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A Novel MECP2 Mutation in a Boy with Neonatal Encephalopathy and Facial Dysmorphism
A Novel MECP2 Mutation in a Boy with Neonatal Encephalopathy and Facial Dysmorphism Kristina Ju¨lich, MD, Denise Horn, MD, Peter Burfeind, PhD, Thomas Erler, MD, and Bernd Auber, MD Methly-CpG-binding protein 2 (MECP2) mutations cause Rett syndrome in females. Here we report on a male infant with neonatal encephalopathy, myoclonic jerks, and irregular breathing patterns caused by a novel frameshift mutation in the MECP2 gene. In addition he has facial dysmorphisms previously not described in these patients. (J Pediatr 2009;155:140-3)
R
ett syndrome (RTT) is an X-linked dominant disorder that occurs with a frequency of 1:10 000 in girls. Typically, affected girls develop normally during the first months of life. Between 6 months and 2 years of age developmental regression becomes evident through loss of speech, purposeful hand movements, and social, as well as cognitive abilities. Patients develop a severe disorder with secondary microcephaly, central respiratory dysregulation, or stereotypic hand movements, and they often have intractable seizures. In most females with RTT, a mutation in the methly-CpGbinding protein 2 (MECP2) gene is detected that encodes the methyl-CpG binding protein 2. MECP2 seems to function in mature rather than developing neurons, but it is presently unknown how a loss of MECP2 function results in the RTT phenotype.1 Although a homozygous or hemizygous MECP2 mutation was previously considered to be lethal, in 1999 a male was reported with neonatal encephalopathy and a MECP2 mutation that caused RTT in female family members.2 Increasing numbers of males with MECP2 mutations have been identified.3-5 The consequences of MECP2 mutations in males can be classified into 3 groups. Some mutations have already been identified to cause RTT in females. Male patients carrying those mutations suffer from severe neonatal encephalopathy and usually die in early childhood. In males with a MECP2 mutation and a 47,XXY karyotype or a somatic mosaicism, the phenotype can present as the classic RTT that is normally seen in female patients. In a second subgroup, mutations have been detected that are not found in girls with RTT because the effects of these mutations are mild in heterozygotes and, hence, remain undetected. These boys suffer from nonspecific X-linked mental retardation with variable phenotypes ranging from mild to severe retardation. In the third subgroup, a duplication of at least the MECP2 gene itself and often of other genes in the vicinity causes a severe pheno-
CGH MECP2 PCR RTT
Comparative genome hybridization Methyl-CpG-binding protein 2 Polymerase chain reaction Rett syndrome
type with mental retardation, infantile hypotonia, and recurrent respiratory infections. No specific dysmorphic signs are known for these groups.
Case Report The affected boy is the second child of healthy unrelated parents. There are no neurodevelopmental abnormalities in the family history. His 4-year old sister developed normally. He was delivered by Caesarian section after 34 weeks gestation because of fetal distress and reduced fetal movements. Apgar scores were 5/7/8, and umbilical cord blood pH was 7.35. His birth weight, length, and head circumference were between the 50th to 90th percentile. During the neonatal period he had frequent apneas and feeding difficulties, which required caffeine therapy and tube feeding for several weeks. His muscle tone was fluctuated from floppiness to opisthotonos. The results of electroencephalography and cranial ultrasonography were normal. Polysomnography revealed irregular breathing patterns (Figure, A). At the age of 3 months the patient’s parents noticed myoclonic jerks several times per day in the right leg. At this time electroencephalography showed multifocal spiking activity without signs of hypsarrhythmia. There were no structural abnormalities on cerebral magnetic resonance imaging. Further laboratory test results for inborn errors of metabolism and neurotransmitters in the cerebrospinal fluid were normal. The karyotype from cultured blood lymphocytes revealed no structural or numerical abnormalities at 500 band resolution (46, XY). Testing for submicroscopic chromosomal aberrations with high-resolution array comparative genome hybridization (CGH) was negative. At the age of 6 months the patient presented with generalized muscular hypotonia and severe developmental delay. He
