Autofluorescence in female carriers with choroideremia: A familial case with a novel mutation in the CHM gene

Choroideremia is an X-linked chorioretinal dystrophy caused by mutations in the CHM gene. The main differential diagnosis is X-linked retinitis pigmentosa. Clinically, male patients that are affected by these two diseases have similar symptoms. This work aims to report a familial case of choroideremia initially diagnosed as X-linked retinitis pigmentosa with a novel mutation in the CHM gene, and the relevance of fundus autofluorescence (FAF) in female carriers. A complete ophthalmological evaluation was done in a 37-year-old woman and her 53-year-old maternal uncle; the uncle had been diagnosed previously with X-linked retinitis pigmentosa. A visual field test, FAF imaging, full-field electroretinography, and a genetic test were performed. In the proband, the fundoscopy revealed diffuse changes in the retinal pigment epithelium in both eyes, and the FAF showed a speckled pattern of low- and high-density. The maternal uncle's ophthalmological evaluation showed choroidal and retinal atrophy consistent with choroideremia. The molecular analysis revealed a pathogenic variant in the CHM gene, c.190–1 G > T. In female carriers of choroideremia and X-linked retinitis pigmentosa, differential diagnosis may be challenging. A speckled pattern of low- and high-density in autofluorescence is commonly found in female carriers of choroideremia. FAF is a powerful tool for making a correct clinical diagnosis because the pattern in FAF is much more apparent than the visible retinal changes obtained by fundoscopy. Although it is crucial to perform molecular analysis to confirm the diagnosis, FAF is useful when genetic testing may not be readily available.

Keywords: X-linked retinitis pigmentosa; choroideremia; fundus autofluorescence; CHM gene

Choroideremia is a rare chorioretinal dystrophy with an incidence ranging from 1:50,000 to 1:100,000; it is characterized by progressive atrophy of the retina and the choroid ([1],[2]). It has an X-linked inheritance pattern that is caused by mutations in the CHM gene located in the Xq21.2 chromosome ([3]). Retinitis pigmentosa (RP) is the most common form of retinal dystrophy, with a worldwide prevalence of approximately 1:4,000 ([4]). The genetic basis of RP is highly heterogenous. X-linked retinitis pigmentosa (XLRP) is the most severe subtype; it represents 10–20% of all RP cases. In XLRP, mutations in the RPGR gene are found in most cases (30% to 80%) ([5]). Both diseases, choroideremia and RP, share clinical characteristics, such as the progressive reduction of visual acuity, nyctalopia, and the progressive loss of the visual field ([6],[7]). Because both conditions also share the X-linked inheritance pattern, clinical diagnosis may be confused ([[8]]). It is also important to consider the clinical differences between female carriers of choroideremia and XLRP. Most female carriers of choroideremia retain good visual function throughout their life, and significant visual impairment is uncommon. Usually, they either do not have any symptoms or they only have mild to moderate night blindness ([[11]]). Instead, female carriers of XLRP show a wide spectrum of clinical features, ranging from mild or moderate abnormalities to severe disease ([14]). In 2015, Comander et al. reported that up to 2% of female XLRP carriers may become legally blind ([15]).

Here, we report a familial case of choroideremia that was initially misdiagnosed as XLRP. We emphasize the fundus autofluorescence (FAF) findings for clinical diagnosis in female carriers of choroideremia. Moreover, we detected a novel pathogenic variant in the CHM gene.

A 37-year-old woman presented for ophthalmic evaluation, complaining of difficult vision at night. She had been experiencing this symptom since the age of 19. Best-corrected visual acuity (BCVA) was 20/20 in both eyes, anterior segment examination was unremarkable, and fundus examination revealed diffuse changes in the retinal pigment epithelium (RPE) in both eyes (Figure 1a). The FAF imaging showed a speckled pattern of low- and high-density (Figure 1b). In accordance with the International Society for Clinical Electrophysiology of Vision (ISCEV) standard protocol, full-field electroretinography was performed, which showed reduced scotopic response with a b-wave decrease in amplitude; other responses were normal. The 30–2 visual fields showed no alterations.

PHOTO (COLOR): Figure 1. (a) Fundus photograph of the right eye of the proband with evidence of hypo- and hyperpigmented changes in the RPE; the optic nerve, the fovea, and the vessels are normal. (b) FAF imaging of the same eye that shows delimited areas of hypo-autofluorescence, mixed with areas of hyper-autofluorescence, an area of hypoautofluorescence is observed at the temporal edge of the optic nerve; at the level of the macula, an area of oval hypoautoflurescence is observed with a greater diameter in the horizontal direction. (c) Fundus photograph of the right eye of the proband's sister without abnormalities. (d) FAF imaging of the same eye without abnormalities

The patient's family medical history showed that she had a 53-year-old maternal uncle with poor vision and nyctalopia; he had been diagnosed with XLRP in childhood. The patient's maternal uncle was re-evaluated, and his fundus examination revealed pallor of the optic nerve, severe choroidal and retinal atrophy, attenuated vessels, visualization of the choroidal vasculature, and areas of total choroidal atrophy in both eyes making it possible to see the sclera (Figure 2). Notably, there were no areas of hyperpigmentation in the form of bone spicules.

