Individual exome analysis in diagnosis and management of paediatric liver failure of indeterminate aetiology

Abstract

Background & aims: In children with liver failure, as many as half remain of indeterminate aetiology. This hinders timely consideration of optimal treatment options. We posit that a significant subset of these children harbour known inherited metabolic liver diseases with atypical presentation or novel inborn errors of metabolism. We investigated the utility of whole-exome sequencing in three children with advanced liver disease of indeterminate aetiology.

Methods: Patient 1 was a 10 year-old female diagnosed with Wilson disease but no detectable ATP7B mutations, and decompensated liver cirrhosis who underwent liver transplant and subsequently developed onset of neurodegenerative disease. Patient 2 was a full-term 2 day-old male with fatal acute liver failure of indeterminate aetiology. Patient 3 was an 8 year-old female with progressive syndromic cholestasis of unknown aetiology since age 3 months.

Results: Unbiased whole-exome sequencing of germline DNA revealed homozygous mutations in MPV17 and SERAC1 as the disease causing genes in patient 1 and 2, respectively. This is the first demonstration of SERAC1 loss-of-function associated fatal acute liver failure. Patient 1 expands the phenotypic spectrum of the MPV17-related hepatocerebral mitochondrial DNA depletion syndrome. Patient 3 was found to have syndromic cholestasis due to bi-allelic NOTCH2 mutations.

Conclusions: Our findings validate the application of whole-exome sequencing in the diagnosis and management of children with advanced liver disease of indeterminate aetiology, with the potential to enhance optimal selection of treatment options and adequate counselling of families. Moreover, whole-exome sequencing revealed a hitherto unrecognized phenotypic spectrum of inherited metabolic liver diseases.

Keywords: Genetic diagnosis; Germline mutations; Inherited metabolic liver diseases; Liver failure of indeterminate aetiology; Whole-exome sequencing.

Figure 1

Liver histological, radiological and genetic findings in patient 1. (A and B) Liver explant with macroscopic nodular cirrhotic appearance at low and high (20x) power, respectively. (C) Well-differentiated hepatocellular carcinoma (100x). (D) Significant copper deposition in many of the hepatocytic nodules, predominantly in the periseptal hepatocytes (400x). (E) Pleomorphic mitochondria with dilated cristae with occasional granular-dense deposits and crystalloid inclusions. (F) Brain magnetic resonance imaging shows bilateral basal ganglia masses with surrounding edema as depicted by asterisks. (G and H) Representative images of brain magnetic resonance showing white matter abnormalities in the brain stem. (I) Missense homozygous mutation in MPV17 (R50W) detected by massively parallel sequencing. (J) Sanger sequence traces of codons 49–51 are shown. Patient 1 is homozygous for a single base change (CGG>TGG, in red) in MPV17, resulting in a R50W (c.148T>C) missense mutation. Proband's parents are heterozygous for the same mutation. (K) Conservation of R50 across species. The amino acid sequence of 42–58 of human MPV70 is shown and compared to the corresponding sequences of nine vertebrate and invertebrate orthologs and paralogs. Amino acid positions identical to human genome are highlighted in yellow.

Figure 2

SERAC1 homozygous splice site mutation detected in patient 2 using WES. Sanger sequence trace of 5' donor splice site of codon 13 is shown. Patient 2 is homozygous for a base change (G>C, red arrow) in SERAC1, resulting in homozygous splice site mutation: IVS13+1G>C (c.1403+1G>C).

Figure 3

Liver histological findings and NOTCH2 compound heterozygous mutations detected in patient 3. (A) Low magnification showing preserved lobular architecture of the liver parenchyma and lack of significant fibrosis (Trichrome stain, 40x). The portal tracts (arrows) seen at this magnification fail to show either significant inflammation or any obvious pathology. (B) Higher magnification (400x) of the portal tract to show absence of the bile duct and also any lack of ductular reaction. Occasional lymphocytes or histocytes can be seen in the portal tract. Few glycogenated hepatocytic nuclei can be seen in the periportal region (H&E stain). (C) Immunostain for CK7 highlights the absence of the native bile duct and shows intense staining of the periportal hepatocytes (200x). (D) Sanger sequence trace of codons 1720–1722 are shown. Patient 3 is heterozygous for a base change (TCA>TTA, in red) in NOTCH2, resulting in a S1741L (c. 5222C>T) missense heterozygous mutation. Proband's father is heterozygous for the same mutation whereas proband's mother is wild-type (WT). (E) Conservation of S1741 across species. The amino acid sequence of 1733–1749 of human NOTCH2 is shown and compared to the corresponding sequences of eight vertebrate orthologs and paralogs. Amino acid positions identical to human genome are highlighted in yellow. (F) Sanger sequence trace of codons 1881–1883 are shown. Patient 3 is heterozygous for a base change (CAC>TAC, in red) in NOTCH2, resulting in a H1882Y (c.5644C>T) missense heterozygous mutation, Proband's mother is heterozygous for the same mutation whereas proband's father is wild-type (WT). (G) Conservation of H1882Y across species. The amino acid sequence of 1874–1890 of human NOTCH2 is shown and compared to the corresponding sequences of eight vertebrate orthologs and paralogs. Amino acid positions identical to human genome are highlighted in yellow. H1882 is conserved among all species examined.