Autosomal Dominant Stargardt-like Macular Dystrophy: Founder Effect and Reassessment of Genetic Heterogeneity

Objectives  To characterize a disease-associated haplotype in 7 families with autosomal dominant Stargardt-like macular dystrophy and to determine whether these families share a common ancestor.

Methods  Twenty-five polymorphic DNA markers spanning known dominant Stargardt-like gene loci were used to determine the haplotype associated with disease. In addition, an extensive genealogical investigation searching for a common ancestor shared by all of the 7 families was performed.

Results  We clinically evaluated 171 patients and genotyped 145 samples. The same DNA haplotype on chromosome 6q16 was shared by all evaluated affected members within the 7 families. In addition, we were able to genealogically join all of the families into one larger family consisting of 31 branches and 2314 individuals. Twenty-seven branches have known living descendants, with 7 branches having affected family members. In addition, we refined the critical region for the gene to approximately 1000 kilobases (kb) and eliminated part or all of 9 candidate disease-causing genes.

Conclusions  Our study indicates that most reported cases of autosomal dominant Stargardt-like macular dystrophy in North America are part of a single larger family associated with a gene locus on chromosome 6q16. Furthermore, the DNA haplotype associated with disease is useful in excluding individuals with phenotypically similar retinal conditions.

Clinical Relevance  The disease-associated haplotype allows for more accurate genetic counseling to be given to individuals with a Stargardt-like phenotype inherited in an autosomal dominant pattern.

FAMILIES affected by rare hereditary diseases are often described independently and are usually unrelated. However, molecular genetic studies can determine whether such families share a related genomic DNA region containing the disease locus. Such findings imply that the disease actually arose in a common ancestor or founder. For example, Fingert and associates¹ found all 27 glaucoma families affected with the GLN386STOP mutation in the myocilin gene appeared to be related through a common ancestor even though they were identified in 4 different patient populations. Equally striking is the observation that all 39 families with radial drusen (malattia leventinese or Doyne honeycomb retinal dystrophy) share a single identical mutation in the EFEMP1 gene containing the same pattern of DNA sequence variation (haplotype).² Thus, the radial drusen mutation appears to have arisen once in an ancestor shared by all 39 families who lived on 3 different continents.

Autosomal dominant Stargardt-like macular dystrophy is another rare hereditary retinal disease reported as occurring independently in several families.^3-13 Clinically, the disease usually presents in the teenage years with decreased visual acuity and atrophy of the macular retinal pigment epithelium with or without surrounding subretinal flecks and progresses rapidly over several years to legal blindness. In 1980, Cibis et al⁸ described one such large family consisting of 98 at-risk members. Several other families with similar clinical features were subsequently described.^3,6,7,9 More recently, we described the clinical and genetic features of 4 large families living in the United States.^10-13 Two of these families were found to share a common set of DNA markers (disease-associated haplotype) in the disease gene region of chromosome 6 as well as paternal ancestors, raising the possibility that other families with dominant Stargardt-like dystrophies might be related.¹² In this study, we show that 7 affected families are part of a single, larger family consisting of more than 2000 individuals whose affected members share an identical disease-associated haplotype spanning the gene responsible for this condition located on chromosome 6q16.

The patients in this study comprise one single family. There are 31 branches of this family, with 7 branches (Figure 1) having affected members. Each of the 7 affected families were thought to be independent of one another before this study, and all were diagnosed as having autosomal dominant Stargardt-like macular dystrophy. We use the term family to refer to the descendants of the top generation of a pedigree known to the authors at the time they reported the pedigree. When independently identified families are found to be related through genetic analysis or genealogical investigation, we refer to the original families as branches of a new larger family.

One hundred seventy-one patients at risk for developing the disease were examined. Patients were considered affected if they showed progressive bilateral visual loss of early onset and if they had atrophic macular lesions as previously described.^10,12 In all cases, the disease status was determined before genotyping and examination. In cases where the patient had died, the disease status was inferred by clinical history or medical and/or governmental records. This study was approved by the institutional review board at each institution.

Family records were searched at the facilities of the Latter Day Saints Family History Center in Salt Lake City, Utah. Additional information was obtained from interviews with family members and from records from city, state, and federal agencies. Marriage, death, cemetery, census, hospital, and church records were also searched. Although more than 3500 family records were obtained, we include only families with direct links to the founder.

Genomic DNA was obtained from peripheral blood and extracted using standard techniques (QIAmp Blood MIDI kit; Qiagen, Inc, Santa Barbara, Calif). Of the 27 branches with living descendants, at least 1 sample was obtained from 16 branches. One hundred forty-five samples were genotyped. Genotypes at polymorphic short tandem repeat markers spanning the disease loci on chromosomes 6q16(13 markers) and 13q34 (12 markers) were determined in selected patients as previously described.^10-12 Haplotypes were constructed manually and/or by using the algorithm used in the GENEHUNTER software package.¹⁴ The disease-associated haplotype is that set of 13 DNA markers on chromosome 6, which segregates in association with dominant Stargardt-like macular dystrophy. The disease penetrance was estimated from the age of disease onset. The penetrances used were as follows: age 0 to 10 years, 0.62; age 11 to 20 years, 0.90; age 21 years or older, 0.99; with a disease allele frequency of 0.000001. Two-point linkage analysis was performed in selected members (60 family members) of branches 13, 14, and 30 using 3 chromosome 6 (D6S286, D6S460, and D6S1609) and 3 chromosome 13 (D13S158, D13S173, and D13S280) markers using the methods as previously described.¹⁰

The phenotypic appearance and the disease-associated haplotype were determined in these 7 apparently unrelated families (Table 1). The disease-associated haplo-type was useful in combining the families into one larger family, excluding families with this diagnosis from other families, and refining the chromosomal location of the gene responsible for this condition. The results correlated with the genealogical analysis as described herein.