From the Department of Pediatric Neurology (K.J.) and the Institute of Medical Genetics (D.H.), Charite´–University Medical Center, Berlin, the Institute of Human Genetics, University of Go¨ttingen (P.B., B.A.), Go¨ttingen, and the Department of Pediatrics, Carl-Thiem Hospital Cottbus (T.E.), Cottbus, Germany The authors declare no conflicts of interest. 0022-3476/$ - see front matter. Copyright ª 2009 Mosby Inc. All rights reserved. 10.1016/j.jpeds.2009.01.035
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Vol. 155, No. 1 July 2009
Figure. A, Polysomnogram obtained when the patient was 2 weeks of age. From his first days of life the patient had irregular breathing patterns with frequent central apneas followed by oxygen desaturation and bradycardia. B, Head growth percentiles for boys in the first months of life (normalized for birth at 35 weeks gestation). Our patient had development of secondary microcephaly at 3 months of age (red circles indicate the patient’s head circumference, lines indicating the 97th–50th–3rd percentile). C, Our patient at the age of 11 months. He had microcephaly, muscular hypotonia, and severe developmental delay. Notable were a prominent forehead, midface hypoplasia, discrete temporal incisions, up-slanting palpebral fissures, thick eyebrows and synophrys, long eye lashes, flat nasal bridge, a deeply grooved philtrum, and large ears. His left ear shows marks from stereotypical ear rubbing. D, Chromatograms of the according genomic MECP2 gene sequence in the mother (top, wild type) and the patient (bottom). The 2 deleted base pairs in exon 3 of the MECP2 gene (c.119_120delAG) are boxed in the wild type sequence. The arrow indicates the position of the deletion in the patient’s sequence.
141
THE JOURNAL OF PEDIATRICS
www.jpeds.com
Vol. 155, No. 1
Table. Clinical features of males with MECP2 mutations and severe neonatal encephalopathy Mutation
No. of patients
c.119_120delAG (p.E40fs) c.378-3_383del9 (p.N126KfsX11)
1 1
+ +
+ +
+ +
+ +
c.469 T > A (p.F157I) c.473C > T (p.T158 M)
1 5*
+ + (3/5)
+ + (5/5)
+ + (3/5)
+ + (3/5)
c.488_489delGG (p.G163fs) c.753_754insG (p.G252fs) c.754_755insC (p.E252fs) c.806delG (p.G269fs) c.808delC (p.R270fs) c.880C > T (p.R294X) c.1154_1185del32 (p.385fs) c.1250A > T (p.K417M) g.58483_65650del; g.65664_66958del{ (p.R9fs)f
1 1 1 3 2 1 2x 1 1
+ n.d. + + (3/3) + (2/2) + + (2/2) + +
+ + + + (3/3) + (1/2) n.d. + (2/2) + +
+ + + + (3/3) + (1/2) + n.d. (2/2) + +
+ + + + (3/3) + (1/2) + (2/2) +
Microcephaly
Respiratory dysfunction
Movement disorder
Seizures/abnormal EEG
Dysmorphic signs Facial dysmorphisms High palate, myopathic-looking face Salt & pepper hair (2/5), high palate (1/5) Salt & pepper hair, high palate n.d. Facial dysmorphisms† (1/2) n.d. (2/2) n.d. Scaphocephaly, myopathic-looking face
Reference This report 5 3-5 3-5 3-5 3,5 3-5 3-5 3-5 5,z 3,5 3,k 5
n.d., Not determined, that is, feature not mentioned in the cited article. Our patient (italicized) has dysmorphic facial features not described in the other patients. Features in brackets were not present in all males carrying this mutation. *One patient had no confirmed mutation, but a brother had the same phenotype and the p.T158M mutation. †Synophrys, up-slanting palpebral fissures, micrognathia. zThis patient has a phenotype resembling classical RTT with loss of acquired skills at older age, but also feeding problems requiring gastrostomy since birth. xOne patient had no confirmed mutation, but a brother had the same phenotype and the p.385fs mutation. kThe pathogenicity of this mutation is unclear, because it is also present in the mother who has a random X-inactivation pattern. {This mutation leads to a large deletion with loss of exons 3 and part of exon 4.
