A Next-Generation Sequencing of the NOTCH3 and HTRA1 Genes in CADASIL Patients

Fernández, Ángela; Gómez, Juan; Alonso, Belén; Iglesias, Sara; Coto, Eliécer

doi:10.1007/s12031-015-0560-3

Cited by 3 publications

(3 citation statements)

References 16 publications

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“…We followed a next‐generation sequencing (NGS) procedure previously used in our laboratory for other complex Mendelian disorders . Briefly, we created DNA pools containing DNA from different patients, and the pools were polymerase chain reaction (PCR) amplified in two tubes with an Ampliseq designed to cover the coding sequences plus at least five intronic flanking nucleotides of the SPG7 gene.…”

Section: Methodsmentioning

confidence: 99%

“…We followed a next-generation sequencing (NGS) procedure previously used in our laboratory for other complex Mendelian disorders. 21,22 Briefly, we created DNA pools containing DNA from different patients, and the pools were polymerase chain reaction (PCR) amplified in two tubes with an Ampliseq designed to cover the coding sequences plus at least five intronic flanking nucleotides of the SPG7gene. Each DNA pool was amplified with the Ion Ampliseq Library Kit in conjunction with Ion Ampliseq Custom Primer Pool protocols, according to the manufacturer procedures (Life Technologies, Carlsbad, CA).…”

Section: Next-generation Sequencing Of the Spg7 Genementioning

confidence: 99%

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Parkinsonism and spastic paraplegia type 7: Expanding the spectrum of mitochondrial Parkinsonism

et al. 2019

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Background Pathogenic variants in the spastic paraplegia type 7 gene cause a complicated hereditary spastic paraplegia phenotype associated with classical features of mitochondrial diseases, including ataxia, progressive external ophthalmoplegia, and deletions of mitochondrial DNA. Objectives To better characterize spastic paraplegia type 7 disease with a clinical, genetic, and functional analysis of a Spanish cohort of spastic paraplegia type 7 patients. Methods Genetic analysis was performed in patients suspecting hereditary spastic paraplegia and in 1 patient with parkinsonism and Pisa syndrome, through next‐generation sequencing, whole‐exome sequencing, targeted Sanger sequencing, and multiplex ligation‐dependent probe analysis, and blood mitochondrial DNA levels determined by quantitative polymerase chain reaction. Results Thirty‐five patients were found to carry homozygous or compound heterozygous pathogenic variants in the spastic paraplegia type 7 gene. Mean age at onset was 40 years (range, 12–63); 63% of spastic paraplegia type 7 patients were male, and three‐quarters of all patients had at least one allele with the c.1529C>T (p.Ala510Val) mutation. Eighty percent of the cohort showed a complicated phenotype, combining ataxia and progressive external ophthalmoplegia (65% and 26%, respectively). Parkinsonism was observed in 21% of cases. Analysis of blood mitochondrial DNA indicated that both patients and carriers of spastic paraplegia type 7 pathogenic variants had markedly lower levels of mitochondrial DNA than control subjects (228 per haploid nuclear DNA vs. 176 vs. 573, respectively; P < 0.001). Conclusions Parkinsonism is a frequent finding in spastic paraplegia type 7 patients. Spastic paraplegia type 7 pathogenic variants impair mitochondrial DNA homeostasis irrespective of the number of mutant alleles, type of variant, and patient or carrier status. Thus, spastic paraplegia type 7 supports mitochondrial DNA maintenance, and variants in the gene may cause parkinsonism owing to mitochondrial DNA abnormalities. Moreover, mitochondrial DNA blood analysis could be a useful biomarker to detect at risk families. © 2019 International Parkinson and Movement Disorder Society

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Section: Methodsmentioning

confidence: 99%

Section: Next-generation Sequencing Of the Spg7 Genementioning

confidence: 99%

Parkinsonism and spastic paraplegia type 7: Expanding the spectrum of mitochondrial Parkinsonism

et al. 2019

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“…In addition, three missense mutations previously reported as pathogenic (C183R [8], R332C [9, 10] and Y465C [11]) but not previously identified in our diagnostics cohort were also detected [17]. In total, six typical CADASIL mutations involving cysteine alterations were identified in seven patients (15.9 %) out of 44 subjects, a detection rate higher than previously reported by Fernandez et al and Bianchi et al [18, 19]. …”

Section: Discussionmentioning

confidence: 47%

Targeted next generation sequencing identifies novel NOTCH3 gene mutations in CADASIL diagnostics patients

et al. 2016

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BackgroundCerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is a monogenic, hereditary, small vessel disease of the brain causing stroke and vascular dementia in adults. CADASIL has previously been shown to be caused by varying mutations in the NOTCH3 gene. The disorder is often misdiagnosed due to its significant clinical heterogeneic manifestation with familial hemiplegic migraine and several ataxia disorders as well as the location of the currently identified causative mutations. The aim of this study was to develop a new, comprehensive and efficient single assay strategy for complete molecular diagnosis of NOTCH3 mutations through the use of a custom next-generation sequencing (NGS) panel for improved routine clinical molecular diagnostic testing.ResultsOur custom NGS panel identified nine genetic variants in NOTCH3 (p.D139V, p.C183R, p.R332C, p.Y465C, p.C597W, p.R607H, p.E813E, p.C977G and p.Y1106C). Six mutations were stereotypical CADASIL mutations leading to an odd number of cysteine residues in one of the 34 NOTCH3 gene epidermal growth factor (EGF)-like repeats, including three new typical cysteine mutations identified in exon 11 (p.C597W; c.1791C>G); exon 18 (p.C977G; c.2929T>G) and exon 20 (p.Y1106C; c.3317A>G). Interestingly, a novel missense mutation in the CACNA1A gene was also identified in one CADASIL patient. All variants identified (novel and known) were further investigated using in silico bioinformatic analyses and confirmed through Sanger sequencing.ConclusionsNGS provides an improved and effective methodology for the diagnosis of CADASIL. The NGS approach reduced time and cost for comprehensive genetic diagnosis, placing genetic diagnostic testing within reach of more patients.Electronic supplementary materialThe online version of this article (doi:10.1186/s40246-016-0093-z) contains supplementary material, which is available to authorized users.

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NOTCH3 mutations in a cohort of Portuguese patients within CADASIL spectrum phenotype

et al. 2021

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A Next-Generation Sequencing of the NOTCH3 and HTRA1 Genes in CADASIL Patients

Cited by 3 publications

References 16 publications

Parkinsonism and spastic paraplegia type 7: Expanding the spectrum of mitochondrial Parkinsonism

Parkinsonism and spastic paraplegia type 7: Expanding the spectrum of mitochondrial Parkinsonism

Targeted next generation sequencing identifies novel NOTCH3 gene mutations in CADASIL diagnostics patients

NOTCH3 mutations in a cohort of Portuguese patients within CADASIL spectrum phenotype

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