusions: The precise definition of the genotype in patients with X-ALD is essential for carriers detection, proper genetic counseling and future therapies.