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A case of mild CHARGE syndrome associated with a splice site mutation in CHD7
European Journal of Medical Genetics 59 (2016) 195e197
Contents lists available at ScienceDirect
European Journal of Medical Genetics journal homepage: http://www.elsevier.com/locate/ejmg
A case of mild CHARGE syndrome associated with a splice site mutation in CHD7 €l Garabedian b, Sylvette Wiener-Vacher c, Constance Wells a, Natalie Loundon b, Noe raldine Goudeffroye a, Tania Attie -Bitach a, e, Marie-Dominique Cordier-Bouvier d, Ge f, * Sandrine Marlin ^pital Necker-Enfants Malades, Paris, France Service d'Histologie, Embryologie et Cytog en etique, Ho ^pital Necker-Enfants Malades, Paris, France Service d'ORL, Ho ^pital Robert Debr Service d'ORL, Ho e, Paris, France d ^pital Trousseau, Paris, France Service de Radiologie, Ho e Universit e Paris Descartes, Sorbonne Paris-Cit e, Paris, France f ^pital Necker-Enfants Malades, Paris, France Centre de r ef erence des surdit es g en etiques, Service de G en etique, Institut Imagine, Ho a
b c
a r t i c l e i n f o
a b s t r a c t
Article history: Received 20 September 2015 Received in revised form 22 February 2016 Accepted 22 February 2016 Available online 24 February 2016
CHARGE syndrome (MIM#214800) (Coloboma, Heart defect, Atresia of choanae, Retarded growth and development, Genital hypoplasia, Ear abnormalities/deafness) is caused by heterozygous mutation of CHD7 transmitted in an autosomal dominant manner. In this report, we describe a patient with bilateral hearing impairment, unusually-shaped ears, no intellectual disability and a patent ductus arteriosus. Further investigation showed abnormal semicircular canals and the presence of olfactory bulbs. He does not fulfill the Blake or the Verloes criteria for CHARGE. A de novo mutation at the donor splice site of intron 33 was identified (c.7164 þ 1G > A). It is of importance to diagnose mildly affected patients for appropriate genetic counselling and to better understand the mild end of the phenotypic spectrum of CHARGE syndrome. © 2016 Elsevier Masson SAS. All rights reserved.
Keywords: CHARGE CHD7 Sensorineural hearing impairment Semicircular canal malformation
1. Introduction CHARGE syndrome (MIM#214800) is a multiorgan genetic autosomal dominant condition affecting from 1/15000 to 1/17000 births (van Ravenswaaij-Arts et al., 2015). CHARGE is an acronym for Coloboma, Heart defect, Atresia of choanae, Retarded growth and development, Genital hypoplasia, Ear abnomalities/deafness. It was first described in 1979 by Hall and Hittner in two separate reports. Heterozygous mutations in the Chromodomain Helicase DNA-binding protein 7 gene (CHD7, MIM#608892) located on 8q12, were found to be the main cause of CHARGE syndrome by Vissers et al., in 2004 (Vissers et al., 2004). Mutations are found in 60e70% of patients with CHARGE syndrome. Most mutations are truncating or frameshift mutations with a premature stop codon (Sanlaville and Verloes, 2007; Schulz et al., 2014) leading to haploinsufficiency. The diagnostic criteria were developed by Blake in
f e rence des surdite s ge ne tiques Institut * Corresponding author. Centre de re ^pital Necker-Enfants Malades, 149 Rue de Se vres, 75015 Paris, France. Imagine, Ho E-mail address: [email protected] (S. Marlin). http://dx.doi.org/10.1016/j.ejmg.2016.02.012 1769-7212/© 2016 Elsevier Masson SAS. All rights reserved.