The clinical course of disease and the phenotypic appearance of the fundus were similar in most patients within the 7 affected families, although some variations were observed (Figure 2).^10,12 Early disease was characterized by subfoveal atrophy of the retinal pigment epithelium with or without the presence of flecks. Later in the disease, the foveal lesions were more pronounced, often with a beaten-metal appearance. At this stage, most of the patients demonstrated subretinal flecks. Patients with late-stage disease often show diffuse geographic atrophy with or without flecks (Figure 2).

An identical chromosome 6q16 pattern of DNA markers (haplotype) segregated with the disease gene (Figure 3)in all affected members of all families studied (branches 5, 10, 13, 14, 20, 24, and 30) but not in any of the unaffected family members. The probability of 2 individuals sharing this same disease-associated haplotype by chance is highly unlikely (approximately 1 in 100 trillion).¹⁰ This result indicates that these 7 affected families (1237 total members) are genetically related through a common ancestor or founder. Nonaffected family members or other family members with juvenile-onset visual loss, including one case of a patient with a childhood intraocular inflammatory disease and one patient with foveal hypoplasia and nystagmus (Table 1; branches 1 and 22), did not share the haplotype.

It became apparent during our genealogical investigation (see the following section) that branch 30 represented a family that was previously described to exhibit linkage between the disease and markers on chromosome 13. Records were identified in this study showing ancestors from this branch lived on a farm adjacent to other family branches (13, 14, and 24) in the early 1800s, confirming the potential for marriage relationships. Maximal 2-point lod scores at a THETAS of 0.0 for the 3 chromosome 6 markers were 7.47, 12.24, and 7.49. Removing branch 30 reduced the lod scores to 5.77, 10.72, and 6.11, respectively. The 2-point lod scores at a THETAS of 0.0/0.1 for the 3 chromosome 13 markers were −27.91/−2.05, −4.88/0.97, and −6.03/0.20, demonstrating exclusion of the chromosome 13 interval. The lod scores without branch 30 were comparable. The chromosome 6 disease-associated haplotype was present in all affected family members of this branch studied.

Analysis of the recombinant individuals within our 7 families with affected individuals enabled us to refine the genomic location of the gene on chromosome 6q16. As illustrated in Figure 4, the critical region is estimated to be approximately 1000 kilobases (kb) using recombinant data from affected and unaffected individuals. The critical region using only affected individuals is also illustrated. Twenty-seven new short tandem repeat markers were developed using genomic sequence from the Sanger Centre (http://www.sanger.ac.uk/) to facilitate the refinement.

During refinement of the critical region, we screened coding sequence in 9 genes. The status of our screening using DNA sequencing of gene exons from 1 unaffected and 1 affected individual is shown in Table 2. At this time, we have excluded coding sequence variations in the human kinase gene (TTK), 3 of 4 exons in a novel protein similar to SH3BGR (75K24.1 in the Sanger database), an unnamed protein (complementary DNA accession AK000712) except for exon 8, and thyroid receptor interacting protein (TRIP7).

Having demonstrated that all of the families studied herein were genetically related through a common founder, we performed an extensive genealogical investigation to identify the relationships between the families. This investigation revealed that all of the families with disease reported herein can be traced to the marriage in 1789 between individuals III:14 and III:15 (Figure 1). This marriage resulted in 31 family branches (Figure 1), giving rise to a total of 2314 descendants (Table 1). The total number of family members ranged from 1 (branches 11, 12, 21, and 23) to 457(branch 24) members per branch (Table 1). Four of the 31 branches (11, 12, 21, and 23) did not give rise to any descendants. Two branches (1 and 22) gave rise to descendants with early-onset visual loss unrelated to macular dystrophy.

Seven branches, designated as 5, 10, 13, 14, 20, 24, and 30, have descendants with autosomal dominant Stargardt-like macular dystrophy (Table 1 and Figure 1). This includes approximately 200 affected individuals (approximately 170 known living). Branch 5 has not been described previously and consists of 66 members. Branches 13 and 14 consist of 238 and 179 members, respectively (Table 1). Branch 20 is composed of 37 members and has not been described previously. Branch 24 is composed of 457 individuals and represents the largest known branch. Branch 30 is composed of 157 members. One branch, designated branch 10, has no known living descendants with the condition. Affected members from this branch were previously evaluated (1964) at the University of Iowa Hospitals and Clinics.

