1. Development of a New Method to Prioritise Gene Analysis in Familial Hypertrophic Cardiomyopathy Jayne Duncan West of Scotland Regional Genetics Service, Glasgow

2. Familial Hypertrophic Cardiomyopathy (FHC) Autosomal dominant disorder showing variable penetrance and age of onset. Affects approximately 1/500 adults and is the most common cause of sudden death in young healthy individuals. So far mutations in over 20 genes have been associated with FHC

3. Primary Clinical Features of FHC Left ventricular hypertrophy, “a thickening of the tissue due to increased size of the constituent cells”. Myocyte/myofibrillar disarray caused by the abnormal shapes, intracellular connections and arrangement of the hypertrophic myocytes and fibrosis. 1 http://www.maxshouse.comhttp://www.maxshouse.com 2 Arad et al 2002 Hum Mol Genet. 11. (20) 2499-2506

4. Genotype Phenotype Correlation

5. The Heterogeneous Nature of FHC HCM is caused by dominant mutations in the sarcomeric genes. de novo mutations occur rarely and account for approximately 10% of cases. Mutations in the sarcomeric genes account for ~55% of cases of HCM. Syndromes such as the Glycogen storage disorders and Friedreich ataxia can mimic HCM.

6. Glasgow Linkage Exclusion Analysis Method (GLEAM) Novel method to prioritise gene analysis in heterogeneous disorders A gene is excluded from analysis when affected relatives are oppositely homozygous for SNPs in and around the gene of interest

7. GLEAM A and B represent alleles at a susceptibility locus for a dominantly inherited disorder affecting individuals II:2, II:3, III:1 and III4. Since III:1 has no allele in common with II:3 or III:4 it effectively rules out this locus as being responsible for the disease in this family. BB AB AA AB BB AB AA

8. Genes analysed in the FHC Project Gene NameChromosomeN o ExonsN o SNPs TTN2363212 MYH7143876 MYH6143777 MYBPC31135147 RAF1316163 PRKAG2716146 TPM11516132 TNNT2115116 MYLK2201294 TNNI319892 MYL33789 MYL212794 CAV33298

9. SNP Analysis Platform 96 fibre optic bundles on each plate Each fibre contains a bead that corresponds to each SNP Image taken from www.Illumina.comwww.Illumina.com Sentrix Array Matrix

10. Results- Raw Data Raw data for one patient sample

11. Results- Raw Data Clustered patient SNP data for a single SNP locus AA BB AB

12. Results- Genotype Comparisons

13. Results RelationshipNumber of pairsAverage number of genes excluded Sibs493 Aunt/Niece Nephew 225 First Cousins107 First Cousins once removed 58 Second Cousins97 Grandparent/ Grandchild 12

14. Results GeneNumber of times gene excluded Percentage number of times gene excluded TTN34/9635% MYH6 and MYH733/9634% MYBPC328/9629% RAF123/9624% PRKAG247/9649% TPM129/9630% TNNT239/9641% MYLK240/9642% TNNI334/9635% MYL333/9634% MYL226/9627% CAV340/9642%

15. Interesting Case H15.1 H15.4 H15.7 H15.14 H15.15 H15.16 H15.12 TNNI3/ TNNI3/ MYBPC3 TNNI3 TNNI3 TNNI3 TNNI3 MYBPC3 MYBPC3

16. Interesting Case Familial mutation in TNNI3 was not excluded in all affected family members. Comparisons between H15.1, H15.4 and H15.7 did not exclude MYBPC3. MYBPC3 was excluded when H15.1, H15.4 and H15.7 were compared against other family members who did not have this mutation. Testing for the TNNI3 mutation in H15.7 would have been negative and suggested a second mutation prompting further analysis.

17. Conclusions For all the pedigrees with one known mutation, this gene was not excluded in any of the analyses performed. More genes tend to be excluded when more distantly related individuals such as first cousins or aunt/niece, nephew pairs are considered, rather than more closely related sibs GLEAM can be used to determine the order in which genes are sequenced in heterogeneous disorders

18. Acknowledgements Scottish Health Innovations Ltd Dr Wai Lee & Dr Stewart Lang, British Heart Foundation, Glasgow Cardiovascular Research Centre, University of Glasgow.  Dr Petros Syrris, Department of Medicine, University College London