Genetic Testing and Clinical Characterization of Patients with Cone–Rod Dystrophy

With interest we read the article by Littink et al., ¹ which was published online June 16, 2010, on genetic screening of 126 probands with cone–rod dystrophy (CRD) for mutations in the ABCA4 gene and mutations in homozygous regions by single-nucleotide polymorphism (SNP) array analysis. The authors found that 31 of 90 patients had mutations detected by APEX (arrayed primer extension) ABCA4 microarray, and eight probands revealed pathogenic mutations in six different genes after sequencing. An additional ophthalmic work-up of these eight patients showed that two (25%) of the eight did not have CRD. The authors excluded these patients from their final calculations and concluded that 27% of the CRD patients had mutations in ABCA4, 1% in PROM1, 2% in CERKL, and 1% in EYS. The authors claimed that the revisions in diagnosis illustrate the complexity of diagnosing retinal dystrophies and the power and utility of molecular genetic testing. ¹  

We question the accuracy of the gene frequencies provided by the authors. First, the diagnosis of the CRD patients who did not receive the additional clinical work-up is uncertain. If two (25%) of eight of the patients who were thoroughly examined after genetic screening did not have CRD, it could mean that the diagnosis in the remaining 118 probands was false in up to 30 persons. The initial CRD diagnosis was made by different ophthalmologists who sent DNA to the laboratory at various stages of the diagnostic process. The laboratory was ignorant of the stage. Some ophthalmologists had drawn blood at the patient's first visit when tests such as ERG, color vision, and other psychophysical tests still had to be performed; others had obtained a blood sample after a definitive diagnosis was established. The great uncertainty in diagnosis does not justify the calculation of gene frequencies in this study group, and the frequencies are not representative of CRD. 

Second, the methodology of genetic testing was not identical in the entire study group. Of the initial probands, 90 (71%) were tested for ABCA4 mutations by APEX microarray, and 95 (75%) were evaluated by one of two SNP arrays (250 K and 6.0; Affymetrix, Santa Clara, CA). Although the SNP arrays enabled detection of ABCA4 mutations in the six largest homozygous regions, homozygous mutations in smaller regions as well as compound heterozygous ABCA4 mutations could never have been detected. Therefore, the authors should have used 90 as the denominator in their frequency calculations of ABCA4. Likewise, frequency calculations of PROM1, CERKL, and EYS should be limited to the group of patients who were screened for these genes. 

To gain insight into the disease pathogenesis, to help guide the diagnostic process in the clinic, and to direct focus for research funding, good genetic epidemiologic data on this incapacitating eye disease are necessary. There is a clear need for unbiased research with uniform inclusion criteria using a gold standard for diagnosis, complete testing of the total study group, and proper statistical analysis. ^2,3 To make this research feasible, good collaboration between clinicians, molecular geneticists, and epidemiologists is warranted.