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A review of the Wilson disease service over the past 15 years

A review of the Wilson disease service over the past 15 years. Miranda Durkie Sheffield Diagnostic Genetics Service miranda.durkie@sch.nhs.uk. Introduction. Wilson disease (WD; OMIM#277900) is an autosomal recessive disorder of copper metabolism Hepatic and/or neurological presentation

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A review of the Wilson disease service over the past 15 years

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  1. A review of the Wilson disease service over the past 15 years Miranda Durkie Sheffield Diagnostic Genetics Service miranda.durkie@sch.nhs.uk

  2. Introduction • Wilson disease (WD; OMIM#277900) is an autosomal recessive disorder of copper metabolism • Hepatic and/or neurological presentation • Treatable if diagnosed early • Over 500 mutations reported in all 21 exons of the ATP7B gene (Cu transporting ATPase) • Significant numbers of reported cases where both mutations cannot be identified (10-30%) • Lots of postulation about second WD gene but no mutations identified in candidate genes so far

  3. WD service at SDGS • WD service available since 1995 (part research) • National (and international) referrals • Published results from SSCP & confirmation DNA sequencing of 52 UK patients in 1999 with detection rate of 70% • SSCP/seq of 3 hotspot exons identified 60% of mutations in British cohort • DNA sequencing replaced SSCP in 1999 and stage 1 screen increased to 7 exons to pick-up 80% mutations • Stage 2 screen = sequencing of all remaining 14 exons

  4. Service Evaluation Aims • Determine mutation spectrum by sequencing • Determine mutation detection frequency in British referrals • Determine if further testing for deletions/duplications and promoter variants is warranted • Assess utility of 2 stage screening approach

  5. Cohort 1 A D B C

  6. Cohort 2 Long time lag between start of project in 2004 and completion of MLPA & promoter work in 2009 (limited resources) Therefore decided to look at 2nd cohort of referrals received between November 2004 and April 2009 Only included cases where 2 mutations had been detected and/or full sequencing had been carried out

  7. Cohort 2 F E

  8. Interesting Cohort 1 caseD • One patient homozygous for common p.His1069Gln, c.3207C>A mutation • Routine family studies in 1999 showed that Dad was heterozygous for this mutation but Mum did not have it • Reported as a possible whole exon deletion or primer SNP. • Alternative primers showed no primer SNPs • In 2007 MLPA kit available for ATP7B (P098 MRC Holland) • MLPA showed no deletion in Mum or index case • Used Promega Powerplex kit to check for sample mix-up

  9. Powerplex results D13S317 Index case 189 Dad 189 Mum 177;193

  10. Interesting caseD cont. • Used fluorescent microsatellite markers along length of chromosome 13 • Found 7 markers between 13q11 and 13q14.2 show biparental inheritance • 6 markers between 13q14.2 to 13q34 (inc ATP7B at 13q14.3) show non-maternal inheritance & inheritance of single paternal allele • Paternal segmental UPD confirmed resulting in autozygosity for p.His1069Gln mutation • First reported case of UPD13 with WD

  11. Putative promoter mutation - Case E Sardinian mutation c.-441_-427del15 c.-442G>A +1 ATG start Cullen et al Clin Genet 2003

  12. Alibaba analysis c.-442G>A Wild-type c.-442G>A YY1 = transcriptional repressor

  13. TFsearch results c.-442G>A Wild-type c.-442G>A

  14. c.-442G>A further studies • Family studies showed that it was inherited from Mum concordant with familial segregation • Not found on 188 normal chromosomes • Putative new binding of YY1/NF-muE1/GATA affecting normal TF binding?? • Needs further work

  15. Interesting cases F • 3 individuals found with 3 putative mutations each • 2 mutations in cis are all different & all missense • 1 is predicted to affect splicing • 1 previously reported case with 3 mutations in literature from isolated mountainous region of Crete with very high WD incidence • Important implications for staged screening & presymptomatic testing

  16. Mutation distribution • Previous SSCP study found mutations in exons 2, 8, 13-15 & 18-19 most prevalent (80%) = Stage 1 screen • However new data shows exons 2, 5, 8, 13-14, 18-20 most prevalent Old stage 1 New stage 1

  17. Summary • 117 different mutations, 36 novel, found in 191 patients • No deletions/duplications detected therefore MLPA is not warranted for routine diagnosis • One promoter variant found therefore promoter analysis is not warranted for routine diagnosis • 1st case of UPD13 found in WD • 2 missense mutations can occur in cis • 60% of negative stage 1 screens said “WD not likely” therefore staged screening warranted but stage 1 exons need to be adjusted • Of 186 patients with confirmed diagnosis of WD the mutation detection frequency is 98% • Second WD locus is unlikely

  18. Thank you! Royal Hallamshire Hospital • Oliver Bandmann • Stefanie Klass Sheffield Children’s NHS Foundation Trust • Stuart Tanner Other • EuroWilson for supporting EMQN WD scheme Sheffield Diagnostic Genetics Service • James Blackburn • Ann Dalton • Anne Goodeve • Ann Lee • Maria Panayi • Everyone at SDGS past & present who worked on WD

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