Biopsy-based analysis of the prosteatotic TM6SF2 p.E167K and PNPLA3 p.I148 M gene variants as potential modifiers of Wilson disease

 

Objectives and study: Wilson disease (WD) is a rare liver disease caused by mutations of the ATP7B gene. The presence of genetic modifiers of WD has for long been suspected, however no common polymorphism affecting the progression of this disorder in pediatric patients has been detected so far. Previous studies demonstrated that the prosteatotic gene variants, PNPLA3 p.I148 M and TM6SF2 p.E167K are associated with increased liver injury in adults with chronic liver diseases. Here we investigate these variants as potential modifiers of liver injury in a large cohort of pediatric WD patients.

Methods: Overall, we recruited 79 children (age 9 ± 5 years, 10 pairs of sibs) with WD among whom 5 required liver transplantation. Genotyping of the TM6SF2 rs58542926 and PNPLA3 rs738409 polymorphisms was performed using TaqMan assays with fluorescence detection. Liver biopsy was performed in 59 children (75%). Steatosis (micro- and macrovesicular) and fibrosis were quantified by pathologists blinded to the genotyping results. All unrelated patients and randomly selected one sib from each pair were included in analyses: association tests were calculated in contingency tables, continuous variables were compared using Mann-Whitney or ANOVA tests.

Results: Among the biopsied patients, 57% displayed steatosis grade 1 and 20% presented with steatosis grade 2. Fibrosis stages 2 – 4 were present in 74% of patients, whereas only 5% showed no fibrosis at liver biopsy. The genotype frequencies of the TM6SF2 p.E167K and PNPLA3 p.I148 M polymorphisms did neither differ from Caucasian population frequencies nor deviate from HWE (P > 0.05). In total, 50% of patients carried at least one copy of the TM6SF2 or PNPLA3 risk allele. The presence of the minor TM6SF2 allele was associated with an increased risk of developing fibrosis stage ≥2 (OR = 9.36, 95% CI 0.51 – 170.92, P = 0.04): All carriers of the TM6SF2 risk allele displayed fibrosis stages ≥2 at biopsy. This variant was however not associated with either liver steatosis or with liver function tests (both P > 0.05). Although the PNPLA3 minor allele was associated with none of the histologic traits (P > 0.05), its carriers had significantly (P = 0.01) increased INR possibly reflecting enhanced liver injury. In total, 4 out 5 (80%) transplanted children carried at least one copy of the PNPLA3 or TM6SF2 risk alleles.

Conclusions: Our results suggest that the TM6SF2 p.E167K, and to a lesser extent, PNPLA3 p.I148 M polymorphisms might modulate liver injury in children with WD. Since fibrosis seems to be frequent in pediatric WD, testing of the TM6SF2 variant could assist in detecting patients with an increased risk of disease progression.