Targeted-Capture Next-Generation Sequencing in Diagnosis Approach of Pediatric Cholestasis

Abstract

Background: Cholestasis is a frequent and severe condition during childhood. Genetic cholestatic diseases represent up to 25% of pediatric cholestasis. Molecular analysis by targeted-capture next generation sequencing (NGS) has recently emerged as an efficient diagnostic tool. The objective of this study is to evaluate the use of NGS in children with cholestasis. Methods: Children presenting cholestasis were included between 2015 and 2020. Molecular sequencing was performed by targeted capture of a panel of 34 genes involved in cholestasis and jaundice. Patients were classified into three categories: certain diagnosis; suggested diagnosis (when genotype was consistent with phenotype for conditions without any available OMIM or ORPHANET-number); uncertain diagnosis (when clinical and para-clinical findings were not consistent enough with molecular findings). Results: A certain diagnosis was established in 169 patients among the 602 included (28.1%). Molecular studies led to a suggested diagnosis in 40 patients (6.6%) and to an uncertain diagnosis in 21 patients (3.5%). In 372 children (61.7%), no molecular defect was identified. Conclusions: NGS is a useful diagnostic tool in pediatric cholestasis, providing a certain diagnosis in 28.1% of the patients included in this study. In the remaining patients, especially those with variants of uncertain significance, the imputability of the variants requires further investigations.

Opinion paper on the diagnosis and treatment of progressive familial intrahepatic cholestasis

“…Diagnosis of PFIC has traditionally been made via clinical assessment (including symptoms, family history, laboratory investigation, liver ultrasound and liver biopsy); however, gene sequencing is now widely accepted as the gold standard for diagnostic confirmation. 15 In addition to direct gene analysis via Sanger sequencing, three methods of next-generation sequencing have been reported in the literature for PFIC diagnosis. 15 , 16 , 17 , 18 Targeted gene panel analysis involves sequencing several selected cholestasis-associated genes in a single analysis, and is frequently used alongside clinical diagnosis when a specific PFIC genotype is suspected.…”

“… 15 In addition to direct gene analysis via Sanger sequencing, three methods of next-generation sequencing have been reported in the literature for PFIC diagnosis. 15 , 16 , 17 , 18 Targeted gene panel analysis involves sequencing several selected cholestasis-associated genes in a single analysis, and is frequently used alongside clinical diagnosis when a specific PFIC genotype is suspected. 15 , 18 This method of next-generation sequencing offers a more time efficient and less costly method of genetic diagnosis compared with other next-generation sequencing techniques.…”

“… 15 , 16 , 17 , 18 Targeted gene panel analysis involves sequencing several selected cholestasis-associated genes in a single analysis, and is frequently used alongside clinical diagnosis when a specific PFIC genotype is suspected. 15 , 18 This method of next-generation sequencing offers a more time efficient and less costly method of genetic diagnosis compared with other next-generation sequencing techniques. 15 The diagnostic yield of targeted gene panel sequencing varies widely in the published literature, from 28.1% to 68%, which is thought to be due to patient selection criteria and the number of genes utilised in the panel.…”

Alagille syndrome: an orphan disease in Colombia and summary of recent advances in treatment and survival – a case report

“…Another significant advance, but in the diagnostic field, is the use of molecular analysis by next-generation sequencing for targeted capture in cases of pediatric cholestasis. Although at present, it only performs the diagnosis in 30% of the cases, since there are cases where no molecular defects are identified 15 . Other work is currently focused on defining pathogenicity variants of NOTCH2 , which are less frequent 16 ; in identifying regulators of JAG1 expression in portal mesenchymal cells 17 , and following up on the results obtained from different treatments 18 .…”

Failure of cholic acid therapy in a child with a bile acid synthesis defect and harboring plectin mutations

“…3 Considering this unexpected cholestatic evolution, additional next-generation sequencing molecular study identified two novel mutations in PLEC (NM_201384; c.3434C>T; p.Thr1145Met; ACMG classification: class 3; gnomAD: 0.032%; CADD phred score: 15.1 and c.7192C>T; p.Arg2398Cys; ACMG classification: class 3; gnomAD: 0.017%; CADD phred score: 25.7), encoding plectin. 4,5 Consanguineous parents were heterozygous for the mutations identified in both genes. PLEC mutations localized in the plakin and coiled coil rod plectin domains were similar to the PLEC mutations already reported in patients with progressive familial intrahepatic cholestasis.…”