1998, reviewed by Verloes in 2005, and updated in 2007 (Sanlaville and Verloes, 2007). The criteria classify patients into typical, incomplete and atypical CHARGE syndrome, depending on the number of abnormal phenotypic features. However, some patients with a CHD7 mutation only fulfill atypical CHARGE criteria or do not fulfill the criteria, with only one or two signs (Hughes et al., 2014; Jongmans et al., 2007). This case report adds to the understanding of the mild end of the phenotypic spectrum of CHD7 mutations. 2. Case report The patient is the second-born of three children of a nonconsanguineous couple. Family history was unremarkable. He was born after an uneventful pregnancy, with a birth weight of 3230 g (20th percentile), a birth length of 50 cm (30th percentile) and a head circumference of 35 cm (40th percentile). Apgar scores were 10/10. Mild feeding problems were noted as a new-born but he did not require medical intervention. He was diagnosed with profound bilateral hearing impairment at the age of 16 months. Clinical features at 2 ½ years old showed a
196
C. Wells et al. / European Journal of Medical Genetics 59 (2016) 195e197
square face with frontal bossing. The ears were mildly low-set, without ear lobes. He also had retractile testes and no micropenis. His growth parameters were on the 75th percentile for height, weight and head circumference. He walked at 18 months old and fell frequently until the age of 3. He had cochlear implants on the right ear at 3 and on the left ear at 4 years old. He had surgery to close a patent ductus arteriosus at 5 years old. Aged 8 years and 8 months (Fig. 1) he showed no significant intellectual disability, behaved normally and went to a regular school. He measured 1.35 m (þ1DS) and weighed 27 kg (þ1DS). He presented no clinical sign of endocrine deficiencies but has not yet reached the age of puberty. Neurological examination did not reveal any cranial nerve dysfunction. He did not complain of hyposmia and did not undergo olfactory testing. Clinical examination and functional vestibular tests (otolithic evoked response and caloric test) indicated a bilateral vestibular are flexia. CT-scan of the temporal bones showed normal cochlea and bilateral symmetrical vestibular abnormalities. The vestibules were dilated. There was a bilateral complete agenesis of the lateral semicircular canal and a rudimentary posterior semicircular canal (Fig. 2). Brain-MRI confirmed the vestibular abnormalities and showed that the cranial nerves VII and VIII were morphologically normal. The olfactory bulbs and olfactory sulci were normal. A renal ultrasound showed two normal-sized kidneys. Ophthalmological examination showed an intermittent divergent strabismus but no coloboma. The patient fulfills one major (Ear abnormality) with one minor criterion (Patent ductus arteriosus) according to the Blake criteria. According to the Verloes criteria (Bergman et al., 2011) he has one major criterion (hypoplasia of semicircular canals) and two minor criteria (heart malformation and sensorineural deafness), classifying him as atypical CHARGE. Because of the typical CHARGE inner ear malformation, CHD7 was screened on DNA extracted from a blood sample, by direct Sanger sequencing of the 37 coding exons and adjacent intronic regions (NM_017780.3) as previously described (Sanlaville and Verloes, 2007). A heterozygous mutation was found at the donor splice site of intron 33: c.7164 þ 1G > A, not reported in UCSC, ensembl, 1000 genome project or the exome variant server. All prediction tools (Alamut software using MaxENtSplice, HSF, NNsplice and Genesplicer) predict that this variant abolishes the donor splice site of intron 33. The most likely effect would be the skipping of exon 33 from the transcript (from c.6937_to c.7164) and absence of 76 aminoacids (Asp2313_to Lys2388) from the protein, however, an alternative splice site within intron 33 could also be used. The parents were also tested with DNA extracted from a blood sample and were shown not to carry the mutation. The
mutation was present in the patient's DNA extracted from urine and saliva with a balanced ratio, and therefore no argument for a somatic mosaicism.
3. Discussion We report a patient with mild CHARGE syndrome, with bilateral profound sensorineural deafness, typical inner ear malformation and a heart defect. He does not fulfill the Blake or the Verloes criteria for CHARGE. Splice site mutations as found in our patient represent around 11% of CHD7 mutations found in CHARGE patients (Sanlaville and Verloes, 2007). The molecular diagnosis of our patient highlights several points. Firstly, we were able to complete clinical examination with complementary examinations (e.g. renal ultrasound, brain MRI). Even though no additional abnormalities were found, the patient will be closely monitored for an early diagnosis of signs which might appear with time, such as hypogonadotrophic hypogonadism and puberty delay or scoliosis. Hypogonadism should be detected early in order to give hormone therapy to prevent osteoporosis. Secondly, an appropriate genetic counselling was given to his parents and will later be available to himself. The molecular diagnosis in our patient was made after the cochlear implantation. However, the presentation of the typical inner ear malformation at imaging before a cochlear implantation should prompt molecular testing before the implantation is performed in order to be informed of other features commonly seen in CHARGE syndrome that may hamper the implantation procedure, such as an aberrant route of the facial nerve and severe hypoplasia of the acoustic nerve, which are common in CHARGE syndrome. Typical CHARGE syndrome is well recognized, but the «mild» end of the spectrum is harder to identify. Those patients with only few symptoms can transmit their mutation and have offspring with a more severe phenotype. Because of the high phenotypic variability, it is important to diagnose such patients to give accurate genetic counselling. Tentative genotypeephenotype correlations have indicated that more severe forms are associated with truncating mutations (Legendre et al., 2012). A report has shown that missense mutations are more frequent in patients with Kallman syndrome due to a CHD7 mutation than in typical CHARGE patients (Marcos et al., 2014). We need to have more descriptions of «mild» CHARGE patients to understand the full spectrum of CHD7 mutations. The effects of the mutation of our patient have not been studied at the RNA and/or protein level. In silico, the splice mutation is predicted to result in an in-frame deletion of exon 33 and a shorter protein lacking 76 aminoacids without any known functional domain.