Our study demonstrates that most families in the United States and Canada with members diagnosed as having autosomal dominant Stargardt-like macular dystrophy are related and comprise a single larger family. One of the unifying clinical features we observed independently was the similar phenotypic appearance and clinical course of this disorder among the various affected members of the 7 families.^10-13 These features included the early onset of progressive visual loss associated with bilateral foveal atrophy with or without fundus flecks. However, a dark choroid on fluorescein angiography was not a feature of this family as has been observed in a substantial fraction of patients with recessive Stargardt macular dystrophy. These relationships among independently described families were further demonstrated by a combination of molecular genetic and genealogical approaches.

A prominent molecular genetic feature of all 7 families was that they shared an identical DNA haplotype on chromosome 6. This result indicated that all 7 families descended from a common ancestor or founder. The chance of 2 individuals having this same disease-associated haplotype is extremely small. A family from which all of these families arose was subsequently identified genealogically.

Although the size of the family reported herein is large, we are aware of other families^9,15,16 that have been reported to link to chromosome 6. Based on our results, it is likely that some of these families may also be related to our family, resulting in an even larger family. This is further supported by our genealogical findings in that we only traced the descendants from one marriage (Figure 1; individuals III:14 and III:15). Since individuals III:14 and III:15 had a total of 16 brothers and sisters, it is likely that several additional families (of either the paternal or maternal line) gave rise to other descendants with this condition. A genealogic investigation of these descendants is currently in progress.

All reported linkage studies^10,12 on families with autosomal dominant Stargardt-like macular dystrophy, which are available to us, have localized the disease gene to chromosome 6. Zhang and associates¹¹ previously reported linkage to chromosome 13 in branch 30. Our results, based on genealogical and molecular genetic findings, indicate that this branch also is part of the family described herein. Ultimately, the genetic defect in this disorder will rely on the identification of the actual disease-causing gene and will help clarify this discrepancy.

Although the genetic defect has not been identified to date in this disorder, our results have narrowed the genetic interval for the disease-causing gene on chromosome 6 to approximately 1000 kb. Several candidate genes (Table 2) in this interval, including Col12A1 (collagen-associated gene), SSP1 (protein kinase gene), MYO6 (myosin 6 gene), TTK (human kinase gene), and TRIP7 (thyroid receptor interacting protein), were screened, and no mutations were identified that correlated with disease status.

Although the current population of the United States is relatively diverse, the early settlement of the country occurred in well-defined movements. One such movement, the Great Scottish-Irish Movement, occurred in the mid-1700s primarily through Pennsylvania before settling in stages to the west and south.¹⁷ All of the families identified in our study, including individuals III:14 and III:15 (Figure 1), appear to have originated from this Scottish-Irish movement. Furthermore, the 31 branches of this family appear to have settled in a relatively narrow region in the United States, with most of the known families living within the same or adjacent states.

In one branch (10), there were no living descendants with macular dystrophy to study. This finding indicates other additional families with this condition may have existed or have not been uncovered to date. Furthermore, it is not always possible to determine the disease status of early ancestors. This is true in our study as well. However, in some cases, it is possible to infer it from vital records. For example, 2 individuals were mustered out of the US Army during the Civil War and received pensions at ages 17 and 18 years because of blindness, implying they inherited the disease genotype.

The identification and characterization of this large family will be useful both clinically and in studies directed toward identifying the gene responsible for this disorder. The finding that many of these patients are related and share an identical disease-associated haplotype will also be useful in counseling families carrying the gene for this condition.

Accepted for publication December 14, 2000.

This study was supported in part by the Henry and Corinne Bower Laboratory for Macular Degeneration, Philadelphia, Pa; the Elizabeth C. King Trust, the estates of Margaret Mercer, Harry B. Wright, Reuben and Mollie Gordon Foundation, and Martha W. S. Rogers, and Research to Prevent Blindness Inc (RPB) (University of Texas Southwestern Medical Center, the University of Iowa, and Wills Eye Hospital); a career development award from RPB and the Foundation Fighting Blindness (Dr Edwards); the Association for Macular Diseases, Macular Degeneration International, the Kyle Curran Memorial Fund for Juvenile Macular Degeneration(Dr Weleber); Foundation Fighting Blindness; and National Institutes of Health grants EY11515 (Dr Hageman), EY10539 (Dr Stone), and EY12699 (Drs Edwards and Donoso). Dr Donoso is the Thomas D. Duane Professor of Ophthalmology, Wills Eye Hospital and Jefferson Medical College, Thomas Jefferson University.

Wallace McMeel, MD, provided helpful discussions and Kang Zhang, MD, provided a fundus photograph. Dale Drake and Robert Andrew provided genealogical assistance. The sequence data were produced by the human chromosome 6 sequencing group at the Sanger Centre, Wellcome Trust Genomic Centre, Hinxton, Cambridge, England.

Corresponding author and reprints: Albert O. Edwards, MD, PhD, Department of Ophthalmology, University of Texas Southwestern Medical Center, 5323 Harry Hines Blvd, Dallas, TX 75390-9057 (e-mail: Albert.Edwards@UTSouthwestern.edu).