Association of Sex With Frequent and Mild ABCA4 Alleles in Stargardt Disease | Genetics and Genomics | JAMA Ophthalmology | JAMA Network
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Figure 1.  Comparison of Sex Distribution Among Patients With and Without a Mild ABCA4 Allele
Comparison of Sex Distribution Among Patients With and Without a Mild ABCA4 Allele

A, More women than men carried the mild allele c.5882G>A (54:25) or c.5603A>T (79:46), whereas the female to male ratio among patients not carrying a mild allele was exactly 1 (142:142). B, In Stargardt disease (STGD1) caused by the mild allele c.5882G>A or c.5603A>T, the proportion of women (68% and 63%, respectively) was significantly higher than in STGD1 not caused by a mild ABCA4 allele (13% [95% CI, 3%-23%; P = .02] vs 18% [95% CI, 6%-30%; P = .005]).

aP < .03.

Figure 2.  Age at Onset by Sex and Carriers of ABCA4 Allele
Age at Onset by Sex and Carriers of ABCA4 Allele

Horizontal lines and error bars represent the median with the interquartile range. No difference in age at onset was observed between men and women in any of the 3 subgroups (ie, biallelic patients not carrying a known mild allele, patients carrying the c.5882G>A allele, and patients carrying the c.5603A>T allele).

Figure 3.  Sex and Other Modifying Factors Associated With Risk for Stargardt Disease (STGD1)
Sex and Other Modifying Factors Associated With Risk for Stargardt Disease (STGD1)

Shown is a modification of the previously proposed ABCA4 disease model that associated genotypes with phenotypes on the basis of the residual activity of the ABCA4 protein.24-26 In individuals heterozygous for 1 severe variant, ABCA4 activity was reduced to 50%, which does not lead to STGD1. In a previous in vitro study, individually tested variants were considered mild if the percentage of wild-type messenger RNA, which we extrapolated to the residual protein activity, was 60% to 80%.21 Therefore, a combination of a severe and a mild allele leads to a total residual activity of approximately 30% to 40% and results in classic STGD1 (mild/severe bar). Because the expression of a mild, low-penetrant allele was variable, its combination with a severe variant could result either in generally late-onset STGD1 or in a normal phenotype (Mild, low penetrant/severe bar). Only in this latter category of individuals might sex and other genetic, epigenetic, and nongenetic modifiers alter the risk for developing STGD1.