Fig. 1. Patient at 8 years 8 months. He has a square face, frontal bossing and no earlobes.
C. Wells et al. / European Journal of Medical Genetics 59 (2016) 195e197
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Fig. 2. Temporal CT-scan images. Complete agenesis of the lateral semicircular canal (b) compared to a control (a).
One of the strategies tested to identify mild CHARGE patients was to sequence CHD7 in patients with only one sign of CHARGE (Bergman et al., 2011). No CHD7 mutations were found in a cohort of atrio-ventricular defects or cono-troncal heart defects (CorstenJanssen et al., 2014). Bergman et al. suggested studying familial cases. There have been 18 families published so far, including a three-generational family (Hughes et al., 2014). Most patients with mild CHARGE were diagnosed after their severely affected child was diagnosed. Somatic mosaicism could also explain clinical variability (Jongmans et al., 2007, 2006). In our patient, three tissues were analysed and did not show any evidence of somatic mosaicism. In her guidelines for CHD7 analysis Bergman states that a temporal bone CT-scan should be performed if only 2 cardinal features or 1 cardinal and 1 supportive feature are present in a patient prior to CHD7 analysis, to evaluate the morphology of semicircular canals (Bergman et al., 2011). Following those guidelines our patient would have been diagnosed (one cardinal feature: abnormal functional vestibular test and one supportive: congenital heart defect). Along with these recommendations in CHARGE (Amiel et al., 2001), the importance of temporal bone imaging in sensorineural deafness should also be emphasized so as to evaluate and guide molecular analysis in patients with hearing impairment. Although the guideline for CHD7 testing presented by Bergman et al. has proven to be very sensitive (Hale et al., 2015), a diagnosis may still be missed in patients presenting with only one major or only one minor clinical feature. Next generation sequencing offers the possibility to diagnose these patients. 4. Conclusion The mild end of the phenotypic spectrum of CHD7 mutations is starting to emerge. We report here another sporadic case with mild CHARGE syndrome, with heart defect, sensorineural deafness and hypoplastic semi-circular canals. It should be emphasized that patients should not be rejected for CHD7 analysis if they do not fulfill criteria for atypical or typical CHARGE as there is a high intraand inter-familial variability. A larger screening of mild cases will be facilitated by the next-generation sequencing. References Amiel, J., Attie-Bitach, T., Marianowski, R., Cormier-Daire, V., Abadie, V., Bonnet, D., Gonzales, M., Chemouny, S., Brunelle, F., Munnich, A., Manach, Y., Lyonnet, S., 2001. Temporal bone anomaly proposed as a major criteria for diagnosis of CHARGE syndrome. Am. J. Med. Genet. 99, 124e127, 10.1002/10968628(20010301)99:2<124::AID-AJMG1114>3.0.CO;2e9.
