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oai:repisalud.isciii.es:20.500.12105/176152024-09-21T23:36:08Zcom_20.500.12105_15322com_20.500.12105_2051col_20.500.12105_16927col_20.500.12105_16963col_20.500.12105_16987

00925njm 22002777a 4500 dc Brasil, Sandra author Leal, Fátima author Vega, Ana author Navarrete, Rosa author Ecay, María Jesús author Desviat, Lourdes R author Riera, Casandra author Padilla, Natàlia author de la Cruz, Xavier author Couce, Mari Luz author Martin-Hernández, Elena author Morais, Ana author Pedrón, Consuelo author Peña-Quintana, Luis author Rigoldi, Miriam author Specola, Norma author de Almeida, Isabel Tavares author Vives, Inmaculada author Yahyaoui, Raquel author Rodríguez-Pombo, Pilar author Ugarte, Magdalena author Pérez-Cerda, Celia author Merinero, Begoña author Pérez, Belén author 2018-07-24 Cellular cobalamin defects are a locus and allelic heterogeneous disorder. The gold standard for coming to genetic diagnoses of cobalamin defects has for some time been gene-by-gene Sanger sequencing of individual DNA fragments. Enzymatic and cellular methods are employed before such sequencing to help in the selection of the gene defects to be sought, but this is time-consuming and laborious. Furthermore some cases remain undiagnosed because no biochemical methods have been available to test for cobalamin absorption and transport defects. This paper reports the use of massive parallel sequencing of DNA (exome analysis) for the accurate and rapid genetic diagnosis of cobalamin-related defects in a cohort of affected patients. The method was first validated in an initial cohort with different cobalamin defects. Mendelian segregation, the frequency of mutations, and the comprehensive structural and functional analysis of gene variants, identified disease-causing mutations in 12 genes involved in the absorption and synthesis of active cofactors of vitamin B12 (22 cases), and in the non-cobalamin metabolism-related genes ACSF3 (in four biochemically misdiagnosed patients) and SUCLA2 (in one patient with an unusual presentation). We have identified thirteen new variants all classified as pathogenic according to the ACGM recommendation but four were classified as variant likely pathogenic in MUT and SUCLA2. Functional and structural analysis provided evidences to classify them as pathogenic variants. The present findings suggest that the technology used is sufficiently sensitive and specific, and the results it provides sufficiently reproducible, to recommend its use as a second-tier test after the biochemical detection of cobalamin disorder markers in the first days of life. However, for accurate diagnoses to be made, biochemical and functional tests that allow comprehensive clinical phenotyping are also needed. 10.1186/s13023-018-0862-y 1750-1172 Orphanet journal of rare diseases http://hdl.handle.net/10668/12749 30041674 http://hdl.handle.net/20.500.12105/17615 Cobalamin disorders Homocystinuria Massive parallel sequencing Methylmalonic aciduria Improving the diagnosis of cobalamin and related defects by genomic analysis, plus functional and structural assessment of novel variants.