Table 1.  Study Participants
Study Participants
Table 2.  Penetrance of Mild ABCA4 Alleles When Accompanied by a Severe ABCA4 Allele
Penetrance of Mild ABCA4 Alleles When Accompanied by a Severe ABCA4 Allele
1.
Blacharski  P. Fundus flavimaculatus. In: Newsome  DA, ed.  Retinal Dystrophies and Degenerations. Vol 135-159. Raven Press; 1988.
2.
Allikmets  R, Singh  N, Sun  H,  et al.  A photoreceptor cell-specific ATP-binding transporter gene (ABCR) is mutated in recessive Stargardt macular dystrophy.   Nat Genet. 1997;15(3):236-246. doi:10.1038/ng0397-236 PubMedGoogle ScholarCrossref
3.
Rotenstreich  Y, Fishman  GA, Anderson  RJ.  Visual acuity loss and clinical observations in a large series of patients with Stargardt disease.   Ophthalmology. 2003;110(6):1151-1158. doi:10.1016/S0161-6420(03)00333-6 PubMedGoogle ScholarCrossref
4.
Fujinami  K, Zernant  J, Chana  RK,  et al.  Clinical and molecular characteristics of childhood-onset Stargardt disease.   Ophthalmology. 2015;122(2):326-334. doi:10.1016/j.ophtha.2014.08.012 PubMedGoogle ScholarCrossref
5.
Lambertus  S, van Huet  RA, Bax  NM,  et al.  Early-onset Stargardt disease: phenotypic and genotypic characteristics.   Ophthalmology. 2015;122(2):335-344. doi:10.1016/j.ophtha.2014.08.032 PubMedGoogle ScholarCrossref
6.
Westeneng-van Haaften  SC, Boon  CJ, Cremers  FP, Hoefsloot  LH, den Hollander  AI, Hoyng  CB.  Clinical and genetic characteristics of late-onset Stargardt’s disease.   Ophthalmology. 2012;119(6):1199-1210. doi:10.1016/j.ophtha.2012.01.005 PubMedGoogle ScholarCrossref
7.
Burke  TR, Tsang  SH, Zernant  J, Smith  RT, Allikmets  R.  Familial discordance in Stargardt disease.   Mol Vis. 2012;18:227-233.PubMedGoogle Scholar
8.
Runhart  EH, Valkenburg  D, Cornelis  SS,  et al.  Late-onset Stargardt disease due to mild, deep-intronic ABCA4 alleles.   Invest Ophthalmol Vis Sci. 2019;60(13):4249-4256. doi:10.1167/iovs.19-27524 PubMedGoogle ScholarCrossref
9.
Valkenburg  D, Runhart  EH, Bax  NM,  et al.  Highly variable disease courses in siblings with Stargardt disease.   Ophthalmology. 2019;126(12):1712-1721. doi:10.1016/j.ophtha.2019.07.010 PubMedGoogle ScholarCrossref
10.
Zernant  J, Lee  W, Collison  FT,  et al.  Frequent hypomorphic alleles account for a significant fraction of ABCA4 disease and distinguish it from age-related macular degeneration.   J Med Genet. 2017;54(6):404-412. doi:10.1136/jmedgenet-2017-104540 PubMedGoogle ScholarCrossref
11.
Runhart  EH, Sangermano  R, Cornelis  SS,  et al.  The common ABCA4 Variant p.Asn1868Ile shows nonpenetrance and variable expression of Stargardt disease when present in trans with severe variants.   Invest Ophthalmol Vis Sci. 2018;59(8):3220-3231. doi:10.1167/iovs.18-23881 PubMedGoogle ScholarCrossref
12.
Zernant  J, Lee  W, Nagasaki  T,  et al.  Extremely hypomorphic and severe deep intronic variants in the ABCA4 locus result in varying Stargardt disease phenotypes.   Cold Spring Harb Mol Case Stud. 2018;4(4):a002733. doi:10.1101/mcs.a002733 PubMedGoogle Scholar
13.
Cornelis  SS, Bax  NM, Zernant  J,  et al.  In silico functional meta-analysis of 5,962 ABCA4 variants in 3,928 retinal dystrophy cases.   Hum Mutat. 2017;38(4):400-408. doi:10.1002/humu.23165 PubMedGoogle ScholarCrossref
14.
Sangermano  R, Garanto  A, Khan  M,  et al.  Deep-intronic ABCA4 variants explain missing heritability in Stargardt disease and allow correction of splice defects by antisense oligonucleotides.   Genet Med. 2019;21(8):1751-1760. doi:10.1038/s41436-018-0414-9 PubMedGoogle ScholarCrossref
15.
Cremers  FPM, Cornelis  SS, Runhart  EH, Astuti  GDN.  Author response: penetrance of the ABCA4 p.Asn1868Ile allele in Stargardt disease.   Invest Ophthalmol Vis Sci. 2018;59(13):5566-5568. doi:10.1167/iovs.18-25944 PubMedGoogle ScholarCrossref
16.
Cremers  FPM, Lee  W, Collin  RWJ, Allikmets  R.  Clinical spectrum, genetic complexity and therapeutic approaches for retinal disease caused by ABCA4 mutations.   Prog Retin Eye Res. 2020;100861. doi:10.1016/j.preteyeres.2020.100861 PubMedGoogle Scholar
17.
Khan  M, Cornelis  SS, Khan  MI,  et al.  Cost-effective molecular inversion probe-based ABCA4 sequencing reveals deep-intronic variants in Stargardt disease.   Hum Mutat. 2019;40(10):1749-1759. doi:10.1002/humu.23787 PubMedGoogle ScholarCrossref
18.
Khan  M, Cornelis  SS, Pozo-Valero  MD,  et al.  Resolving the dark matter of ABCA4 for 1054 Stargardt disease probands through integrated genomics and transcriptomics.   Genet Med. 2020;22(7):1235-1246. doi:10.1038/s41436-020-0787-4 PubMedGoogle ScholarCrossref
19.
World Medical Association.  World Medical Association Declaration of Helsinki: ethical principles for medical research involving human subjects.   JAMA. 2013;310(20):2191-2194. doi:10.1001/jama.2013.281053PubMedGoogle ScholarCrossref
20.
Karczewski  KJ, Francioli  LC, Tiao  G,  et al; Genome Aggregation Database Consortium.  The mutational constraint spectrum quantified from variation in 141,456 humans.   Nature. 2020;581(7809):434-443. doi:10.1038/s41586-020-2308-7PubMedGoogle ScholarCrossref
21.
Sangermano  R, Khan  M, Cornelis  SS,  et al.  ABCA4 midigenes reveal the full splice spectrum of all reported noncanonical splice site variants in Stargardt disease.   Genome Res. 2018;28(1):100-110. doi:10.1101/gr.226621.117 PubMedGoogle ScholarCrossref
22.
Pisano  A, Preziuso  C, Iommarini  L,  et al.  Targeting estrogen receptor β as preventive therapeutic strategy for Leber’s hereditary optic neuropathy.   Hum Mol Genet. 2015;24(24):6921-6931. doi:10.1093/hmg/ddv396 PubMedGoogle Scholar
23.
Oostra  RJ, Kemp  S, Bolhuis  PA, Bleeker-Wagemakers  EM.  No evidence for ‘skewed’ inactivation of the X-chromosome as cause of Leber’s hereditary optic neuropathy in female carriers.   Hum Genet. 1996;97(4):500-505. doi:10.1007/BF02267075 PubMedGoogle ScholarCrossref
24.
Cremers  FP, van de Pol  DJ, van Driel  M,  et al.  Autosomal recessive retinitis pigmentosa and cone-rod dystrophy caused by splice site mutations in the Stargardt’s disease gene ABCR.   Hum Mol Genet. 1998;7(3):355-362. doi:10.1093/hmg/7.3.355 PubMedGoogle ScholarCrossref
25.
van Driel  MA, Maugeri  A, Klevering  BJ, Hoyng  CB, Cremers  FP.  ABCR unites what ophthalmologists divide(s).   Ophthalmic Genet. 1998;19(3):117-122. doi:10.1076/opge.19.3.117.2187 PubMedGoogle ScholarCrossref
26.
Maugeri  A, van Driel  MA, van de Pol  DJ,  et al.  The 2588G→C mutation in the ABCR gene is a mild frequent founder mutation in the Western European population and allows the classification of ABCR mutations in patients with Stargardt disease.   Am J Hum Genet. 1999;64(4):1024-1035. doi:10.1086/302323 PubMedGoogle ScholarCrossref
27.
Schulz  HL, Grassmann  F, Kellner  U,  et al.  Mutation spectrum of the ABCA4 gene in 335 Stargardt disease patients from a multicenter German cohort - impact of selected deep intronic variants and common SNPs.   Invest Ophthalmol Vis Sci. 2017;58(1):394-403. doi:10.1167/iovs.16-19936 PubMedGoogle ScholarCrossref
28.
Franconi  F, Campesi  I, Colombo  D, Antonini  P.  Sex-gender variable: methodological recommendations for increasing scientific value of clinical studies.   Cells. 2019;8(5):E476. doi:10.3390/cells8050476 PubMedGoogle Scholar
29.
Gerdts  E, Regitz-Zagrosek  V.  Sex differences in cardiometabolic disorders.   Nat Med. 2019;25(11):1657-1666. doi:10.1038/s41591-019-0643-8 PubMedGoogle ScholarCrossref
30.
De Bellis  A, De Angelis  G, Fabris  E, Cannatà  A, Merlo  M, Sinagra  G.  Gender-related differences in heart failure: beyond the “one-size-fits-all” paradigm.   Heart Fail Rev. 2020;25(2):245-255. doi:10.1007/s10741-019-09824-y PubMedGoogle ScholarCrossref
31.
Billi  AC, Kahlenberg  JM, Gudjonsson  JE.  Sex bias in autoimmunity.   Curr Opin Rheumatol. 2019;31(1):53-61.PubMedGoogle ScholarCrossref
32.
Ventura-Clapier  R, Moulin  M, Piquereau  J,  et al.  Mitochondria: a central target for sex differences in pathologies.   Clin Sci (Lond). 2017;131(9):803-822. doi:10.1042/CS20160485 PubMedGoogle ScholarCrossref
33.
Sampathkumar  NK, Bravo  JI, Chen  Y,  et al.  Widespread sex dimorphism in aging and age-related diseases.   Hum Genet. 2020;139(3):333-356. doi:10.1007/s00439-019-02082-w PubMedGoogle ScholarCrossref
34.
Wickham  LA, Gao  J, Toda  I, Rocha  EM, Ono  M, Sullivan  DA.  Identification of androgen, estrogen and progesterone receptor mRNAs in the eye.   Acta Ophthalmol Scand. 2000;78(2):146-153. doi:10.1034/j.1600-0420.2000.078002146.x PubMedGoogle ScholarCrossref
35.
Marin-Castaño  ME, Elliot  SJ, Potier  M,  et al.  Regulation of estrogen receptors and MMP-2 expression by estrogens in human retinal pigment epithelium.   Invest Ophthalmol Vis Sci. 2003;44(1):50-59. doi:10.1167/iovs.01-1276 PubMedGoogle ScholarCrossref
36.
Hulsman  CA, Westendorp  IC, Ramrattan  RS,  et al.  Is open-angle glaucoma associated with early menopause? The Rotterdam Study.   Am J Epidemiol. 2001;154(2):138-144. doi:10.1093/aje/154.2.138 PubMedGoogle ScholarCrossref
37.
Prokai-Tatrai  K, Xin  H, Nguyen  V,  et al.  17β-estradiol eye drops protect the retinal ganglion cell layer and preserve visual function in an in vivo model of glaucoma.   Mol Pharm. 2013;10(8):3253-3261. doi:10.1021/mp400313u PubMedGoogle ScholarCrossref
38.
Elliot  SJ, Catanuto  P, Espinosa-Heidmann  DG,  et al.  Estrogen receptor beta protects against in vivo injury in RPE cells.   Exp Eye Res. 2010;90(1):10-16. doi:10.1016/j.exer.2009.09.001 PubMedGoogle ScholarCrossref
39.
Du  M, Mangold  CA, Bixler  GV,  et al.  Retinal gene expression responses to aging are sexually divergent.   Mol Vis. 2017;23:707-717.PubMedGoogle Scholar
40.
Li  B, Gografe  S, Munchow  A, Lopez-Toledano  M, Pan  ZH, Shen  W.  Sex-related differences in the progressive retinal degeneration of the rd10 mouse.   Exp Eye Res. 2019;187:107773. doi:10.1016/j.exer.2019.107773 PubMedGoogle Scholar
41.
Guarneri  R, Russo  D, Cascio  C,  et al.  Retinal oxidation, apoptosis and age- and sex-differences in the mnd mutant mouse, a model of neuronal ceroid lipofuscinosis.   Brain Res. 2004;1014(1-2):209-220. doi:10.1016/j.brainres.2004.04.040 PubMedGoogle ScholarCrossref
42.
Poloschek  CM, Bach  M, Lagrèze  WA,  et al.  ABCA4 and ROM1: implications for modification of the PRPH2-associated macular dystrophy phenotype.   Invest Ophthalmol Vis Sci. 2010;51(8):4253-4265. doi:10.1167/iovs.09-4655 PubMedGoogle ScholarCrossref
43.
Lee  W, Paavo  M, Zernant  J,  et al.  Modification of the PROM1 disease phenotype by a mutation in ABCA4.   Ophthalmic Genet. 2019;40(4):369-375. doi:10.1080/13816810.2019.1660382 PubMedGoogle ScholarCrossref
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    Original Investigation
    August 20, 2020

