Developing a diagnostic service for Stargardt disease – a feasibility study

1. Developing a diagnostic service for Stargardt disease – a feasibility study Emily Packham Oxford Regional Molecular Genetics Laboratory

2. Introduction • Inherited eye disorders • Services currently available for some of the AD and X-linked conditions • Limited services currently for AR conditions (Asper Ophthalmics offer commercial genotyping of some genes) • Why? • Significant clinical overlap • Genetically heterogeneous • Stargardt disease may be feasible

3. Stargardt disease • Autosomal recessive juvenile macular degeneration • Prevalence of 1 in 10,000

4. Stargardt disease/Fundus Flavimaculatus (STGD/FF) Characterised by yellow-white flecks and atrophy STGD FF

5. Symptoms • Age of onset varies from early childhood to twenties • Early stages – difficulty reading, watching TV, missing patches in vision, photophobia, slow dark adaption • Later stages – always disturbance of central vision and sometimes: peripheral disturbance, increasing photophobia or problems with dark vision

6. Diagnosis • Clinical diagnosis • Sophisticated imaging but dependent on tests performed, experience and stage of disorder • Late stage shows clinical overlap • Genetic diagnosis • Support or confirm diagnosis • Provide prognosis information • Aid genetic counselling • Therapeutic intervention

7. Genetics of Stargardts disease • ABCA4 (1p13-p22) • 50 exons (6819bp ORF) • Highly polymorphic • No mutation hotspots • 500+ variants identified • Most common seen in ~ <10% of patients • Many missense variants

8. ABCA4 protein (ABCR / Rim) • ATP-binding cassette (ABC) transporter superfamily • Transmembrane proteins involved in transportation of compounds across cell membranes • 2273 amino acid protein expressed in cones and rods

10. ABCA4 function and disease pathology • Actively ‘flips’ Ret-PE across disc rim membrane • Enables retinal signalling to continue • Loss-of-function mutations • Loss of/reduction in ABCA4 function results in accumulation of toxic lipofuscin deposits • Destroys retinal pigment epithelium and rod and cone cells, resulting in visual loss

11. ABCA4 and other retinopathies • Stargardt disease • AR cone-rod dystrophy • AR retinitis pigmentosa • Age-related macular degeneration? • Genotype/phenotype correlation model based on residual activity of protein

12. Screening strategy • 30 patients selected for testing • Highly polymorphic, 50 exon gene with no particular hotspots • Bi-directional sequencing • Robotics approach –5 patients per batch • Pathogenicity investigations performed • MLPA

13. Results • 37 different potential pathogenic variants detected in 26 patients • 13/20 patients with two or more variants had all of them classified as either likely or highly likely • Most common seen in 4 patients

14. Results

15. Results • Extensive published data • MLPA normal in all 10 patients tested

16. Feasibility • Clinical sensitivity • 67% or 43% (+/- intermediate variants) • Higher than literature • Different screening methods and patient selection • Clinical utility • Able to interpret most variants • Supports clinical diagnosis, aids counselling and therapy • Improves equity of access

17. What next? • Report our 30 patients • Determine if variants are in trans • Submit gene dossier • Collaborate with BRC retinal research project • Evaluating use of high throughput sequencing to test numerous inherited retinal conditions NHS lab BRC

18. Acknowledgements • Oxford Molecular Genetics Laboratory • Anneke Seller • Treena Cranston • Tina Bedenham, Louise Williams, Kate Gibson • Oxford Clinical Genetics and BRC • Andrea Nemeth • Oxford eye hospital • Susan Downes