Bergman, J.E.H., Janssen, N., Hoefsloot, L.H., Jongmans, M.C.J., Hofstra, R.M.W., van Ravenswaaij-Arts, C.M.A., 2011. CHD7 mutations and CHARGE syndrome: the clinical implications of an expanding phenotype. J. Med. Genet. 48, 334e342. http://dx.doi.org/10.1136/jmg.2010.087106. Corsten-Janssen, N., du Marchie Sarvaas, G.J., Kerstjens-Frederikse, W.S., Hoefsloot, L.H., van Beynum, I.M., Kapusta, L., van Ravenswaaij-Arts, C.M.A., 2014. CHD7 mutations are not a major cause of atrioventricular septal and conotruncal heart defects. Am. J. Med. Genet. Part A 164, 3003e3009. http:// dx.doi.org/10.1002/ajmg.a.36747. Hale, C.L., Niederriter, A.N., Green, G.E., Martin, D.M., 2015. Atypical phenotypes associated with pathogenic CHD7 variants and a proposal for broadening CHARGE syndrome clinical diagnostic criteria. Am. J. Med. Genet. Part A 1e11. http://dx.doi.org/10.1002/ajmg.a.37435. Hughes, S.S., Welsh, H.I., Safina, N.P., Bejaoui, K., Ardinger, H.H., 2014. Family history and clefting as major criteria for CHARGE syndrome. Am. J. Med. Genet. Part A 164, 48e53. http://dx.doi.org/10.1002/ajmg.a.36192. Jongmans, M.C.J., Admiraal, R.J., van der Donk, K.P., Vissers, L.E.L.M., Baas, A.F., Kapusta, L., van Hagen, J.M., Donnai, D., de Ravel, T.J., Veltman, J.A., Geurts van Kessel, A., De Vries, B.B.A., Brunner, H.G., Hoefsloot, L.H., van Ravenswaaij, C.M.A., 2006. CHARGE syndrome: the phenotypic spectrum of mutations in the CHD7 gene. J. Med. Genet. 43, 306e314. http://dx.doi.org/ 10.1136/jmg.2005.036061. Jongmans, M.C.J., Hoefsloot, L.H., van der Donk, K.P., Admiraal, Ronald J., Magee, A., van de Laar, I., Hendriks, Y., Verheij, J.B., Walpole, I., Brunner, H.G., Van Ravenswaaij, C.M.A., 2007. Familial CHARGE syndrome and the CHD7 gene: a recurrent missense mutation, intrafamilial recurrence and variability. Am. J. Med. Genet. A 143A, 2106e2112. http://dx.doi.org/10.1002/ajmg.a. Legendre, M., Gonzales, M., Goudefroye, G., Bilan, F., Parisot, P., Perez, M.-J., re, M., Bessie res, B., Martinovic, J., Delezoide, A.-L., Jossic, F., FalletBonnie Bianco, C., Bucourt, M., Tantau, J., Loget, P., Loeuillet, L., Laurent, N., Leroy, B., lin, C., Bazin, A., Alby, C., Ichkou, A., Salhi, H., Bigi, N., Rouleau, C., Guimiot, F., Que Gesny, R., Kitzis, A., Ville, Y., Lyonnet, S., Razavi, F., Gilbert-Dussardier, B., -Bitach, T., 2012. Antenatal spectrum of CHARGE syndrome Vekemans, M., Attie in 40 fetuses with CHD7 mutations. J. Med. Genet. 49, 698e707. http:// dx.doi.org/10.1136/jmedgenet-2012-100926. Marcos, S., Sarfati, J., Leroy, C., Fouveaut, C., Parent, P., Metz, C., Wolczynski, S., rard, M., Bieth, E., Kurtz, F., Verier-Mine, O., Perrin, L., Archambeaud, F., Ge Cabrol, S., Rodien, P., Hove, H., Prescott, T., Lacombe, D., Christin-Maitre, S., Touraine, P., Hieronimus, S., Dewailly, D., Young, J., Pugeat, M., Hardelin, J.P., , C., 2014. The prevalence of CHD7 missense versus truncating mutations is Dode higher in patients with Kallmann syndrome than in typical CHARGE patients. J. Clin. Endocrinol. Metab. 99, E2138eE2143. http://dx.doi.org/10.1210/jc.20142110. Sanlaville, D., Verloes, A., 2007. CHARGE syndrome: an update. Eur. J. Hum. Genet. 15, 389e399. http://dx.doi.org/10.1038/sj.ejhg.5201778. Schulz, Y., Wehner, P., Opitz, L., Salinas-Riester, G., Bongers, E.M.H.F., Van Ravenswaaij-Arts, C.M.A., Wincent, J., Schoumans, J., Kohlhase, J., Borchers, A., Pauli, S., 2014. CHD7, the gene mutated in CHARGE syndrome, regulates genes involved in neural crest cell guidance. Hum. Genet. 133, 997e1009. http:// dx.doi.org/10.1007/s00439-014-1444-2. van Ravenswaaij-Arts, C.M. van, Blake, K., Hoefsloot, L., Verloes, A., 2015. Clinical utility gene card for: CHARGE syndrome e update 2015. Eur. J. Hum. Genet. 23, 1e4. http://dx.doi.org/10.1038/ejhg.2014.170. Vissers, L.E.L.M., van Ravenswaaij, C.M. a, Admiraal, R., Hurst, J. a, de Vries, B.B. a, Janssen, I.M., van der Vliet, W. a, Huys, E.H.L.P.G., de Jong, P.J., Hamel, B.C.J., Schoenmakers, E.F.P.M., Brunner, H.G., Veltman, J. a, van Kessel, A.G., 2004. Mutations in a new member of the chromodomain gene family cause CHARGE syndrome. Nat. Genet. 36, 955e957. http://dx.doi.org/10.1038/ng1407.
