Improving the diagnosis of cobalamin and related defects by genomic analysis, plus functional and structural assessment of novel variants

Abstract

BackgroundCellular 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.ResultsThis 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.ConclusionsThe 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.Electronic supplementary materialThe online version of this article (10.1186/s13023-018-0862-y) contains supplementary material, which is available to authorized users.

A Review of Disparities and Unmet Newborn Screening Needs over 33 Years in a Cohort of Mexican Patients with Inborn Errors of Intermediary Metabolism

“…A systematized public health program and budget for sequencing large numbers of patients would expedite IEiM diagnoses, especially for those associated with more than one gene (i.e., PA, MSUD, or MMA) or without specific biomarkers for a biochemical diagnosis such as glycogen storage disease (GSD). As other authors have demonstrated, it is necessary to improve access to WES or WGS in this group of patients [ 77 , 78 , 79 ].…”

Combined Malonic and Methylmalonic Aciduria Due to ACSF3 Variants Results in Benign Clinical Course in Three Chinese Patients

“…The locations of these variants in ACSF3 are graphically displayed in Figure 5. Most of the variants are missense, mainly located at the carboxyl terminal of the protein (Table 1) (2,(4)(5)(6)(7)(8)(9). The most common ACSF3 variants reported so far are c.1075G>A (p.E359K) and c.1672C>T (p.R558W), which are also the most common variants in asymptomatic patients.…”

“…Signs and symptoms reported so far are involved in neurological abnormalities in adults and infection induced encephalopathy in pediatric patients. Nevertheless, asymptomatic patients with normal outcomes strongly suggest that CMAMMA can be a benign state (2,(4)(5)(6)(7)(8)(9). No case of the disease has been reported in China so far.…”