could neither roll over nor hold his head for more than a few seconds. He did not cry, established only infrequent eye-toeye contact and was unable to grasp objects purposefully. He had developed secondary microcephaly and had frequent respiratory infections. He still had feeding difficulties, and his weight had dropped below the third percentile (Figure, B). No abnormal movements such as stereotypic hand-rubbing or focal neurological deficits were noted. Treatment with sulthiame, a carboanhydrase inhibitor used for the treatment of benign focal epilepsies of childhood and seizures in West syndrome (not registered in the United States), suppressed the myoclonic jerks, but in the EEG spiking patterns were still present. We also noted dysmorphic signs, which were not present in other family members (Figure, C). He started to vocalize at the age of 9 months. At 11 months his motor development had not progressed. He displayed only little (but symmetric) general movements. In addition, he developed stereotypic hand movements (ie, rubbing of his ears). Deep tendon reflexes—normal previously—were absent now. The combination of severe developmental delay, seizures, secondary microcephaly, central breathing dysfunction, and stereotypic hand movements led us to suspect an RTT-related disorder despite the more severe phenotype compared with girls. Our patient scored 8/12 on a clinical score developed by Huppke et al6 for diagnosing RTT in girls. A score of 8 or higher is recommended by the authors as a cutoff to test for MECP2 mutations. Mutation analysis of the MECP2 gene revealed a deletion of 2 base pairs in exon 3 (c.119_120delAG). This novel mutation results in a frameshift and early premature stop codon 8 base pairs downstream of the site of the mutation (p.E40fs) (Figure, D). Molecular genetic analysis of the 142
mother revealed a wild type sequence of the corresponding region of the MECP2 gene. In 50 alleles of healthy male control subjects, the deletion described here could not be detected.
Methods MECP2 Analysis Total genomic DNA was prepared from blood lymphocytes with standard techniques. All coding regions and flanking intronic sequences of the MECP2 gene were amplified by polymerase chain reaction (PCR). Primer sequences and the PCR conditions are available on request. PCR products were separated on 1.5% agarose gels. PCR products were purified by use of a PCR purification kit (Millipore, Eschborn, Germany) and sequenced bidirectionally on a Megabace 1000 sequencer (GE Healthcare, Freiburg, Germany) with the DYEnemic ET Terminator cycle sequencing kit (GE Healthcare) used according to the manufacturer’s instructions.
Array CGH Array CGH was performed with the Agilent Human Genome CGH Microarray Kit 244 K (Agilent Technologies, Inc, Santa Clara, California) as previously described.7
Polysomnography Several polysomnograms were recorded during night sleep in a sleep laboratory. Heart rate, oxygen saturation, and respiratory events (central apnea, obstructive apnea, mixed apnea, hypopnea) were recorded. Ju¨lich et al
July 2009
Discussion The boy presented with signs of neonatal encephalopathy, severe developmental delay, seizures, microcephaly, and central breathing disorder. Other causes for mental retardation and seizures in infancy such as inborn errors of metabolism, intrauterine infections, and hypothyroidism were excluded, and no chromosomal aberrations were detected. There are reports of 20 other male patients presenting with severe neonatal or early-onset encephalopathy and confirmed (18 patients) or suspected (2 patients) MECP2 mutations. All of these patients had various degrees of respiratory insufficiency and died in infancy (except for one who is ventilator-dependent at the age of 6 years). Most of them had microcephaly and had some form of movement disorder and seizures. Except for 2 cases with unusual grayish hair color, there is just 1 patient reported with discrete dysmorphic facial features (Table).2-5 The facial dysmorphism seen in our patient could also be due to an additional mutation. However, we did not detect any chromosomal deletions/duplications in an array CGH analysis. In addition, there are several reports on mild facial dysmorphisms in males with MECP2 mutations and mental retardation,8,9 indicating that this can be part of the phenotype of MECP2 mutations in male patients. The exact molecular mechanism leading to RTT is not yet understood. Because MECP2 levels are low during embryogenesis and increase progressively during postnatal development, it may function in mature neurons rather than affecting neuronal precursors. Its levels obviously need to be regulated tightly because mice expressing increased or decreased levels of MECP2 both show neurodevelopmental abnormalities.1 Exons 3 and 4 of MECP2 encode different functional domains involved in DNA and RNA binding, transcriptional repression, and splicing-factor binding. Because of the frameshift mutation and a subsequent premature stop codon in exon 3, none of the functional domains are present in our patient, which suggests a complete loss of functional MECP2 protein. This might explain the severity of the case. Genotype-phenotype relations in RTT have been discussed extensively. Charman et al10 described an association between early truncating mutations and a severe outcome and between late truncating or missense mutations and a better outcome in
CLINICAL AND LABORATORY OBSERVATIONS female patients. However, because of the small number of affected individuals, one can only speculate about genotypephenotype relations in male patients. In males, mutations in the MECP2 gene are found in a variety of phenotypes ranging from mild mental retardation with autistic disorders to classical RTT or severe encephalopathy. The incidence of pathogenic MECP2 mutations in the population of mentally retarded males is estimated between 1.3% and 1.7%.4 Male patients with severe developmental delay, symptoms of central respiratory regulation, microcephaly, and seizures should be considered for MECP2 mutational analysis. n Submitted for publication Aug 14, 2008; last revision received Dec 19, 2008; accepted Jan 9, 2009. Reprint requests: Kristina Ju¨lich, MD, Department of Pediatric Neurology, Charite´ University Hospital, Augustenburger Platz 1, 13353 Berlin, Germany. E-mail: [email protected].
