Targeted Next Generation Sequencing in Patients with Inborn Errors of Metabolism

Abstract

BackgroundNext-generation sequencing (NGS) technology has allowed the promotion of genetic diagnosis and are becoming increasingly inexpensive and faster. To evaluate the utility of NGS in the clinical field, a targeted genetic panel approach was designed for the diagnosis of a set of inborn errors of metabolism (IEM). The final aim of the study was to compare the findings for the diagnostic yield of NGS in patients who presented with consistent clinical and biochemical suspicion of IEM with those obtained for patients who did not have specific biomarkers.MethodsThe subjects studied (n = 146) were classified into two categories: Group 1 (n = 81), which consisted of patients with clinical and biochemical suspicion of IEM, and Group 2 (n = 65), which consisted of IEM cases with clinical suspicion and unspecific biomarkers. A total of 171 genes were analyzed using a custom targeted panel of genes followed by Sanger validation.ResultsGenetic diagnosis was achieved in 50% of patients (73/146). In addition, the diagnostic yield obtained for Group 1 was 78% (63/81), and this rate decreased to 15.4% (10/65) in Group 2 (X2 = 76.171; p < 0.0001).ConclusionsA rapid and effective genetic diagnosis was achieved in our cohort, particularly the group that had both clinical and biochemical indications for the diagnosis.

Multiplex targeted high‐throughput sequencing in a series of 352 patients with congenital limb malformations

“…Overall, in this series of 352 patients, 138 pathogenic or likely pathogenic variants were identified, among which 26 CNVs, and 22 variants of uncertain significance. Our approach revealed to be a useful strategy, providing a molecular diagnosis in 35.2% of patients, similar to other heterogeneous conditions (50% for inborn errors of metabolism (Yubero et al, ), 47.3% for myopathies and muscular dystrophies (Stehlikova et al, ), 39% for intellectual disability (Martinez et al, ), 28.5% for epileptic encephalopathy (Kothur et al, ), 28.1% for disorders of sexual development (Fan et al, ), 24.5% for developmental eye disorders (Patel et al, ), 20% for primary arrhythmia syndrome and cardiomyopathy (Robyns et al, ), and 18% for inherited polyneuropathy (Wang et al, ). A 7.7% increase was observed in our diagnostic performance, compared to our previous workflow (diagnostic yield 27.5% before 2014, data not shown).…”

“…CLM are a diagnostic challenge due to their variable expressivity, clinical overlap between the different syndromes (e.g., radial anomalies) and their genetic heterogeneity. NGS approaches are increasingly being used for genetic diagnosis in routine clinical practice and many publications report successful use in heterogeneous disorders such as inborn errors of metabolism (Yubero et al, ), neuromuscular disorders (Stehlikova et al, ), and intellectual disability (Martinez et al, ). To our knowledge, no report of the diagnostic impact in patients affected with CLM using high‐throughput sequencing has been published so far.…”

Cerebrospinal fluid monoamines, pterins, and folate in patients with mitochondrial diseases: systematic review and hospital experience

“…The different mtDNA mutations were studied by specific procedures, including Sanger sequencing, Southern blot, and real-time PCR, as previously reported (O'Callaghan et al 2015;Montiel-Sosa et al 2013). Nuclear DNA was assessed either by Sanger sequencing, depending on the suspected causative gene, or, over the past 4 years, by nextgeneration sequencing (NGS) using customized (Yubero et al 2016) or commercial panels (TruSight One Sequencing Panel, Illumina) in MiSeq or NextSeq500 sequencers (Illumina). Progenitor studies were performed to evaluate the inheritance model and to establish the molecular diagnosis.…”

Aplicabilidade clínica da técnica de sequenciamento de nova geração com enfoque em displasias esqueléticas

“…Mas talvez a questão mais fundamental do teste clínico esteja no fato de o resultado não ser submetido a um conjunto de revisores especializados (como no caso de testes de pesquisa), embora tenha imediata relevância para a saúde do paciente. Shashi et al, 2014;Wang et al, 2014;Yang et al, 2014;Jamuar e Tan, 2015;Jamie et al, 2016;Retterer et al, 2016;Yubero et al, 2016;Celestino-Soper et al, 2017 …”

“…A sensibilidade diagnóstica de pouco mais de 60% obtida na casuística de doenças ósseas genéticas foi homogênea para os três métodos (Sanger, painel NGS e WES) no presente estudo e acima da média reportada na literatura, tanto para amostras de doenças ósseas (Polla et al, 2015;Bae et al, 2016;Retterer et al, 2016), como para amostras com outros tipos de doenças genéticas realizadas por NGS. Já a sensibilidade diagnóstica na casuística de RASopatias foi de 46% para WES, que é próxima à média da literatura Lepri et al, 2014;Wang et al, 2014;Yang et al, 2014;Taillandier et al, 2015;Kuhn et al, 2016;Jamie et al, 2016;Retterer et al, 2016;Yubero et al, 2016;Celestino-Soper et al, 2017;Bhoj et al, 2017) (Naslavsky, Yamamoto et al, 2017). No caso das doenças ósseas genéticas, foi criado recentemente um banco de dados chamado 'SkeletonGenetics', para armazenar as variantes já descritas associadas a doenças ósseas genéticas …”