PHOTO (COLOR): Figure 2. Fundus photographs of the proband's uncle, right (a) and left eye (b). There are areas of RPE atrophy and choroid that allow sclera to be seen around the papilla predominating in the lower sector. In this area, some large remaining choroid vessels, yellow in color, suggest choroidal sclerosis. Outside these regions, areas of hyperpigmentation with poorly defined edges, which are characteristic of choroideremia, are observed

The proband's mother was asymptomatic, and the fundus examination revealed diffuse changes in the RPE in both eyes. The proband's sister was asymptomatic, and no fundoscopy changes were identified (Figure 1c,d). Other family members had no visual problems.

In the proband, the speckled pattern of low- and high-density in FAF was consistent with the findings commonly found in female carriers of choroideremia. In her maternal uncle, the characteristics of the retina were all suggestive of choroideremia instead of XLRP.

The molecular test adhered to the tenets of the Declaration of Helsinki. After obtaining approval from the Ethics Committee and informed consent from the patients, blood samples were drawn. Then, genomic DNA was extracted from the blood leukocytes according to standard procedures. Sanger sequencing of the entire CHM coding region was performed using the BigDye™ Terminator Cycle Sequencing Kit (Applied Biosystems, Foster City, CA, USA) following a protocol previously detailed ([16]). All samples were analyzed in a 3130 Genetic Analyzer (Applied Biosystems, Foster City, CA, USA).

A pathogenic c.190–1 G > T (intron 3–4) CHM variant was identified heterozygously in DNA from the proband and hemizygously in DNA from her uncle. This genetic variant has not been reported in the GnomAD, 1000 genomes, and LOVD genes databases. The pathogenic potential of the mutation was reviewed through analysis in Human Splicing Finder, Mutation Taster, and FATMM-MKL, where it is reported that the variant critically modifies the acceptor site for exon 4, causing deletion of the first 8 nucleotides of that exon. This genetic variant is considered to be pathogenic according to the American Genetic Counseling Guidelines standards ([17]).

In this case report, the symptoms of nyctalopia, RPE changes, and the FAF findings, combined with the family history and the pathogenic variant detected in the CHM gene, confirmed the diagnosis of choroideremia. Previously, the maternal uncle had an erroneous diagnosis of XLRP.

Male patients affected with choroideremia suffer from progressive symptoms of reduced visual acuity, nyctalopia, and loss of the peripheral visual field. In contrast, usually, female carriers are asymptomatic. In male patients, the symptomatology is similar for choroideremia and RP or other inherited retinal dystrophies. Approximately 6% of individuals diagnosed with RP might have choroideremia ([18]).

During the initial approach in patients with night blindness, probably affected by inherited retinal dystrophy, easily accessible tools can be used. FAF is considered to be due to lipofuscin deposits in the cells of the RPE ([18]). The different patterns of FAF in the central fundus depend on the stage and type of the inherited disease. Thus, FAF imaging is a non-invasive tool that can be helpful in the diagnosis of female carriers of an X-linked inherited retinal dystrophy. Usually, small changes in the lipofuscin content of RPE are not visible on ophthalmoscopy ([19]).

In FAF imaging, female carriers of choroideremia show a characteristic speckled pattern of low- and high-density; this finding is not seen in any other inherited retinal disease ([[19]]). Female carriers of XLRP also show changes in FAF patterns. In 2014, Wegscheider et al. described a radial pattern that has not been described in other inheritance patterns of RP ([[22]]). The difference in FAF patterns between these diseases makes FAF imaging a powerful clinical tool for achieving an accurate clinical diagnosis between X-linked chorioretinal diseases. Although genetic testing is becoming more accessible as a tool for a definitive diagnosis, FAF imaging has a particular value for directing it.

Genetic testing is important for patients with choroideremia because they are often misdiagnosed as having RP. The CHM gene is the only gene known to be associated with choroideremia; it encodes the geranylgeranyl transferase Rab escort protein-1 (REP-1). REP-1 deficiency affects intracellular vesicular trafficking, resulting in cellular dysfunction and premature cell death ([17]). According to The Human Gene Mutations Database (http://www.hgmd.cf.ac.uk/ac/index.php), more than 300 mutations in the CHM gene have been reported to be associated with choroideremia. Of these, the most frequent are point mutations (missense or nonsense mutations) that directly introduce premature stop codons. Furthermore, in this mutations database it has been reported that there are 40 mutations that affect the splicing site. In this study, we detected a novel mutation in a splice site in exon 4 of the CHM gene, causing deletion in the first 8 nucleotides of that exon.

In conclusion, to differentiate female carriers of XLRP from choroideremia, FAF imaging can detect a specific pattern that can aid in directing the clinical diagnosis. A molecular test will provide the patient and their relatives with adequate genetic counseling, including detection of female carriers. Moreover, the early and precise diagnosis of choroideremia is vitally important for enrolling patients in gene therapies that are currently being developed.

The authors report no conflicts of interest. The authors alone are responsible for the content and writing of this article.