References 1. Chahrour M, Zoghbi HY. The story of Rett syndrome: from clinic to neurobiology. Neuron 2007;56:422-37. 2. Wan M, Lee SS, Zhang X, Houwink-Manville I, Song HR, Amir RE, et al. Rett syndrome and beyond: recurrent spontaneous and familial MECP2 mutations at CpG hotspots. Am J Hum Genet 1999;65:1520-9. 3. Kankirawatana P, Leonard H, Ellaway C, Scurlock J, Mansour A, Makris CM, et al. Early progressive encephalopathy in boys and MECP2 mutations. Neurology 2006;67:164-6. 4. Villard L. MECP2 mutations in males. J Med Genet 2007;44:417-23. 5. Schu¨le B, Armstrong DD, Vogel H, Oviedo A, Francke U. Severe congenital encephalopathy caused by MECP2 null mutations in males: central hypoxia and reduced neuronal dendritic structure. Clin Genet 2008; 74:116-26. 6. Huppke P, Ko¨hler K, Laccone F, Hanefeld F. Indication for genetic testing: a checklist for Rett syndrome. J Pediatr 2003;142:332-5. 7. Martinet D, Filges I, Besuchet Schmutz N, Morris MA, Gaide AC, Dahoun S, et al. Subtelomeric 6p deletion: clinical and arrah-CGH characterization in two patients. Am J Med Genet A 2008;146A:2094-102. 8. Imessaoudene B, Bonnefont JP, Royer G, Cormier-Daire V, Lyonnet S, Lyon G, et al. MECP2 mutation in non-fatal, non-progressive encephalopathy in a male. J Med Genet 2001;38:171-4. 9. Dotti MT, Orrico A, De Stefano N, Battisti C, Siccurelli F, Severi N, et al. A Rett syndrome MECP2 mutation that causes mental retardation in men. Neurology 2002;58:226-30. 10. Charman T, Neilson TC, Mash V, Archer H, Gardiner MT, Knudsen GP, et al. Dimensional phenotypic analysis and functional categorisation of mutations reveal novel genotype-phenotype associations in Rett syndrome. Eur J Hum Genet 2005;13:1121-30.
A Novel MECP2 Mutation in a Boy with Neonatal Encephalopathy and Facial Dysmorphism
143
R
ett syndrome (RTT) is an X-linked dominant disorder that occurs with a frequency of 1:10 000 in girls. Typically, affected girls develop normally during the first months of life. Between 6 months and 2 years of age developmental regression becomes evident through loss of speech, purposeful hand movements, and social, as well as cognitive abilities. Patients develop a severe disorder with secondary microcephaly, central respiratory dysregulation, or stereotypic hand movements, and they often have intractable seizures. In most females with RTT, a mutation in the methly-CpGbinding protein 2 (MECP2) gene is detected that encodes the methyl-CpG binding protein 2. MECP2 seems to function in mature rather than developing neurons, but it is presently unknown how a loss of MECP2 function results in the RTT phenotype.1 Although a homozygous or hemizygous MECP2 mutation was previously considered to be lethal, in 1999 a male was reported with neonatal encephalopathy and a MECP2 mutation that caused RTT in female family members.2 Increasing numbers of males with MECP2 mutations have been identified.3-5 The consequences of MECP2 mutations in males can be classified into 3 groups. Some mutations have already been identified to cause RTT in females. Male patients carrying those mutations suffer from severe neonatal encephalopathy and usually die in early childhood. In males with a MECP2 mutation and a 47,XXY karyotype or a somatic mosaicism, the phenotype can present as the classic RTT that is normally seen in female patients. In a second subgroup, mutations have been detected that are not found in girls with RTT because the effects of these mutations are mild in heterozygotes and, hence, remain undetected. These boys suffer from nonspecific X-linked mental retardation with variable phenotypes ranging from mild to severe retardation. In the third subgroup, a duplication of at least the MECP2 gene itself and often of other genes in the vicinity causes a severe pheno-
CGH MECP2 PCR RTT
Comparative genome hybridization Methyl-CpG-binding protein 2 Polymerase chain reaction Rett syndrome
type with mental retardation, infantile hypotonia, and recurrent respiratory infections. No specific dysmorphic signs are known for these groups.