    Association of Sex With Frequent and Mild ABCA4 Alleles in Stargardt Disease

    Author Affiliations
    • 1Department of Ophthalmology, Radboud University Medical Center, Nijmegen, the Netherlands
    • 2Donders Institute for Brain, Cognition and Behavior, Radboud University Medical Center, Nijmegen, the Netherlands
    • 3Department of Human Genetics, Radboud University Medical Center, Nijmegen, the Netherlands
    • 4Department of Genetics, Instituto de Investigación Sanitaria–Fundación Jiménez Díaz University Hospital, Universidad Autónoma de Madrid, Madrid, Spain
    • 5Center for Biomedical Network Research on Rare Diseases, Instituto de Salud Carlos III, Madrid, Spain
    • 6Centre for Ophthalmology and Visual Science, The University of Western Australia, Nedlands, Western Australia, Australia
    • 7Australian Inherited Retinal Disease Registry and DNA Bank, Department of Medical Technology and Physics, Sir Charles Gairdner Hospital, Nedlands, Western Australia, Australia
    • 8Research Unit for Rare Diseases, Department of Pediatrics and Adolescent Medicine, First Faculty of Medicine, Charles University and General University Hospital in Prague, Prague, Czech Republic
    • 9Department of Ophthalmology, First Faculty of Medicine, Charles University and General University Hospital in Prague, Prague, Czech Republic
    • 10University of Cape Town/MRC Genomic and Precision Medicine Research Unit, Division of Human Genetics, Department of Pathology, Institute of Infectious Disease and Molecular Medicine, Faculty of Health Sciences, University of Cape Town, Cape Town, South Africa
    • 11Institute of Human Genetics, University of Regensburg, Regensburg, Germany
    • 12Department of Ophthalmology, Erasmus Medical Center, Rotterdam, the Netherlands
    • 13Department of Epidemiology, Erasmus Medical Center, Rotterdam, the Netherlands
    • 14University Lille, Inserm, CHU Lille, U1172-LilNCog-Lille Neuroscience & Cognition, Lille, France
    JAMA Ophthalmol. 2020;138(10):1035-1042. doi:10.1001/jamaophthalmol.2020.2990
    Key Points

    Question  Are mild ABCA4 alleles associated with the patient’s sex in Stargardt disease?

    Findings  In this cross-sectional study of 550 patients with genetically confirmed Stargardt disease, a female predilection was observed among patients who carried a mild ABCA4 genotype compared with patients who harbored (moderately) severe ABCA4 genotypes.

    Meaning  This study found that, in the 25% of patients with Stargardt disease who carried a combination of a mild and a severe allele, sex may play a modifying role in the disease.

    Abstract

    Importance  The mechanisms behind the phenotypic variability and reduced penetrance in autosomal recessive Stargardt disease (STGD1), often a blinding disease, are poorly understood. Identification of the unknown disease modifiers can improve patient and family counseling and provide valuable information for disease management.

    Objective  To assess the association of incompletely penetrant ABCA4 alleles with sex in STGD1.

    Design, Setting, and Participants  Genetic data for this cross-sectional study were obtained from 2 multicenter genetic studies of 1162 patients with clinically suspected STGD1. Unrelated patients with genetically confirmed STGD1 were selected. The data were collected from June 2016 to June 2019, and post hoc analysis was performed between July 2019 and January 2020.

    Main Outcomes and Measures  Penetrance of reported mild ABCA4 variants was calculated by comparing the allele frequencies in the general population (obtained from the Genome Aggregation Database) with the genotyping data in the patient population (obtained from the ABCA4 Leiden Open Variation Database). The sex ratio among patients with and patients without an ABCA4 allele with incomplete penetrance was assessed.

    Results  A total of 550 patients were included in the study, among which the mean (SD) age was 45.7 (18.0) years and most patients were women (311 [57%]). Five of the 5 mild ABCA4 alleles, including c.5603A>T and c.5882G>A, were calculated to have incomplete penetrance. The women to men ratio in the subgroup carrying c.5603A>T was 1.7 to 1; the proportion of women in this group was higher compared with the subgroup not carrying a mild allele (difference, 13%; 95% CI, 3%-23%; P = .02). The women to men ratio in the c.5882G>A subgroup was 2.1 to 1, and the women were overrepresented compared with the group carrying no mild allele (difference, 18%; 95% CI, 6%-30%; P = .005).

    Conclusions and Relevance  This study found an imbalance in observed sex ratio among patients harboring a mild ABCA4 allele, which concerns approximately 25% of all patients with STGD1, suggesting that STGD1 should be considered a polygenic or multifactorial disease rather than a disease caused by ABCA4 gene mutations alone. The findings suggest that sex should be considered as a potential disease-modifying variable in both basic research and clinical trials on STGD1.

    Introduction

    Autosomal recessive Stargardt disease (STGD1) is the most frequent inherited macular dystrophy.1 It is caused by mutations in both copies of the gene encoding the transmembrane ATP-binding cassette transporter type A4 (ABCA4 gene [OMIM 601691]).2 Patients generally report progressive central loss of vision in the first or second decade of life,3-5 but cases with a much later disease onset, even past the age of 80 years, have been described as well.6 This large clinical variability has also been observed in siblings sharing the same ABCA4 gene mutations and has remained largely unexplained to date.7-9 The lack of knowledge about the mechanisms underlying this clinical variability impairs not only patient and family counseling but also informed decision-making regarding study design and patient inclusion. Moreover, deciphering these mechanisms might lead to new targets for treating this incurable disease.

    Late-onset STGD1 and clinical variability are associated with mild variants, which generally are clinically relevant only when in trans with a severe ABCA4 variant.8,10-12 To date, 8 mild ABCA4 variants have been reported.8,10-13 Four coding variants—c.2588G>C, p.[Gly863Ala,Gly863del]; c.3113C>T, p.(Ala1038Val); c.5882G>A, p.(Gly1961Glu); and c.6089G>A, p.(Arg2030Gln)—and 1 noncanonical splice site variant—c.5714+5G>A, p.[=,Glu1863Leufs*33]—were deemed mild on the basis of their considerably lower than expected homozygous occurrence in patients, considering the respective allele frequencies in the general population.13 An additional mild coding variant—ie, c.5603A>T, p.(Asn1868Ile)—and 2 mild deep-intronic variants—c.769-784C>T, p.[=,Leu257Aspfs*3] and c.4253+43G>A, p.[=,Ile1377Hisfs*3]—were identified.10,12,14 Variant c.5603A>T was calculated to cause disease only in a small proportion of individuals (approximately 5%) who carried this allele in combination with a severe one.11,15 Variant c.2588G>C is likely benign and only has complete penetrance when present in the same gene copy as c.5603A>T (c.[2588G>C;5603A>T]).10 Similarly, in most cases of STGD1, with the deep-intronic variant c.769-784C>T, and in some cases with c.4253+43G>A, c.5603A>T was found in the same gene copy, and arguably only the complex alleles (c.[769-784C>T;5603A>T] and c.[4253+43G>A;5603A>T]) have complete penetrance.8 The mechanisms underlying incomplete penetrance, found for some ABCA4 genotypes, are still unknown; both cis- or trans-acting genetic modifiers and environmental factors might alter ABCA4 gene expression and the clinical presentation of STGD1.16

    In a small cohort of patients carrying the c.5603A>T allele, 22 of 34 were female patients (65%) and all 3 siblings unaffected by compound heterozygous were male patients.11 In another study, which compared siblings with STGD1, a sex difference appeared to be present; in 5 of 5 families in which a wide variation in age at onset was observed (13-39 years), the patient with the latest onset was male.9 A sex ratio imbalance in STGD1 has never been established despite abundant research into this disease. We hypothesized that only a subgroup of patients (ie, those carrying a low penetrant ABCA4 allele) might harbor sex-related risk factors for STGD1. Because sex-related differences are associated with the causes and severity of many common diseases as well as the treatment outcomes, we investigated the sex distribution in a large STGD1 patient cohort. To assess the penetrance of mild ABCA4 alleles and the sex distribution among patients with biallelic mutations, we used data from 2 recent genetic studies in which the ABCA4 gene of 1162 patients with a potential ABCA4-associated retinal dystrophy was fully sequenced.17,18 The ABCA4 Disease Consortium Study Group allowed us to validate the sex ratio imbalance for c.5603A>T and to identify the incomplete penetrance and sex ratio imbalance for c.5882G>A, another ABCA4 allele.