Contents lists available at ScienceDirect
European Journal of Medical Genetics journal homepage: http://www.elsevier.com/locate/ejmg
A case of mild CHARGE syndrome associated with a splice site mutation in CHD7 €l Garabedian b, Sylvette Wiener-Vacher c, Constance Wells a, Natalie Loundon b, Noe raldine Goudeffroye a, Tania Attie -Bitach a, e, Marie-Dominique Cordier-Bouvier d, Ge f, * Sandrine Marlin ^pital Necker-Enfants Malades, Paris, France Service d'Histologie, Embryologie et Cytog en etique, Ho ^pital Necker-Enfants Malades, Paris, France Service d'ORL, Ho ^pital Robert Debr Service d'ORL, Ho e, Paris, France d ^pital Trousseau, Paris, France Service de Radiologie, Ho e Universit e Paris Descartes, Sorbonne Paris-Cit e, Paris, France f ^pital Necker-Enfants Malades, Paris, France Centre de r ef erence des surdit es g en etiques, Service de G en etique, Institut Imagine, Ho a
b c
a r t i c l e i n f o
a b s t r a c t
Article history: Received 20 September 2015 Received in revised form 22 February 2016 Accepted 22 February 2016 Available online 24 February 2016
CHARGE syndrome (MIM#214800) (Coloboma, Heart defect, Atresia of choanae, Retarded growth and development, Genital hypoplasia, Ear abnormalities/deafness) is caused by heterozygous mutation of CHD7 transmitted in an autosomal dominant manner. In this report, we describe a patient with bilateral hearing impairment, unusually-shaped ears, no intellectual disability and a patent ductus arteriosus. Further investigation showed abnormal semicircular canals and the presence of olfactory bulbs. He does not fulfill the Blake or the Verloes criteria for CHARGE. A de novo mutation at the donor splice site of intron 33 was identified (c.7164 þ 1G > A). It is of importance to diagnose mildly affected patients for appropriate genetic counselling and to better understand the mild end of the phenotypic spectrum of CHARGE syndrome. © 2016 Elsevier Masson SAS. All rights reserved.
Keywords: CHARGE CHD7 Sensorineural hearing impairment Semicircular canal malformation
1. Introduction CHARGE syndrome (MIM#214800) is a multiorgan genetic autosomal dominant condition affecting from 1/15000 to 1/17000 births (van Ravenswaaij-Arts et al., 2015). CHARGE is an acronym for Coloboma, Heart defect, Atresia of choanae, Retarded growth and development, Genital hypoplasia, Ear abnomalities/deafness. It was first described in 1979 by Hall and Hittner in two separate reports. Heterozygous mutations in the Chromodomain Helicase DNA-binding protein 7 gene (CHD7, MIM#608892) located on 8q12, were found to be the main cause of CHARGE syndrome by Vissers et al., in 2004 (Vissers et al., 2004). Mutations are found in 60e70% of patients with CHARGE syndrome. Most mutations are truncating or frameshift mutations with a premature stop codon (Sanlaville and Verloes, 2007; Schulz et al., 2014) leading to haploinsufficiency. The diagnostic criteria were developed by Blake in
f e rence des surdite s ge ne tiques Institut * Corresponding author. Centre de re ^pital Necker-Enfants Malades, 149 Rue de Se vres, 75015 Paris, France. Imagine, Ho E-mail address: [email protected] (S. Marlin). http://dx.doi.org/10.1016/j.ejmg.2016.02.012 1769-7212/© 2016 Elsevier Masson SAS. All rights reserved.