Case Report The affected boy is the second child of healthy unrelated parents. There are no neurodevelopmental abnormalities in the family history. His 4-year old sister developed normally. He was delivered by Caesarian section after 34 weeks gestation because of fetal distress and reduced fetal movements. Apgar scores were 5/7/8, and umbilical cord blood pH was 7.35. His birth weight, length, and head circumference were between the 50th to 90th percentile. During the neonatal period he had frequent apneas and feeding difficulties, which required caffeine therapy and tube feeding for several weeks. His muscle tone was fluctuated from floppiness to opisthotonos. The results of electroencephalography and cranial ultrasonography were normal. Polysomnography revealed irregular breathing patterns (Figure, A). At the age of 3 months the patient’s parents noticed myoclonic jerks several times per day in the right leg. At this time electroencephalography showed multifocal spiking activity without signs of hypsarrhythmia. There were no structural abnormalities on cerebral magnetic resonance imaging. Further laboratory test results for inborn errors of metabolism and neurotransmitters in the cerebrospinal fluid were normal. The karyotype from cultured blood lymphocytes revealed no structural or numerical abnormalities at 500 band resolution (46, XY). Testing for submicroscopic chromosomal aberrations with high-resolution array comparative genome hybridization (CGH) was negative. At the age of 6 months the patient presented with generalized muscular hypotonia and severe developmental delay. He
From the Department of Pediatric Neurology (K.J.) and the Institute of Medical Genetics (D.H.), Charite´–University Medical Center, Berlin, the Institute of Human Genetics, University of Go¨ttingen (P.B., B.A.), Go¨ttingen, and the Department of Pediatrics, Carl-Thiem Hospital Cottbus (T.E.), Cottbus, Germany The authors declare no conflicts of interest. 0022-3476/$ - see front matter. Copyright ª 2009 Mosby Inc. All rights reserved. 10.1016/j.jpeds.2009.01.035
140
Vol. 155, No. 1 July 2009
Figure. A, Polysomnogram obtained when the patient was 2 weeks of age. From his first days of life the patient had irregular breathing patterns with frequent central apneas followed by oxygen desaturation and bradycardia. B, Head growth percentiles for boys in the first months of life (normalized for birth at 35 weeks gestation). Our patient had development of secondary microcephaly at 3 months of age (red circles indicate the patient’s head circumference, lines indicating the 97th–50th–3rd percentile). C, Our patient at the age of 11 months. He had microcephaly, muscular hypotonia, and severe developmental delay. Notable were a prominent forehead, midface hypoplasia, discrete temporal incisions, up-slanting palpebral fissures, thick eyebrows and synophrys, long eye lashes, flat nasal bridge, a deeply grooved philtrum, and large ears. His left ear shows marks from stereotypical ear rubbing. D, Chromatograms of the according genomic MECP2 gene sequence in the mother (top, wild type) and the patient (bottom). The 2 deleted base pairs in exon 3 of the MECP2 gene (c.119_120delAG) are boxed in the wild type sequence. The arrow indicates the position of the deletion in the patient’s sequence.
