Elsevier

European Journal of Medical Genetics

Volume 51, Issue 5, September–October 2008, Pages 417-425
European Journal of Medical Genetics

Original article
Exon copy number alterations of the CHD7 gene are not a major cause of CHARGE and CHARGE-like syndrome

https://doi.org/10.1016/j.ejmg.2008.03.003Get rights and content

Abstract

CHARGE syndrome is a multiple congenital anomaly syndrome caused by mutations in the CHD7 gene. Mutations in this gene are found in 60–70% of patients suspected of having CHARGE syndrome. However, if only typical CHARGE patients are taken into account, mutations in the CHD7 gene are found in over 90% of cases. The remaining 10% might be caused by hitherto undetected alterations of the CHD7 gene, including whole exon duplications and deletions that are missed by the currently used diagnostic procedures. Therefore we looked for these kinds of alterations by multiplex ligation-dependent probe amplification in 54 patients suspected of having CHARGE syndrome without a CHD7 mutation. In one patient a partial deletion of the CHD7 gene (exons 13–38) was identified, while in the other patients no abnormalities were found. The frequency of exon deletions in our cohort was 1.9% (1/54) and 5.6% (1/18) in all patients and in typical CHARGE patients, respectively. We conclude that exon copy number alterations of the CHD7 gene are not a major cause of CHARGE and CHARGE-like syndrome.

Introduction

In 2004 the underlying gene defect for CHARGE syndrome (the CHD7 gene, OMIM #214800) was identified on chromosome 8 (8q12.1) [16]. This multiple congenital anomaly syndrome was originally described as a combination of coloboma, heart defects, atresia of choanae, retardation of growth and/or development, genital hypoplasia and ear anomalies and/or deafness [12]. Later, additional congenital malformations were recognised, of which agenesis of the semicircular canals and arhinencephaly were found to be present in nearly all patients [1], [2], [13]. The prevalence is approximately 1 in 10,000. CHARGE syndrome is diagnosed on clinical grounds according to two different sets of criteria as proposed by Blake et al. [5] and Verloes [15], and/or analysis of the CHD7 (chromodomain helicase DNA binding protein 7) gene. Three large, independent studies found heterozygous mutations in the CHD7 gene in 60–70% of patients suspected of having CHARGE syndrome [3], [8], [10]. However, if the clinical criteria for CHARGE syndrome [5], [15] are strictly applied, mutations in CHD7 are present in over 90% of the typical CHARGE patients.

The two sets of criteria used for diagnosing CHARGE syndrome, consist of phenotypic features that do not fully overlap (Table 1). Blake et al. consider a patient as having typical CHARGE syndrome if four major, or three major and three minor, criteria are present [5]. Based on new diagnostic insights, Verloes refined the criteria and included abnormalities of the semicircular canals [15]. Besides, Verloes distinguished three categories of CHARGE syndrome patients: typical, partial and atypical. Verloes considered typical CHARGE patients to present with three major, or two major and two minor, criteria. Patients were considered to have partial CHARGE syndrome when two major and one minor criteria were present, while atypical CHARGE patients have two major and no minor criteria, or one major and two minor criteria.

Most of the CHD7 mutations that have been described in CHARGE syndrome are truncating (nonsense and frameshift), but missense mutations have also been found. Most mutations are unique, although some recurrent de novo mutations have been found. There seem to be no real mutation hot spots, but some exons (2, 3, 31 and 34) are more frequently mutated than others (own unpublished data). The higher mutation frequency of these exons appears to be related to their size, with the largest exon 2 most frequently mutated. Until now, only one intragenic deletion has been reported in a CHARGE patient [14]. Whole gene deletions of CHD7, although present in the two patients who contributed to the discovery of the CHD7 gene [16], were not found in two large cohorts of CHARGE patients [8], [10]. However, single exon deletion or duplication would have been missed with the techniques used in these studies. Since the underlying defect in the remaining 10% of typical CHARGE patients has not yet been discovered, we hypothesised that exon copy number alterations of the CHD7 gene might contribute to CHARGE syndrome. We identified a cohort of 54 patients (suspected of) having CHARGE syndrome who did not have a CHD7 mutation. We screened for single exon deletion or duplication in the CHD7 gene using multiplex ligation-dependent probe amplification (MLPA).

Section snippets

Materials and methods

Fifty-four patients were selected from a group of patients who were referred for mutation analysis of CHD7 because of clinical features suggestive of CHARGE syndrome. Mutation screening performed by polymerase chain reaction (PCR) followed by direct sequencing had not revealed any CHD7 alterations in these patients [8]. The DNA samples were subsequently screened for exon deletions and/or duplications of the CHD7 gene by MLPA analysis. Half of the patients (see Table 2) were analysed using

Results

In our cohort of 54 patients we found a deletion of exons 13–38 in patient no. 1 (Fig. 1). In all other patients no exon copy number changes in the CHD7 gene were found.

An overview of the clinical features of patient no. 1 is given in Table 2 (see also Fig. 2). The patient was born at 42 weeks of gestation with a birth weight of 3.685 kg. At birth right facial nerve palsy and dysmorphic ears were noted. In addition he had partial palsy of cranial nerves IX and X, for which he required tube

Discussion

A defect in the CHD7 gene is found in 60–70% of all patients suspected of having CHARGE syndrome and in over 90% of typical CHARGE patients [8]. In the remaining 10% of typical patients the cause of CHARGE syndrome remains elusive. Genetic heterogeneity could be present, but the only other gene that has been implicated in CHARGE syndrome, the SEMA3E gene, was found to be mutated in only one CHARGE patient [11]. So far, mutations in this gene have not been reported in other CHARGE patients. It

Acknowledgements

We thank Jackie Senior for her help in preparing this manuscript. Dr. Bergman was supported by a grant from the Netherlands Organisation for Health Research.

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