    Methods

    This cross-sectional study adhered to the tenets of the Declaration of Helsinki.19 Approval from institutional research ethics committees were obtained from all hospitals and medical facilities that participated in the 2 studies we analyzed (specified at the end of the article), and patients provided written informed consent before participation. No compensation or incentive was offered to patients for participation in the study. Data were collected from June 2016 to June 2019.

    Patient Selection

    Patients with genetically confirmed STGD1 were selected from 2 studies that involved 21 international and 4 national centers.17,18 In these studies, unrelated patients with a potential ABCA4-associated retinal dystrophy (eg, STGD; cone-rod dystrophy; unspecified macular dystrophies; and pattern dystrophy, a well-known phenocopy) underwent clinical examination by a local ophthalmologist specializing in inherited retinal diseases. The coding and noncoding regions of the ABCA4 gene were sequenced using 3866 single-molecule molecular inversion probes. In 555 of 1162 cases, STGD1 was genetically confirmed by the presence of either 2 or more (likely) pathogenic ABCA4 variants or c.5603A>T combined with a (likely) pathogenic ABCA4 variant. In addition, we collected data on sex, age, and age at onset. Age at onset was defined as the onset of initial symptoms reported by the patient. Patients with missing data on sex were excluded.

    In this study, we used the term sex, defined by biological characteristics, rather than gender, which refers to a person’s identity and the sociocultural expectations of behavior associated with a given sex. However, the 2 terms could potentially be used interchangeably in the given context.

    Penetrance Calculation

    As previously described for variants c.5603A>T, p.(Asn1868Ile); c.769-784C>T, p.[=,Leu257Aspfs*3]; and c.4253+43G>A, p.[=,Ile1377Hisfs*3],8,11,15 we aimed to first assess whether additional mild variants could have reduced penetrance. For this purpose, we performed penetrance calculations for 5 other alleles, which were deemed mild because of their absence or low occurrence in a homozygous state in patients; these alleles were c.5882G>A, p.(Gly1961Glu); c.5714+5G>A, p.[=,Glu1863Leufs*33]; c.6089G>A, p.(Arg2030Gln); c.2588G>C, p.[Gly863Ala,Gly863del]; and c.3113C>T, p.(Ala1038Val).13

    Penetrance was calculated by comparing the ABCA4 allele frequency data in the general population that were obtained from gnomAD (Genome Aggregation Database, version 2.1.1.),20 with the genotyping data in the patient population that were obtained from the ABCA4 LOVD (Leiden Open Variation Database, version 3.0). Given that most patients in the LOVD have a non-Finnish European ethnic background, only the non-Finnish European population in gnomAD was considered, which included data on 7718 genomes and 56 885 exomes. Patients in the LOVD who were reported to have an ethnic origin other than non-Finnish European were excluded, as were patients in whom fewer than 2 ABCA4 variants had been identified. Penetrance was calculated according to the same methods used in previous studies,8,11,15 which could be summarized as the ratio of the observed frequency of patients (cases) with the genotype of interest to the theoretically expected frequency of patients with this genotype. Expected frequency was calculated according to the respective allele frequency (AF) in the general population (controls). Thus, the formula, given an estimated STGD1 prevalence of 1:10 000,1 was as follows:

    Image description not available.

    The genotype of interest consisted of the combination mild variant of interest and severe variant. Variants in the gnomAD and LOVD were annotated as severe if they constituted stop mutations, frameshift mutations, or canonical splice site variants (eTable 1 in the Supplement). We also added noncanonical splice site variants21 and deep-intronic variants shown to result in less than 25% normal messenger RNA, which we deemed to be a conservative cutoff for allocating variants to the status of severe.21

    Statistical Analysis

    To examine whether the proportion of men and women in the patient group harboring an allele with reduced penetrance was different from the patient group without any of these alleles, we conducted a 2-tailed Fisher exact test. For the statistical analysis, the 2 most frequent mild variants c.5603A>T and c.5882G>A were selected from the total of 8 mild alleles. We applied Bonferroni correction for multiple testing in the main analysis, which consisted of 2 tests (α = .025). For patient groups who showed a sex ratio imbalance, we tested whether the age at onset differed between men and women using the Mann-Whitney test. A 2-tailed α = .05 indicated statistical significance.

    Analyses were performed with SPSS for Windows, version 25 (SPSS IBM), between July 2019 and January 2020.

    Results
    Description of the Cohort

    After exclusion of 5 patients for missing data on sex, we included a total of 550 patients in this study. In this cohort, the mean (SD) age was 45.7 (18.0) years, 456 patients (83%) were of non-Finnish European descent (eTable 2 in the Supplement), and 311 patients were women (57%) (Table 1). Mild alleles were frequently identified in 266 (48%) participants, often in women (169 of 266 [64%]). Because c.5603A>T was formerly not considered to be a causal variant, it was (not unexpectedly) found to be the most frequent variant in this cohort, present as a noncomplex allele (ie, no additional potentially pathogenic variant on the same gene copy) in 125 patients (23%). The other most frequent variant was c.5882G>A, which was found in a noncomplex configuration in 79 patients (14%). The 6 other mild variants were identified in a total of 64 patients (12%). These 6 variants were c.4253+43G>A, c.5714+5G>A, c.6089G>A, c.3113C>T, c.769-784C>T, and c.2588G>C.

    Reduced Penetrance of 5 Additional Mild Variants

    A total of 2031 cases in the LOVD met the criteria of having a non-Finnish European ethnic background and at least 2 ABCA4 variants. For variants c.5882G>A, c.5714+5G>A, and c.6089G>A, dividing the frequency of the genotype of interest in cases by the frequency of this genotype in controls and then multiplying the numbers by the STGD1 prevalence of 1:10 000 yielded reduced penetrance of 50.4% for c.5882G>A, 58.6% for c.5714+5G>A, and 36.4% for c.6089G>A (Table 2). For variants c.3113C>T and c.2588G>C, which can be part of frequent complex alleles c.[1622T>C;3113C>T] and c.[2588G>C;5603A>T], penetrance was calculated in the same manner, not taking potential cis-variants into consideration. The relative proportion of these complex alleles is generally higher in patients than in controls. Consequently, both the observed and the expected numbers of patients carrying the noncomplex alleles were overestimated, but the observed number was overestimated to a greater extent. Therefore, the penetrance 16.8% for c.3113C>T and 11.0% for c.2588G>C should be interpreted as an overestimation of the actual penetrance.

    Sex Ratio in STGD1 Associated With c.5603A>T and c.5882G>A

    The sex ratio in the subgroup of patients with none of the mild alleles was exactly 1 to 1. The women to men ratio in the subgroup carrying c.5603A>T was 1.7 to 1; the proportion of women in this group was higher compared with the subgroup not harboring a mild allele (difference, 13%; 95% CI, 3%-23%; P = .02) (Figure 1A and B). Furthermore, the sex ratio in the c.5882G>A group was 2.1 to 1 and the women were overrepresented compared with the group carrying no mild allele (difference, 18%; 95% CI, 6%-30%; P = .005) (Figure 1A and B). Among the patients who harbored any of the 6 other mild alleles (c.4253+43G>A, c.5714+5G>A, c.6089G>A, c.3113C>T, c.769-784C>T, and c.2588G>C), the overall women to men ratio was 1.4 to 1.

    To assess whether a survival bias from longer life expectancy of women could have a statistically significant implication for this study, we compared the current age of the women and men in the subgroups. The median age was not statistically significantly different between the sexes in any of the subgroups (eFigure 1 in the Supplement). Among patients harboring c.5603A>T or c.5882G>A, the sex ratio imbalance was observed regardless of age and was pronounced before the age of 60 years (eFigure 2 in the Supplement).