141
THE JOURNAL OF PEDIATRICS
www.jpeds.com
Vol. 155, No. 1
Table. Clinical features of males with MECP2 mutations and severe neonatal encephalopathy Mutation
No. of patients
c.119_120delAG (p.E40fs) c.378-3_383del9 (p.N126KfsX11)
1 1
+ +
+ +
+ +
+ +
c.469 T > A (p.F157I) c.473C > T (p.T158 M)
1 5*
+ + (3/5)
+ + (5/5)
+ + (3/5)
+ + (3/5)
c.488_489delGG (p.G163fs) c.753_754insG (p.G252fs) c.754_755insC (p.E252fs) c.806delG (p.G269fs) c.808delC (p.R270fs) c.880C > T (p.R294X) c.1154_1185del32 (p.385fs) c.1250A > T (p.K417M) g.58483_65650del; g.65664_66958del{ (p.R9fs)f
1 1 1 3 2 1 2x 1 1
+ n.d. + + (3/3) + (2/2) + + (2/2) + +
+ + + + (3/3) + (1/2) n.d. + (2/2) + +
+ + + + (3/3) + (1/2) + n.d. (2/2) + +
+ + + + (3/3) + (1/2) + (2/2) +
Microcephaly
Respiratory dysfunction
Movement disorder
Seizures/abnormal EEG
Dysmorphic signs Facial dysmorphisms High palate, myopathic-looking face Salt & pepper hair (2/5), high palate (1/5) Salt & pepper hair, high palate n.d. Facial dysmorphisms† (1/2) n.d. (2/2) n.d. Scaphocephaly, myopathic-looking face
Reference This report 5 3-5 3-5 3-5 3,5 3-5 3-5 3-5 5,z 3,5 3,k 5
n.d., Not determined, that is, feature not mentioned in the cited article. Our patient (italicized) has dysmorphic facial features not described in the other patients. Features in brackets were not present in all males carrying this mutation. *One patient had no confirmed mutation, but a brother had the same phenotype and the p.T158M mutation. †Synophrys, up-slanting palpebral fissures, micrognathia. zThis patient has a phenotype resembling classical RTT with loss of acquired skills at older age, but also feeding problems requiring gastrostomy since birth. xOne patient had no confirmed mutation, but a brother had the same phenotype and the p.385fs mutation. kThe pathogenicity of this mutation is unclear, because it is also present in the mother who has a random X-inactivation pattern. {This mutation leads to a large deletion with loss of exons 3 and part of exon 4.
could neither roll over nor hold his head for more than a few seconds. He did not cry, established only infrequent eye-toeye contact and was unable to grasp objects purposefully. He had developed secondary microcephaly and had frequent respiratory infections. He still had feeding difficulties, and his weight had dropped below the third percentile (Figure, B). No abnormal movements such as stereotypic hand-rubbing or focal neurological deficits were noted. Treatment with sulthiame, a carboanhydrase inhibitor used for the treatment of benign focal epilepsies of childhood and seizures in West syndrome (not registered in the United States), suppressed the myoclonic jerks, but in the EEG spiking patterns were still present. We also noted dysmorphic signs, which were not present in other family members (Figure, C). He started to vocalize at the age of 9 months. At 11 months his motor development had not progressed. He displayed only little (but symmetric) general movements. In addition, he developed stereotypic hand movements (ie, rubbing of his ears). Deep tendon reflexes—normal previously—were absent now. The combination of severe developmental delay, seizures, secondary microcephaly, central breathing dysfunction, and stereotypic hand movements led us to suspect an RTT-related disorder despite the more severe phenotype compared with girls. Our patient scored 8/12 on a clinical score developed by Huppke et al6 for diagnosing RTT in girls. A score of 8 or higher is recommended by the authors as a cutoff to test for MECP2 mutations. Mutation analysis of the MECP2 gene revealed a deletion of 2 base pairs in exon 3 (c.119_120delAG). This novel mutation results in a frameshift and early premature stop codon 8 base pairs downstream of the site of the mutation (p.E40fs) (Figure, D). Molecular genetic analysis of the 142
mother revealed a wild type sequence of the corresponding region of the MECP2 gene. In 50 alleles of healthy male control subjects, the deletion described here could not be detected.
Methods MECP2 Analysis Total genomic DNA was prepared from blood lymphocytes with standard techniques. All coding regions and flanking intronic sequences of the MECP2 gene were amplified by polymerase chain reaction (PCR). Primer sequences and the PCR conditions are available on request. PCR products were separated on 1.5% agarose gels. PCR products were purified by use of a PCR purification kit (Millipore, Eschborn, Germany) and sequenced bidirectionally on a Megabace 1000 sequencer (GE Healthcare, Freiburg, Germany) with the DYEnemic ET Terminator cycle sequencing kit (GE Healthcare) used according to the manufacturer’s instructions.
