OPHTHALMIC MOLECULAR GENETICS Clinical and Functional Findings in Choroideremia Due to Complete Deletion of the CHM Gene Marco Mura, MD; Christina Sereda, BS; Monica M. Jablonski, PhD; Ian M. MacDonald, MD; Alessandro Iannaccone, MD, MS Objective: To report the clinical, functional, and in vivo illaris atrophy. The carrier mother had diffuse elevation microanatomic characteristics of a family with choroi- of 650-nm dark-adapted thresholds. deremia with a deletion of the entire gene that encodes for the Rab escort protein 1 (CHM). Conclusions: Deletion of the CHM gene causes severe choroideremia. Results of serial ERGs and fundus ex- Methods: We performed clinical examination, flash elec- aminations documented progression first of rod and then troretinography (ERG), light- and dark-adapted perim- of cone disease. Fundus appearance deteriorated rap- etry, and optical coherence tomography; reviewed medi- idly, in excess of the severity of the ERG decline. Opti- cal records; and obtained the medical history of the cal coherence tomography findings explained this ob- proband and 3 other family members. servation, at least in part. Results: At 4 years of age, the proband had a hypopig- Clinical Relevance: To our knowledge, this is the ear- mented fundus and retinal pigment epithelium mot- liest clinical, microanatomic, and ERG longitudinal tling, and dark-adapted ERGs were reduced. Severe reti- phenotypic documentation in molecularly character- nal pigment epithelium and choriocapillaris atrophy ized choroideremia and the first documentation of im- developed by 6 years of age, paralleled by a lesser ERG decline. Optical coherence tomography findings showed paired dark-adapted cone function in carriers. The pres- normal neural retinas overlying mild changes in the reti- ervation of the neural retina has mechanistic, prognostic, nal pigment epithelium and thinned neural retina with and therapeutic implications. impaired lamination, yet the neural retina was fairly pre- served over retinal pigment epithelium and choriocap- Arch Ophthalmol. 2007;125(8):1107-1113 HOROIDEREMIA IS A PRO- tions in molecularly characterized pa- gressive X-linked reces- tients; and the disease mechanisms remain sive retinal disease caused incompletely understood. by mutations in the CHM Herein we report the phenotype of gene, which encodes for members of a family resulting from the un- Cthe Rab escort protein 1 (REP-1).1 In af- common lack of the entire CHM gene. Spe- fected males, choroideremia is character- cifically, we had the opportunity to char- acterize disease expression and progression Author Affiliations: Retinal ized clinically by areas of scalloped atro- Degeneration and Ophthalmic phy of the retinal pigment epithelium in the proband in his early childhood years Genetics Service, Hamilton Eye (RPE), choriocapillaris (CC), and retina.2,3 by clinical criteria, functional testing, and in vivo imaging methods. We also con- Institute, University of Although the exact function and cellular ducted in-depth functional studies on an Tennessee Health Science localization of REP-1 in the eye remain in- Center, Memphis (Drs Mura, obligate carrier, documenting abnormali- Jablonski, and Iannaccone); and completely understood, REP-1 appears to ties of visual function in the dark- Department of Ophthalmology, be necessary in cellular processes as a regu- adapted state that, to our knowledge, had Ophthalmic Genetics lator of signal transduction, protein traf- not been previously reported in carriers. Laboratory, University of ficking, and endocytosis.4 Most CHM mu- Alberta, Edmonton (Ms Sereda tations result in a truncated REP-1 product and Dr MacDonald). Dr Mura is and include complete gene deletions.5,6 METHODS now with the Department of Ophthalmology, Academic Genotype-phenotype correlations re- Medical Center, University of main unclear, with instances of clinical The pedigree is illustrated in Figure 1. On re- 6 Amsterdam, Amsterdam, heterogeneity ; limited information is avail- ferral, the proband (individual V:1) did not have the Netherlands. able on the early phenotypic manifesta- visual symptoms. The family history was posi- (REPRINTED) ARCH OPHTHALMOL / VOL 125 (NO. 8), AUG 2007 WWW.ARCHOPHTHALMOL.COM 1107 ©2007 American Medical Association. All rights reserved. Downloaded From: https://jamanetwork.com/ on 09/29/2021 images from comparable retinal locations were obtained and analyzed from an age-matched control subject. I:1 I:2 The resulting images were analyzed manually with Na- tional Institutes of Health Image J software (version 1.36; http: //rsb.info.nih.gov/ij/). The correct scale was set by calibrating the software to the retinal thickness measured at the fovea by II:1 II:2 II:3 the OCT automated retinal thickness software (version 4.0.1; StratusOCT), which does not include the first hyperreflective band (shown in red in the false-color OCT maps) of approxi- III:1 III:2 III:3 