    Based on the observed sex ratio imbalance and the calculated penetrance, the sex-specific penetrance of c.5603A>T and c.5882G>A, when in trans with a severe allele, was approximately 6% for women and 4% for men with c.5603A>T, and 68% for women and 32% for men with c.5882G>A (eFigure 3 in the Supplement).

    Age at Disease Onset Among Patients

    No difference was observed in the age of disease onset between the sexes in any of the genotypic subgroups (Figure 2). Among patients who were carrying c.5603A>T, women reported initial symptoms at a median (range) age of 40.0 (9.0-86.0) years, whereas men reported symptoms at a median (range) age of 43.0 (5.0-65.0) years (difference, 1.0 year; 95% CI, –7.0 to 10.0 years; P = .78). In the c.5882G>A subgroup, age at onset was slightly right-skewed, with a median (range) of 21.0 (1.0-51.0) years in women and 19.5 (10.0-40.0) years in men (difference, 1.0 year; 95% CI, –7.0 to 9.0 years; P = .82) (Figure 2; eFigure 2B in the Supplement). As expected, the age at onset among patients who were carrying none of the mild alleles was the lowest, with a median (range) of 13.0 (5.0-82.0) years in women and 12.0 (1.0-52.0) years in men (difference, 2.0 years; 95% CI, 0.0-4.0 years; P = .27) (Figure 2). In this latter subgroup, the distribution of age at onset was clearly right-skewed, likely because most of these individuals carried 2 (moderately) severe alleles and only some variants could behave like mild variants (eFigure 2C in the Supplement).

    Discussion

    To our knowledge, no sex ratio imbalance has been established before in STGD1, nor has it been observed in other non-sex-related inherited retinal diseases, except for the maternally inherited Leber hereditary optic neuropathy, in which incomplete penetrance was observed and more men than women were affected.22,23 This international collaboration, complete ABCA4 sequencing data, and advanced understanding of mildly pathogenic ABCA4 alleles allowed us to establish that, in a genetically predefined patient group, those who harbored ABCA4 alleles with potential incomplete penetrance were predominantly women. This observation corroborated the evidence of reduced penetrance in STGD1 and pointed to the existence of sex-related modifiers of the expression of reduced penetrant ABCA4 alleles (Figure 3).24-26

    Women were overrepresented among patients who were carrying 1 of the 2 most frequent alleles, c.5603A>T or c.5882G>A, compared with patients who were not carrying a known mild allele. Approximately 10%10,11 of all patients with STGD1 harbored c.5603A>T and 15%13,27 had c.5882G>A as 1 of the alleles. A search for the mechanisms underlying incomplete penetrance and female preponderance in approximately 25% of all patients with STGD1 may yield new hints for the pathogenesis and management of this blinding disease. Although many future studies are likely needed to elucidate the cause of sex differences in STGD1 and other diseases, these findings warrant careful consideration of sex or gender as a variable in future preclinical and clinical trials. Currently, medical practice in general (diagnosis and treatment) is less evidence based for women than for men because of the underrepresentation of women in biomedical research.28 However, considering sex as a study variable is indispensable in this era of personalized medicine, given that sex differences in the prevalence, manifestation, progression, and outcome of multifactorial disorders are increasingly identified. For instance, studies have found that ischemic heart disease has a younger onset, a higher incidence, and a different manifestation in men vs women, and dilated cardiomyopathy has a higher prevalence, younger onset, and worse outcome in men, whereas left ventricular hypertrophy is more prevalent and less modifiable in women.29,30 Female predilection has been shown in many autoimmune diseases, most strikingly (7-10:1) in Sjögren syndrome, systemic lupus erythematosus, and thyroiditis.31

    The biological mechanisms underlying the sex-based differences in multifactorial diseases are not completely understood, but among the likely factors are sex chromosomes, sex hormones, and mitochondria.29,32,33 As estrogen, progesterone, and androgen receptors’ messenger RNAs have been identified in the human retina and specifically the retinal pigment epithelium,34,35 each of these sex hormones may play a causal role in sexual differences in retinal health and pathology. Research to date has mainly focused on estrogen, which had neuroprotective properties for retinal ganglion cells in a glaucoma rat model.36,37 Estrogen receptor β protected mice against retinal pigment epithelium injury.38 Moreover, sex hormone receptors, which are nuclear transcription factors that bind to DNA, are known to regulate gene expression. Sex-based differences in retinal gene expression have been observed in male and female mice.39

    Mitochondria are involved in steroid hormone synthesis, production of reactive oxygen species, and cell death.32 Sex hormones, in turn, regulate mitochondrial function. Two retinal disease models, PDE6B-associated retinitis pigmentosa40 and neuronal ceroid lipofuscinoses,41 showed an increased susceptibility to photoreceptor degeneration of female mice compared with age-matched male mice.

    In addition to biological processes, socioeconomic and behavioral factors seem obvious variables to consider in future studies of modifiers for STGD1. Environmental factors that could directly be associated with the pathophysiological processes (eg, accumulation of vitamin A derivatives and phototoxic reaction) are diet and sunlight exposure.

    Genetic modifiers independent of sex exist that could also explain why some individuals harboring a mild and a severe allele develop STGD1 whereas others do not. First, despite sequencing of the complete 128-kb ABCA4 gene using a targeted, short-read sequencing method, the presence of deep-intronic variants, inversions or insertions in cis with a small proportion of the mild variants that make them fully penetrant, could not be ruled out.18 Second, sequence variants within or outside the ABCA4 locus could change disease expression, for instance, through effects on the transcription machinery (eg, owing to variants in enhancer or silencer regions) or through protein-protein interaction (eg, variants in PROM1 [OMIM *604365], ROM1 [OMIM *180721], and PRPH2 [OMIM *179605]42,43). However, these genetic sequence variants are not likely to be factors in the observed sex ratio imbalance.

    Although we hypothesized that the factors associated with a female predilection in STGD1 could also be associated with a more severe or earlier onset phenotype in women, the data on age at onset did not support this hypothesis. Instead, specific disease modifiers might play a role at specific stages in life and could be different for men and women. Therefore, female patients are not necessarily expected to have more severe cases.

    Limitations

    This study has limitations that are mostly associated with its cross-sectional design. Cause and effect could not be determined. Selection bias (eg, potential sex differences in health seeking behavior or willingness to participate in research) could not be eliminated but was unlikely a major factor given that a reference group of patients from the same cohort was used. In addition, the finding that age at onset was not associated with sex argues against a substantial difference in health-seeking behavior that explains the sex ratio imbalance. This study corroborated a finding of a small, independent cohort study that a female predilection existed in the c.5603A>T genotypic subgroup.11 The female predilection in c.5882G>A cases was a novel finding and should be corroborated in an independent cohort. Self-reported age at onset might not reflect the actual disease onset, although this method is often used because of lack of a superior, accessible measure. The limitations of penetrance calculations have been previously discussed.11,15 These calculations rely on assumptions regarding disease prevalence, ethnic diversity, and absence of unidentified causal variants in cis with the mild variants. If disease prevalence was higher, the actual penetrance would be higher. If patients carried unidentified causal variants in cis with the mild variant, the actual penetrance of the mild variant would be lower. Thus, these calculations provided robust indications of incomplete penetrance rather than the exact penetrance rates.

    Conclusions

    This cross-sectional study demonstrated a female predilection in STGD1 among all patients who were carrying either of the 2 most frequent and mild ABCA4 alleles compared with patients who were not carrying a putative mild ABCA4 allele. This finding adds to the evidence of reduced penetrance for some frequent ABCA4 genotypes and highlights the potentially crucial role of sex in human health and disease that needs to be taken into consideration in future studies of STGD1.

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    Article Information

    Accepted for Publication: July 1, 2020.

    Published Online: August 20, 2020. doi:10.1001/jamaophthalmol.2020.2990

    Correction: This article was corrected on February 25, 2021, to add the online supplement containing the names of the members of the ABCA4 Disease Consortium Study Group so that those collaborators’ names will all appear on PubMed.

