Development of a Genotype Assay for Age-Related Macular Degeneration: The EYE-RISK Consortium

Anita de Breuk, Ilhan E Acar, Eveline Kersten, Mascha M V A P Schijvenaars, Johanna M Colijn, Lonneke Haer-Wigman, Bjorn Bakker, Sarah de Jong, Magda A Meester-Smoor, Timo Verzijden, Tom O A R Missotten, Jordi Monés, Marc Biarnés, Daniel Pauleikhoff, Hans W Hense, Rufino Silva, Sandrina Nunes, Joana B Melo, Sascha Fauser, Carel B HoyngMarius Ueffing, Marieke J H Coenen, Caroline C W Klaver, Anneke I den Hollander,

Research output: Contribution to journalArticleResearchpeer-review


PURPOSE: To develop a genotype assay to assess associations with common and rare AMD risk variants, to calculate an overall genetic risk score (GRS), and to identify potential misdiagnoses with inherited macular dystrophies that mimic AMD.

DESIGN: Case-control study.

PARTICIPANTS: Individuals (N=4,740) from five European cohorts.

METHODS: We designed single molecule molecular inversion probes (smMIPs) for target selection and used next generation sequencing (NGS) to sequence eighty-seven single nucleotide polymorphisms (SNPs), coding and splice-site regions of ten AMD-(related) genes (ARMS2, C3, C9, CD46, CFB, CFH, CFI, HTRA1, TIMP3, SLC16A8), and three genes that cause inherited macular dystrophies (ABCA4, CTNNA1, PRPH2). GRS for common AMD risk variants were calculated based on effect size and genotype of 52 AMD-associated variants. Frequency of rare variants was compared between late AMD cases and control individuals with logistic regression analysis.

MAIN OUTCOME MEASURES: GRS, association of genetic variants with AMD, genotype-phenotype correlations.

RESULTS: We observed high concordance rates between our platform and other genotyping platforms for the 69 successfully genotyped SNPs (96.77-97.28%) and for the rare variants (99.81%). We observed a higher GRS for patients with late AMD compared to patients with early/intermediate AMD (p<0.001) and individuals without AMD (p<0.001). A higher proportion of rare loss-of-function variants and variants with a Combined Annotation Dependent Depletion score ≥20 in the CFH (50 [2.92%] vs 8 [1.02%], OR=2.88 [1.36-6.11], p=0.006), CFI (38 [2.22%] vs 4 [0.51%], OR=4.45 [1.58-12.50], p=0.005) and C3 (56 [3.27%] vs 4 [0.51%], OR=6.56 [2.37-18.17], p=0.0003) genes was observed in late AMD cases compared to control individuals. In nine patients we identified pathogenic variants in the PRPH2, ABCA4 and CTNNA1 genes, which allowed reclassification of these patients as inherited macular dystrophy.

CONCLUSIONS: This study reports a high-throughput and comprehensive genotype assay for common and rare AMD genetic variants. This test can identify individuals at intermediate to high genetic risk of late AMD, and enables differential diagnosis of AMD mimicking dystrophies. Our study supports sequencing of CFH, CFI and C3 genes as they harbor rare high-risk loss-of-function variants. Carriers of these variants could be amendable for new treatments for AMD that are currently under development.

Original languageEnglish
Early online date24 Jul 2020
Publication statusPublished - Jul 2020

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    de Breuk, A., Acar, I. E., Kersten, E., Schijvenaars, M. M. V. A. P., Colijn, J. M., Haer-Wigman, L., Bakker, B., de Jong, S., Meester-Smoor, M. A., Verzijden, T., Missotten, T. O. A. R., Monés, J., Biarnés, M., Pauleikhoff, D., Hense, H. W., Silva, R., Nunes, S., Melo, J. B., Fauser, S. (2020). Development of a Genotype Assay for Age-Related Macular Degeneration: The EYE-RISK Consortium. Ophthalmology.