III:4 III:5 III:6 III:7 III:8 mately 20 to 25 µm attributable to photoreceptor inner and outer segments. For the purpose of comparing our findings more di- rectly with those of Jacobson et al,11 measurements of total neu- ral retinal thickness were conducted at the fovea and at 1.3 mm IV:1 IV:2 IV:3 IV:4 IV:5 IV:6 of eccentricity from the fovea on magnified views of the im- ages over adjacent pixel columns at these locations. However, ? ? ? our total neural retinal thickness measurements included the V:1 V:2 V:3 V:4 V:5 V:6 first hyperreflective photoreceptor inner and outer segment band P but not the second one, which is instead attributable to the RPE. The thickness of the outermost hyporeflective band in the OCT Male Affected P Proband Monozygotic profile attributable to the outer nuclear layer (ONL) was also twins estimated. We performed identical measurements at 3.0 mm Female Unaffected Obligate Dizygotic temporal to the fovea, in an area of neural retina overlying a carrier twins large nummular area of RPE/CC atrophy. All measurements were Unknown Deceased Unknown Divorced sex status averaged and are expressed as mean±SD. Comparisons are given with our normal control image and the reference ranges that could be derived from Jacobson et al.11 Also, because in the lat- Figure 1. Pedigree of the family with choroideremia. ter study total retinal thickness measurements included the RPE peak,11 we enhanced comparability between their study and ours by reducing the published reference ranges by 30 µm, which tive for night blindness and a progressive X-linked recessive is our estimated average thickness of the RPE hyperreflective retinal dystrophy. In addition to the proband, 3 other subjects band and the intervening thin yellow-white band above it. (his mother [IV:2] and his 2 male siblings [V:2 and V:3]) were examined. The gathering of a medical history and a review of medical records were used to obtain information about the 5 MOLECULAR GENETICS males in generation III with advanced disease. After obtaining informed consent, a whole-blood sample was FUNCTIONAL TESTING collected by venipuncture from the proband, his 2 male sib- lings, and their mother. Genomic DNA was extracted with a Full-field flash electroretinograms (ERGs) were recorded with a commercially available kit (QIAamp 50 Maxi Kit; Qiagen Inc, digital system (Nicolet Spirit; Viasys Nicolet Biomedical, Madi- Valencia, California) according to the manufacturer’s specifi- son, Wisconsin), in response to flashes ranging from −2.1 to 1.0 cations. Molecular genetic analyses of the CHM gene were con- log candela (cd)·s/m2 in intensity, with monopolar contact lens ducted at the University of Alberta. The CHM primers were modi- electrodes (ERG Jet; Universo Plastique AS, Grenchen, Switzer- fied from van Bokhoven et al.5 Reactions were performed in total land) as reported previously.7 The International Society for Clini- volumes of 25 µL (2.5 µL 10ϫNEB buffer [New England Bio- cal Electrophysiology of Vision “standard flash”8 was 0.32 log cd·s/ labs, Ipswich, Massachusetts], 2.5 µL of 2mM deoxyribonucleo- m2. In individual V:1, responses were obtained with the person tide triphosphates, 0.125 µL bovine serum albumin, 60-ng for- under full anesthesia (intravenous propofol infusion). ward primer, 60-ng reverse primer, double-distilled water to Monochromatic automated perimetry was performed as pre- volume, 50-ng genomic DNA, and 1 U Taq [New England Bio- viously reported,7 based on the methods developed by Jacobson labs]). The cycling settings were as follows: 95°C for 5 min- et al9 and Apa´thy et al.10 In brief, red/green cone-mediated thresh- utes (95°C for 1 minute, ␹°C for 1 minute, and 72°C for 1 olds were determined in the light-adapted state (white standard minute)ϫ40 cycles, and 72°C for 7 minutes where the anneal- background) with a 600-nm (orange) stimulus. Dark-adapted ing temperature ␹ was 50°C (exons 2, 5B, 6, 7, 8, 9, 14, and thresholds were measured after 40 minutes of dark adaptation 15), 52°C (exons 3, 5A, 11, 12, and 13), and 58°C (exons 1, 4, in response to 650-nm (red) and 500-nm (blue-green) stimuli. and 10). Polymerase chain reactions (PCRs) were run on 1% agarose gels with a 1-kilobase (kb) pair DNA ladder. OPTICAL COHERENCE TOMOGRAPHY All research studies conducted in this investigation were approved by the institutional review boards of the participat- Retinal microanatomy was investigated in individual V:1 with ing institutions and were in compliance with the Declaration an optical coherence tomography (OCT) device (StratusOCT; of Helsinki. Carl Zeiss Meditec, Dublin, California). To retain maximal spa- tial resolution with each scan, 3 or four 6-mm images were ob- RESULTS tained along the horizontal meridian (each with a density of 512 A-scans), to include areas of clinically intact macular reti- nal RPE and CC and areas of ophthalmoscopically visible at- PROBAND (V:1) rophy.