    Corresponding Author: Claire-Marie Dhaenens, PharmD, PhD, University Lille, Inserm, CHU Lille, U1172-LilNCog-Lille Neuroscience & Cognition, 59045 Lille, France (claire-marie.dhaenens@inserm.fr).

    Author Contributions: Drs Runhart and Dhaenens had full access to all of the data in the study and take responsibility for the integrity of the data and the accuracy of the data analysis.

    Concept and design: Runhart, Cremers, Dhaenens.

    Acquisition, analysis, or interpretation of data: All authors.

    Drafting of the manuscript: Runhart, Khan, Dhaenens.

    Critical revision of the manuscript for important intellectual content: Runhart, Cornelis, Roosing, Del Pozo-Valero, Lamey, Liskova, Roberts, Stöhr, Klaver, Hoyng, Cremers, Dhaenens.

    Statistical analysis: Runhart, Cornelis, Klaver.

    Obtained funding: Hoyng, Cremers, Dhaenens.

    Administrative, technical, or material support: Khan, Lamey, Liskova, Roberts, Stöhr, Dhaenens.

    Supervision: Roosing, Klaver, Hoyng, Cremers, Dhaenens.

    Other: Del Pozo-Valero.

    Conflict of Interest Disclosures: None reported.

    Funding/Support: This work was performed within the framework of ERN-EYE (European Reference Network). This study was funded by grants from the Federal Ministry of Education and Research and programs SVV/260367/2017 by the Spanish National Organization of the Blind. The Spanish cases were supported by grants CIBERER (06/07/0036), IIS-FJD Biobank PT13/0010/0012, and FIS (PI16/00425) from the Instituto de Salud Carlos III from the Spanish Ministry of Health as well as grant CAM, B2017/BMD-3721 from the regional government of Madrid and RAREGenomics-CM, which are partially supported by the European Regional Development Fund. Drs Runhart and Cremers, Dr Khan, and Ms Cornelis were supported by grant GR591 from the RetinaUK; grant FB18CRE from Fighting Blindness Ireland, a program of Horizon 2020, Marie Sklodowska-Curie Innovative Training Network StarT (European Training Network to Diagnose, Understand and Treat Stargardt Disease, a Frequent Inherited Blinding Disorder); grant PPA-0517-0717-RAD from the Foundation Fighting Blindness USA; the Rotterdamse Stichting Blindenbelangen, the Stichting Blindenhulp, and the Stichting tot Verbetering van het Lot der Blinden; the Landelijke Stichting voor Blinden en Slechtzienden, Macula Degeneratie fonds, and the Stichting Blinden-Penning, which contributed through Uitzicht 2016-12; and the Algemene Nederlandse Vereniging ter Voorkoming van Blindheid and Landelijke Stichting voor Blinden en Slechtzienden, which contributed through UitZicht 2014-13. Ms Del Pozo-Valero was sponsored by the Conchita Rábago Foundation. Dr Liskova was supported by programs UNCE 204064 and PROGRES Q26 of the Charles University and grant NU20-07-00182 from the Ministry of Health of the Czech Republic. Dr Dhaenens was supported by Groupement de Coopération Sanitaire Interrégional G4 qui réunit les Centres Hospitaliers Universitaires Amiens, Caen, Lille, et Rouen and by the Fondation Stargardt France.

    Role of the Funder/Sponsor: The funders had no role in the design and conduct of the study; collection, management, analysis, and interpretation of the data; preparation, review, or approval of the manuscript; and decision to submit the manuscript for publication.

    Group Information: Members of the ABCA4 Disease Consortium Study Group who participated in this study are listed in Supplement 2. They include the following: Alaa AlTabishi, MD, PhD (St John of Jerusalem Eye Hospital Group, East Jerusalem, Palestine); Carmen Ayuso, MD, PhD (Department of Genetics, IIS-Fundación Jiménez Díaz, CIBERER, Madrid, Spain); Sandro Banfi, MD (Department of Precision Medicine, University of Campania Luigi Vanvitelli, Naples and Telethon Institute of Genetics and Medicine [TIGEM], Pozzuoli, Italy); Tamar Ben-Yosef, PhD (Ruth and Bruce Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa, Israel); L. Ingeborgh van den Born, MD, PhD (The Rotterdam Eye Hospital and The Rotterdam Ophthalmic Institute, Rotterdam, The Netherlands); Ana Fakin, MD, PhD (Eye Hospital, University Medical Centre Ljubljana, Slovenia); G. Jane Farrar, PhD (The School of Genetics & Microbiology, Trinity College Dublin, Dublin, Ireland); Juliana Maria Ferraz Sallum, MD, PhD (Universidade Federal de São Paulo and Instituto de Genética Ocular, São Paulo, SP, Brazil); Kaoru Fujinami, MD, PhD (UCL Institute of Ophthalmology and Moorfields Eye Hospital, London, UK; Laboratory of Visual Physiology, Division of Vision Research, National Institute of Sensory Organs, National Hospital Organization Tokyo Medical Center, Tokyo, Japan; Graduate School of Health Management, Keio University, Tokyo, Japan); Michael B. Gorin, MD, PhD (Department of Ophthalmology and Department of Human Genetics, David Geffen School of Medicine, Stein Eye Institute, UCLA [University of California, Los Angeles], USA); Lucia Hlavata, MD (Research Unit for Rare Diseases, Department of Pediatrics and Adolescent Medicine, First Faculty of Medicine, Charles University and General University Hospital in Prague, Prague, Czech Republic); Smaragda Kamakari, PhD (Ophthalmic Genetics Unit, OMMA Ophthalmological Institute of Athens, Athens, Greece); Bohdan Kousal, MD (Research Unit for Rare Diseases of the Department of Pediatrics and Adolescent Medicine and Department of Ophthalmology, First Faculty of Medicine, Charles University and General University Hospital in Prague, Prague, Czech Republic); Ian M. MacDonald, MD, CM (Departments of Ophthalmology and Medical Genetics, University of Alberta, Edmonton, Alberta, Canada); Terri L. McLaren, BSc (Centre for Ophthalmology and Visual Science, The University of Western Australia, Nedlands, Western Australia, Australia; Australian Inherited Retinal Disease Registry and DNA Bank, Department of Medical Technology and Physics, Sir Charles Gairdner Hospital, Nedlands, Western Australia, Australia); Anna Matynia, PhD (Departments of Ophthalmology and Human Genetics, David Geffen School of Medicine, Stein Eye Institute, UCLA, USA); Monika Oldak, MD, PhD (Department of Histology and Embryology, Medical University of Warsaw, Warsaw, Poland); Osvaldo L. Podhajcer, PhD (Laboratory of Molecular and Cellular Therapy, Fundacion Instituto Leloir-CONICET, Buenos Aires, Argentina); Raj Ramesar, MBA, PhD (MRC Genomic and Precision Medicine Research Unit, Division of Human Genetics, Department of Pathology, Institute of Infectious Disease and Molecular Medicine [IDM]), Faculty of Health Sciences, University of Cape Town, Cape Town, South Africa); John N. De Roach, PhD (Centre for Ophthalmology and Visual Science, The University of Western Australia, Nedlands, Western Australia, Australia; Australian Inherited Retinal Disease Registry and DNA Bank, Department of Medical Technology and Physics, Sir Charles Gairdner Hospital, Nedlands, Western Australia, Australia); Dror Sharon, PhD (Department of Ophthalmology, Hadassah Medical Center, Faculty of Medicine, The Hebrew University of Jerusalem, Jerusalem, Israel); Francesca Simonelli, MD (Eye Clinic, Multidisciplinary Department of Medical, Surgical and Dental Sciences, University of Campania Luigi Vanvitelli, Naples, Italy); Francesco Testa, MD, PhD (Eye Clinic, Multidisciplinary Department of Medical, Surgical and Dental Sciences, University of Campania Luigi Vanvitelli, Naples, Italy); Jennifer A. Thompson, PhD (Australian Inherited Retinal Disease Registry and DNA Bank, Department of Medical Technology and Physics, Sir Charles Gairdner Hospital, Nedlands, Western Australia, Australia); Anna M. Tracewska, PhD (Łukasiewicz Research Network–PORT Polish Center for Technology Development, Wroclaw, Poland); Andrea L. Vincent, MBChB, MD, (Department of Ophthalmology, New Zealand National Eye Centre, Faculty of Medical and Health Sciences, The University of Auckland, Grafton, Auckland, New Zealand; Eye Department, Greenlane Clinical Centre, Auckland District Health Board, Auckland, New Zealand); and Bernhard H.F. Weber, PhD (Institute of Human Genetics, University of Regensburg, and Institute of Clinical Human Genetics, University Hospital Regensburg, Regensburg, Germany).