1998, reviewed by Verloes in 2005, and updated in 2007 (Sanlaville and Verloes, 2007). The criteria classify patients into typical, incomplete and atypical CHARGE syndrome, depending on the number of abnormal phenotypic features. However, some patients with a CHD7 mutation only fulfill atypical CHARGE criteria or do not fulfill the criteria, with only one or two signs (Hughes et al., 2014; Jongmans et al., 2007). This case report adds to the understanding of the mild end of the phenotypic spectrum of CHD7 mutations. 2. Case report The patient is the second-born of three children of a nonconsanguineous couple. Family history was unremarkable. He was born after an uneventful pregnancy, with a birth weight of 3230 g (20th percentile), a birth length of 50 cm (30th percentile) and a head circumference of 35 cm (40th percentile). Apgar scores were 10/10. Mild feeding problems were noted as a new-born but he did not require medical intervention. He was diagnosed with profound bilateral hearing impairment at the age of 16 months. Clinical features at 2 ½ years old showed a
196
C. Wells et al. / European Journal of Medical Genetics 59 (2016) 195e197
square face with frontal bossing. The ears were mildly low-set, without ear lobes. He also had retractile testes and no micropenis. His growth parameters were on the 75th percentile for height, weight and head circumference. He walked at 18 months old and fell frequently until the age of 3. He had cochlear implants on the right ear at 3 and on the left ear at 4 years old. He had surgery to close a patent ductus arteriosus at 5 years old. Aged 8 years and 8 months (Fig. 1) he showed no significant intellectual disability, behaved normally and went to a regular school. He measured 1.35 m (þ1DS) and weighed 27 kg (þ1DS). He presented no clinical sign of endocrine deficiencies but has not yet reached the age of puberty. Neurological examination did not reveal any cranial nerve dysfunction. He did not complain of hyposmia and did not undergo olfactory testing. Clinical examination and functional vestibular tests (otolithic evoked response and caloric test) indicated a bilateral vestibular are flexia. CT-scan of the temporal bones showed normal cochlea and bilateral symmetrical vestibular abnormalities. The vestibules were dilated. There was a bilateral complete agenesis of the lateral semicircular canal and a rudimentary posterior semicircular canal (Fig. 2). Brain-MRI confirmed the vestibular abnormalities and showed that the cranial nerves VII and VIII were morphologically normal. The olfactory bulbs and olfactory sulci were normal. A renal ultrasound showed two normal-sized kidneys. Ophthalmological examination showed an intermittent divergent strabismus but no coloboma. The patient fulfills one major (Ear abnormality) with one minor criterion (Patent ductus arteriosus) according to the Blake criteria. According to the Verloes criteria (Bergman et al., 2011) he has one major criterion (hypoplasia of semicircular canals) and two minor criteria (heart malformation and sensorineural deafness), classifying him as atypical CHARGE. Because of the typical CHARGE inner ear malformation, CHD7 was screened on DNA extracted from a blood sample, by direct Sanger sequencing of the 37 coding exons and adjacent intronic regions (NM_017780.3) as previously described (Sanlaville and Verloes, 2007). A heterozygous mutation was found at the donor splice site of intron 33: c.7164 þ 1G > A, not reported in UCSC, ensembl, 1000 genome project or the exome variant server. All prediction tools (Alamut software using MaxENtSplice, HSF, NNsplice and Genesplicer) predict that this variant abolishes the donor splice site of intron 33. The most likely effect would be the skipping of exon 33 from the transcript (from c.6937_to c.7164) and absence of 76 aminoacids (Asp2313_to Lys2388) from the protein, however, an alternative splice site within intron 33 could also be used. The parents were also tested with DNA extracted from a blood sample and were shown not to carry the mutation. The
mutation was present in the patient's DNA extracted from urine and saliva with a balanced ratio, and therefore no argument for a somatic mosaicism.
