Development of a UK diagnostic service for Meckel-Gruber syndrome

1. Development of a UK diagnostic service for Meckel-Gruber syndrome Helen Lindsay, Kimberley Flintoff, David Cockburn, Ruth Charlton and Colin Johnson Yorkshire Regional DNA Laboratory

2. Overview • Meckel-Gruber syndrome (MKS) • Clinical features • Genetic aspects • MKS mutation spectrum • Screening test strategy • Achievements to date

3. Meckel-Gruber syndrome (MKS) Clinical features • Lethal developmental disorder • UK incidence approximately 1 in 30,000 • ‘Classic triad’ of features (Salonen, 1984): • occipital encephalocele (or other CNS abnormality) • bilateral large multicystic kidneys • fibrotic changes in the liver • other features: bilateral postaxial polydactyly, microphthalmia, cleft lip and palate, heart defects, genetic abnormalities, bowing of long bones, situs inversus, low set ears etc. Diagnosis • Via ultrasound at 11-14 weeks • Often an autopsy is necessary

4. Genetic aspects • Associated with the dysfunction of primary cilia • Autosomal recessive • Genetic and clinical heterogeneity • Oligogenic inheritance

5. Genetic aspects • Associated with the dysfunction of primary cilia • Autosomal recessive • Genetic and clinical heterogeneity • Oligogenic inheritance

6. MKS1 and MKS3 mutations MKS3/Meckelin MKS1 B9 domain Khaddour et al. (2007) Human Mutation 28(5); 523-4

7. The need for a diagnostic service for MKS • Prior to this project, no CPA accredited laboratory offered MKS testing • Mutation scanning performed on a research basis by Dr Colin Johnson at the Leeds Institute of Molecular Medicine • approximately 50 requests, nationally and internationally, for screening annually • In the local population the incidence of MKS may be as high as 1 in 3000 • A diagnostic service would allow • accurate diagnosis • confirmation of research results • carrier testing in at-risk individuals • prenatal testing

8. Referrals to DNA Lab Pakistani origin Other origin Screen for common MKS3 splice-site mutations Autozygosity or linkage analysis at MKS1 & MKS3 Targeted mutation screen by sequencing Proposed test strategy • Clinical sensitivity of testing for mutations in MKS1 and MKS3 in the general population is approximately 15% • In the local Pakistani population sensitivity for the MKS3 c.1575+1G>A mutation alone is estimated at 40%

9. The story so far... • Bidirectional sequencing optimised for: • entire coding region of MKS1 (18 exons) • exons 1-18 of MKS3 • Microsatellite analysis for MKS1 and MKS3 loci optimised: • MKS1 17q22D17S1853 and D17S1290 • MKS3 8q22 D8S1818 and D8S1699 • Reports issued: • 35 confirmations of research findings • 30 locally • 5 nationally • it is anticipated that these results will lead to cascade carrier testing and prenatal diagnosis requests • Gene dossier to be submitted to UKGTN April 2008

10. Exon16 | Intron 16 Mutations reported 482 483 484 485 486 487 488 Thr ThrGly Thr Val Thr Phe MKS1 • c.1451_1453dupGGCA (p.Thr485fs) • Pakistani • 4bp duplication in exon 16 MKS3 • c.1674+1G>A • Pakistani • mutation abolishes exon 16 splice donor site Thr ThrGly Arg His Cys His Upper panel: wild-type Lower panel: homozygous mutant

11. Intron 8 | Exon 9 Mutations reported Exon 15 | Intron 15 MKS3 • c.1575+1G>A • Pakistani • mutation abolishes exon 15 splice donor site • Estimated allele frequency in the local Pakistani population is 0.016; carrier frequency approximately 1/32 • Variable phenotype e.g. CNS, polydactyly. Inter- and intra-familial variation • c.870-2A>G • Pakistani • mutation abolishes exon 9 splice acceptor site Upper panel: wild-type Lower panel: homozygous mutant

12. MKS testing costs Please contact the laboratory for further information on testing for Meckel-Gruber syndrome. helen.lindsay@leedsth.nhs.uk

13. Acknowledgements Leeds Institute of Molecular Medicine • Colin Johnson Yorkshire Regional DNA Laboratory • Kim Flintoff, David Cockburn, Ruth Charlton Yorkshire Clinical Genetics Service • Chris Bennett