    Additional Information: The following institutional research ethics committees provided approval: Research and Ethics Committee of the Hospital de Pediatría Professor Dr Juan P. Garrahan and by the Bioethics Committee of Fundación Instituto Leloir, Buenos Aires, Argentina; Sir Charles Gairdner Hospital Human Research Ethics Committee, Australia; Research Ethics Committee of UNIFESP, Brazil; Health Research Ethics Board, University of Alberta, Edmonton, Canada; Ethics Committee of the General University Hospital, Prague, Czech; In France and Germany, informed consents were obtained in accordance with the French bioethics law and the German Genetic Diagnostics Act; Human Subjects Review Committee at the University Hospital of Heraklion, Crete, Greece; Royal Victoria Eye and Ear Hospital, Ireland; Hadassah Medical Center Institutional Review Board, Israel; the Israeli Health Ministry and local institutional review boards at Bnai Zion (Haifa), Sourasky (Tel Aviv), and Rambam (Haifa) Medical Centers, Israel; Ethics Committee of the Azienda Ospediaiera Universitaria della Seconda Università di Napoli, Italy; Institutional Review Board of National Hospital Organization Tokyo Medical Center, Tokyo, Japan; Ethics Committee of the Erasmus Medical Center, Rotterdam, the Netherlands; Ministry of Health, Health and Disability Ethic Committee, and Auckland District Health Board, New Zealand; St John Eye Hospital Ethics Committee, Palestine; Bioethics Committee at the Medical University of Warsaw, Wroclaw Medical University Bioethics Committee, and the Bioethics Committee at the Children's Memorial Health Institute, Poland; Republic of Slovenia National Medical Ethics Committee, Ministry of Health, Ljubljana, Slovenia; Human Research Ethics Committee, University of Cape Town, South Africa; Health Research Fundacion Jimenez Diaz University Hospital, Spain; and Institutional Review Board of UCLA, Los Angeles, California.