3. Discussion We report a patient with mild CHARGE syndrome, with bilateral profound sensorineural deafness, typical inner ear malformation and a heart defect. He does not fulfill the Blake or the Verloes criteria for CHARGE. Splice site mutations as found in our patient represent around 11% of CHD7 mutations found in CHARGE patients (Sanlaville and Verloes, 2007). The molecular diagnosis of our patient highlights several points. Firstly, we were able to complete clinical examination with complementary examinations (e.g. renal ultrasound, brain MRI). Even though no additional abnormalities were found, the patient will be closely monitored for an early diagnosis of signs which might appear with time, such as hypogonadotrophic hypogonadism and puberty delay or scoliosis. Hypogonadism should be detected early in order to give hormone therapy to prevent osteoporosis. Secondly, an appropriate genetic counselling was given to his parents and will later be available to himself. The molecular diagnosis in our patient was made after the cochlear implantation. However, the presentation of the typical inner ear malformation at imaging before a cochlear implantation should prompt molecular testing before the implantation is performed in order to be informed of other features commonly seen in CHARGE syndrome that may hamper the implantation procedure, such as an aberrant route of the facial nerve and severe hypoplasia of the acoustic nerve, which are common in CHARGE syndrome. Typical CHARGE syndrome is well recognized, but the «mild» end of the spectrum is harder to identify. Those patients with only few symptoms can transmit their mutation and have offspring with a more severe phenotype. Because of the high phenotypic variability, it is important to diagnose such patients to give accurate genetic counselling. Tentative genotypeephenotype correlations have indicated that more severe forms are associated with truncating mutations (Legendre et al., 2012). A report has shown that missense mutations are more frequent in patients with Kallman syndrome due to a CHD7 mutation than in typical CHARGE patients (Marcos et al., 2014). We need to have more descriptions of «mild» CHARGE patients to understand the full spectrum of CHD7 mutations. The effects of the mutation of our patient have not been studied at the RNA and/or protein level. In silico, the splice mutation is predicted to result in an in-frame deletion of exon 33 and a shorter protein lacking 76 aminoacids without any known functional domain.
Fig. 1. Patient at 8 years 8 months. He has a square face, frontal bossing and no earlobes.
C. Wells et al. / European Journal of Medical Genetics 59 (2016) 195e197
197
Fig. 2. Temporal CT-scan images. Complete agenesis of the lateral semicircular canal (b) compared to a control (a).
One of the strategies tested to identify mild CHARGE patients was to sequence CHD7 in patients with only one sign of CHARGE (Bergman et al., 2011). No CHD7 mutations were found in a cohort of atrio-ventricular defects or cono-troncal heart defects (CorstenJanssen et al., 2014). Bergman et al. suggested studying familial cases. There have been 18 families published so far, including a three-generational family (Hughes et al., 2014). Most patients with mild CHARGE were diagnosed after their severely affected child was diagnosed. Somatic mosaicism could also explain clinical variability (Jongmans et al., 2007, 2006). In our patient, three tissues were analysed and did not show any evidence of somatic mosaicism. In her guidelines for CHD7 analysis Bergman states that a temporal bone CT-scan should be performed if only 2 cardinal features or 1 cardinal and 1 supportive feature are present in a patient prior to CHD7 analysis, to evaluate the morphology of semicircular canals (Bergman et al., 2011). Following those guidelines our patient would have been diagnosed (one cardinal feature: abnormal functional vestibular test and one supportive: congenital heart defect). Along with these recommendations in CHARGE (Amiel et al., 2001), the importance of temporal bone imaging in sensorineural deafness should also be emphasized so as to evaluate and guide molecular analysis in patients with hearing impairment. Although the guideline for CHD7 testing presented by Bergman et al. has proven to be very sensitive (Hale et al., 2015), a diagnosis may still be missed in patients presenting with only one major or only one minor clinical feature. Next generation sequencing offers the possibility to diagnose these patients. 4. Conclusion The mild end of the phenotypic spectrum of CHD7 mutations is starting to emerge. We report here another sporadic case with mild CHARGE syndrome, with heart defect, sensorineural deafness and hypoplastic semi-circular canals. It should be emphasized that patients should not be rejected for CHD7 analysis if they do not fulfill criteria for atypical or typical CHARGE as there is a high intraand inter-familial variability. A larger screening of mild cases will be facilitated by the next-generation sequencing. References Amiel, J., Attie-Bitach, T., Marianowski, R., Cormier-Daire, V., Abadie, V., Bonnet, D., Gonzales, M., Chemouny, S., Brunelle, F., Munnich, A., Manach, Y., Lyonnet, S., 2001. Temporal bone anomaly proposed as a major criteria for diagnosis of CHARGE syndrome. Am. J. Med. Genet. 99, 124e127, 10.1002/10968628(20010301)99:2<124::AID-AJMG1114>3.0.CO;2e9.
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