    References
    1.
    Blacharski  P. Fundus flavimaculatus. In: Newsome  DA, ed.  Retinal Dystrophies and Degenerations. Vol 135-159. Raven Press; 1988.
    2.
    Allikmets  R, Singh  N, Sun  H,  et al.  A photoreceptor cell-specific ATP-binding transporter gene (ABCR) is mutated in recessive Stargardt macular dystrophy.   Nat Genet. 1997;15(3):236-246. doi:10.1038/ng0397-236 PubMedGoogle ScholarCrossref
    3.
    Rotenstreich  Y, Fishman  GA, Anderson  RJ.  Visual acuity loss and clinical observations in a large series of patients with Stargardt disease.   Ophthalmology. 2003;110(6):1151-1158. doi:10.1016/S0161-6420(03)00333-6 PubMedGoogle ScholarCrossref
    4.
    Fujinami  K, Zernant  J, Chana  RK,  et al.  Clinical and molecular characteristics of childhood-onset Stargardt disease.   Ophthalmology. 2015;122(2):326-334. doi:10.1016/j.ophtha.2014.08.012 PubMedGoogle ScholarCrossref
    5.
    Lambertus  S, van Huet  RA, Bax  NM,  et al.  Early-onset Stargardt disease: phenotypic and genotypic characteristics.   Ophthalmology. 2015;122(2):335-344. doi:10.1016/j.ophtha.2014.08.032 PubMedGoogle ScholarCrossref
    6.
    Westeneng-van Haaften  SC, Boon  CJ, Cremers  FP, Hoefsloot  LH, den Hollander  AI, Hoyng  CB.  Clinical and genetic characteristics of late-onset Stargardt’s disease.   Ophthalmology. 2012;119(6):1199-1210. doi:10.1016/j.ophtha.2012.01.005 PubMedGoogle ScholarCrossref
    7.
    Burke  TR, Tsang  SH, Zernant  J, Smith  RT, Allikmets  R.  Familial discordance in Stargardt disease.   Mol Vis. 2012;18:227-233.PubMedGoogle Scholar
    8.
    Runhart  EH, Valkenburg  D, Cornelis  SS,  et al.  Late-onset Stargardt disease due to mild, deep-intronic ABCA4 alleles.   Invest Ophthalmol Vis Sci. 2019;60(13):4249-4256. doi:10.1167/iovs.19-27524 PubMedGoogle ScholarCrossref
    9.
    Valkenburg  D, Runhart  EH, Bax  NM,  et al.  Highly variable disease courses in siblings with Stargardt disease.   Ophthalmology. 2019;126(12):1712-1721. doi:10.1016/j.ophtha.2019.07.010 PubMedGoogle ScholarCrossref
    10.
    Zernant  J, Lee  W, Collison  FT,  et al.  Frequent hypomorphic alleles account for a significant fraction of ABCA4 disease and distinguish it from age-related macular degeneration.   J Med Genet. 2017;54(6):404-412. doi:10.1136/jmedgenet-2017-104540 PubMedGoogle ScholarCrossref
    11.
    Runhart  EH, Sangermano  R, Cornelis  SS,  et al.  The common ABCA4 Variant p.Asn1868Ile shows nonpenetrance and variable expression of Stargardt disease when present in trans with severe variants.   Invest Ophthalmol Vis Sci. 2018;59(8):3220-3231. doi:10.1167/iovs.18-23881 PubMedGoogle ScholarCrossref
    12.
    Zernant  J, Lee  W, Nagasaki  T,  et al.  Extremely hypomorphic and severe deep intronic variants in the ABCA4 locus result in varying Stargardt disease phenotypes.   Cold Spring Harb Mol Case Stud. 2018;4(4):a002733. doi:10.1101/mcs.a002733 PubMedGoogle Scholar
    13.
    Cornelis  SS, Bax  NM, Zernant  J,  et al.  In silico functional meta-analysis of 5,962 ABCA4 variants in 3,928 retinal dystrophy cases.   Hum Mutat. 2017;38(4):400-408. doi:10.1002/humu.23165 PubMedGoogle ScholarCrossref
    14.
    Sangermano  R, Garanto  A, Khan  M,  et al.  Deep-intronic ABCA4 variants explain missing heritability in Stargardt disease and allow correction of splice defects by antisense oligonucleotides.   Genet Med. 2019;21(8):1751-1760. doi:10.1038/s41436-018-0414-9 PubMedGoogle ScholarCrossref
    15.
    Cremers  FPM, Cornelis  SS, Runhart  EH, Astuti  GDN.  Author response: penetrance of the ABCA4 p.Asn1868Ile allele in Stargardt disease.   Invest Ophthalmol Vis Sci. 2018;59(13):5566-5568. doi:10.1167/iovs.18-25944 PubMedGoogle ScholarCrossref
    16.
    Cremers  FPM, Lee  W, Collin  RWJ, Allikmets  R.  Clinical spectrum, genetic complexity and therapeutic approaches for retinal disease caused by ABCA4 mutations.   Prog Retin Eye Res. 2020;100861. doi:10.1016/j.preteyeres.2020.100861 PubMedGoogle Scholar
    17.
    Khan  M, Cornelis  SS, Khan  MI,  et al.  Cost-effective molecular inversion probe-based ABCA4 sequencing reveals deep-intronic variants in Stargardt disease.   Hum Mutat. 2019;40(10):1749-1759. doi:10.1002/humu.23787 PubMedGoogle ScholarCrossref
    18.
    Khan  M, Cornelis  SS, Pozo-Valero  MD,  et al.  Resolving the dark matter of ABCA4 for 1054 Stargardt disease probands through integrated genomics and transcriptomics.   Genet Med. 2020;22(7):1235-1246. doi:10.1038/s41436-020-0787-4 PubMedGoogle ScholarCrossref
    19.
    World Medical Association.  World Medical Association Declaration of Helsinki: ethical principles for medical research involving human subjects.   JAMA. 2013;310(20):2191-2194. doi:10.1001/jama.2013.281053PubMedGoogle ScholarCrossref
    20.
    Karczewski  KJ, Francioli  LC, Tiao  G,  et al; Genome Aggregation Database Consortium.  The mutational constraint spectrum quantified from variation in 141,456 humans.   Nature. 2020;581(7809):434-443. doi:10.1038/s41586-020-2308-7PubMedGoogle ScholarCrossref
    21.
    Sangermano  R, Khan  M, Cornelis  SS,  et al.  ABCA4 midigenes reveal the full splice spectrum of all reported noncanonical splice site variants in Stargardt disease.   Genome Res. 2018;28(1):100-110. doi:10.1101/gr.226621.117 PubMedGoogle ScholarCrossref
    22.
    Pisano  A, Preziuso  C, Iommarini  L,  et al.  Targeting estrogen receptor β as preventive therapeutic strategy for Leber’s hereditary optic neuropathy.   Hum Mol Genet. 2015;24(24):6921-6931. doi:10.1093/hmg/ddv396 PubMedGoogle Scholar
    23.
    Oostra  RJ, Kemp  S, Bolhuis  PA, Bleeker-Wagemakers  EM.  No evidence for ‘skewed’ inactivation of the X-chromosome as cause of Leber’s hereditary optic neuropathy in female carriers.   Hum Genet. 1996;97(4):500-505. doi:10.1007/BF02267075 PubMedGoogle ScholarCrossref
    24.
    Cremers  FP, van de Pol  DJ, van Driel  M,  et al.  Autosomal recessive retinitis pigmentosa and cone-rod dystrophy caused by splice site mutations in the Stargardt’s disease gene ABCR.   Hum Mol Genet. 1998;7(3):355-362. doi:10.1093/hmg/7.3.355 PubMedGoogle ScholarCrossref
    25.
    van Driel  MA, Maugeri  A, Klevering  BJ, Hoyng  CB, Cremers  FP.  ABCR unites what ophthalmologists divide(s).   Ophthalmic Genet. 1998;19(3):117-122. doi:10.1076/opge.19.3.117.2187 PubMedGoogle ScholarCrossref
    26.
    Maugeri  A, van Driel  MA, van de Pol  DJ,  et al.  The 2588G→C mutation in the ABCR gene is a mild frequent founder mutation in the Western European population and allows the classification of ABCR mutations in patients with Stargardt disease.   Am J Hum Genet. 1999;64(4):1024-1035. doi:10.1086/302323 PubMedGoogle ScholarCrossref
    27.
    Schulz  HL, Grassmann  F, Kellner  U,  et al.  Mutation spectrum of the ABCA4 gene in 335 Stargardt disease patients from a multicenter German cohort - impact of selected deep intronic variants and common SNPs.   Invest Ophthalmol Vis Sci. 2017;58(1):394-403. doi:10.1167/iovs.16-19936 PubMedGoogle ScholarCrossref
    28.
    Franconi  F, Campesi  I, Colombo  D, Antonini  P.  Sex-gender variable: methodological recommendations for increasing scientific value of clinical studies.   Cells. 2019;8(5):E476. doi:10.3390/cells8050476 PubMedGoogle Scholar
    29.
    Gerdts  E, Regitz-Zagrosek  V.  Sex differences in cardiometabolic disorders.   Nat Med. 2019;25(11):1657-1666. doi:10.1038/s41591-019-0643-8 PubMedGoogle ScholarCrossref
    30.
    De Bellis  A, De Angelis  G, Fabris  E, Cannatà  A, Merlo  M, Sinagra  G.  Gender-related differences in heart failure: beyond the “one-size-fits-all” paradigm.   Heart Fail Rev. 2020;25(2):245-255. doi:10.1007/s10741-019-09824-y PubMedGoogle ScholarCrossref
    31.
    Billi  AC, Kahlenberg  JM, Gudjonsson  JE.  Sex bias in autoimmunity.   Curr Opin Rheumatol. 2019;31(1):53-61.PubMedGoogle ScholarCrossref
    32.
    Ventura-Clapier  R, Moulin  M, Piquereau  J,  et al.  Mitochondria: a central target for sex differences in pathologies.   Clin Sci (Lond). 2017;131(9):803-822. doi:10.1042/CS20160485 PubMedGoogle ScholarCrossref
    33.
    Sampathkumar  NK, Bravo  JI, Chen  Y,  et al.  Widespread sex dimorphism in aging and age-related diseases.   Hum Genet. 2020;139(3):333-356. doi:10.1007/s00439-019-02082-w PubMedGoogle ScholarCrossref
    34.
    Wickham  LA, Gao  J, Toda  I, Rocha  EM, Ono  M, Sullivan  DA.  Identification of androgen, estrogen and progesterone receptor mRNAs in the eye.   Acta Ophthalmol Scand. 2000;78(2):146-153. doi:10.1034/j.1600-0420.2000.078002146.x PubMedGoogle ScholarCrossref
    35.
    Marin-Castaño  ME, Elliot  SJ, Potier  M,  et al.  Regulation of estrogen receptors and MMP-2 expression by estrogens in human retinal pigment epithelium.   Invest Ophthalmol Vis Sci. 2003;44(1):50-59. doi:10.1167/iovs.01-1276 PubMedGoogle ScholarCrossref
    36.
    Hulsman  CA, Westendorp  IC, Ramrattan  RS,  et al.  Is open-angle glaucoma associated with early menopause? The Rotterdam Study.   Am J Epidemiol. 2001;154(2):138-144. doi:10.1093/aje/154.2.138 PubMedGoogle ScholarCrossref
    37.
    Prokai-Tatrai  K, Xin  H, Nguyen  V,  et al.  17β-estradiol eye drops protect the retinal ganglion cell layer and preserve visual function in an in vivo model of glaucoma.   Mol Pharm. 2013;10(8):3253-3261. doi:10.1021/mp400313u PubMedGoogle ScholarCrossref
    38.
    Elliot  SJ, Catanuto  P, Espinosa-Heidmann  DG,  et al.  Estrogen receptor beta protects against in vivo injury in RPE cells.   Exp Eye Res. 2010;90(1):10-16. doi:10.1016/j.exer.2009.09.001 PubMedGoogle ScholarCrossref
    39.
    Du  M, Mangold  CA, Bixler  GV,  et al.  Retinal gene expression responses to aging are sexually divergent.   Mol Vis. 2017;23:707-717.PubMedGoogle Scholar
    40.
    Li  B, Gografe  S, Munchow  A, Lopez-Toledano  M, Pan  ZH, Shen  W.  Sex-related differences in the progressive retinal degeneration of the rd10 mouse.   Exp Eye Res. 2019;187:107773. doi:10.1016/j.exer.2019.107773 PubMedGoogle Scholar
    41.
    Guarneri  R, Russo  D, Cascio  C,  et al.  Retinal oxidation, apoptosis and age- and sex-differences in the mnd mutant mouse, a model of neuronal ceroid lipofuscinosis.   Brain Res. 2004;1014(1-2):209-220. doi:10.1016/j.brainres.2004.04.040 PubMedGoogle ScholarCrossref
    42.
    Poloschek  CM, Bach  M, Lagrèze  WA,  et al.  ABCA4 and ROM1: implications for modification of the PRPH2-associated macular dystrophy phenotype.   Invest Ophthalmol Vis Sci. 2010;51(8):4253-4265. doi:10.1167/iovs.09-4655 PubMedGoogle ScholarCrossref
    43.
    Lee  W, Paavo  M, Zernant  J,  et al.  Modification of the PROM1 disease phenotype by a mutation in ABCA4.   Ophthalmic Genet. 2019;40(4):369-375. doi:10.1080/13816810.2019.1660382 PubMedGoogle ScholarCrossref
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