Array CGH Array CGH was performed with the Agilent Human Genome CGH Microarray Kit 244 K (Agilent Technologies, Inc, Santa Clara, California) as previously described.7
Polysomnography Several polysomnograms were recorded during night sleep in a sleep laboratory. Heart rate, oxygen saturation, and respiratory events (central apnea, obstructive apnea, mixed apnea, hypopnea) were recorded. Ju¨lich et al
July 2009
Discussion The boy presented with signs of neonatal encephalopathy, severe developmental delay, seizures, microcephaly, and central breathing disorder. Other causes for mental retardation and seizures in infancy such as inborn errors of metabolism, intrauterine infections, and hypothyroidism were excluded, and no chromosomal aberrations were detected. There are reports of 20 other male patients presenting with severe neonatal or early-onset encephalopathy and confirmed (18 patients) or suspected (2 patients) MECP2 mutations. All of these patients had various degrees of respiratory insufficiency and died in infancy (except for one who is ventilator-dependent at the age of 6 years). Most of them had microcephaly and had some form of movement disorder and seizures. Except for 2 cases with unusual grayish hair color, there is just 1 patient reported with discrete dysmorphic facial features (Table).2-5 The facial dysmorphism seen in our patient could also be due to an additional mutation. However, we did not detect any chromosomal deletions/duplications in an array CGH analysis. In addition, there are several reports on mild facial dysmorphisms in males with MECP2 mutations and mental retardation,8,9 indicating that this can be part of the phenotype of MECP2 mutations in male patients. The exact molecular mechanism leading to RTT is not yet understood. Because MECP2 levels are low during embryogenesis and increase progressively during postnatal development, it may function in mature neurons rather than affecting neuronal precursors. Its levels obviously need to be regulated tightly because mice expressing increased or decreased levels of MECP2 both show neurodevelopmental abnormalities.1 Exons 3 and 4 of MECP2 encode different functional domains involved in DNA and RNA binding, transcriptional repression, and splicing-factor binding. Because of the frameshift mutation and a subsequent premature stop codon in exon 3, none of the functional domains are present in our patient, which suggests a complete loss of functional MECP2 protein. This might explain the severity of the case. Genotype-phenotype relations in RTT have been discussed extensively. Charman et al10 described an association between early truncating mutations and a severe outcome and between late truncating or missense mutations and a better outcome in
CLINICAL AND LABORATORY OBSERVATIONS female patients. However, because of the small number of affected individuals, one can only speculate about genotypephenotype relations in male patients. In males, mutations in the MECP2 gene are found in a variety of phenotypes ranging from mild mental retardation with autistic disorders to classical RTT or severe encephalopathy. The incidence of pathogenic MECP2 mutations in the population of mentally retarded males is estimated between 1.3% and 1.7%.4 Male patients with severe developmental delay, symptoms of central respiratory regulation, microcephaly, and seizures should be considered for MECP2 mutational analysis. n Submitted for publication Aug 14, 2008; last revision received Dec 19, 2008; accepted Jan 9, 2009. Reprint requests: Kristina Ju¨lich, MD, Department of Pediatric Neurology, Charite´ University Hospital, Augustenburger Platz 1, 13353 Berlin, Germany. E-mail: [email protected].
References 1. Chahrour M, Zoghbi HY. The story of Rett syndrome: from clinic to neurobiology. Neuron 2007;56:422-37. 2. Wan M, Lee SS, Zhang X, Houwink-Manville I, Song HR, Amir RE, et al. Rett syndrome and beyond: recurrent spontaneous and familial MECP2 mutations at CpG hotspots. Am J Hum Genet 1999;65:1520-9. 3. Kankirawatana P, Leonard H, Ellaway C, Scurlock J, Mansour A, Makris CM, et al. Early progressive encephalopathy in boys and MECP2 mutations. Neurology 2006;67:164-6. 4. Villard L. MECP2 mutations in males. J Med Genet 2007;44:417-23. 5. Schu¨le B, Armstrong DD, Vogel H, Oviedo A, Francke U. Severe congenital encephalopathy caused by MECP2 null mutations in males: central hypoxia and reduced neuronal dendritic structure. Clin Genet 2008; 74:116-26. 6. Huppke P, Ko¨hler K, Laccone F, Hanefeld F. Indication for genetic testing: a checklist for Rett syndrome. J Pediatr 2003;142:332-5. 7. Martinet D, Filges I, Besuchet Schmutz N, Morris MA, Gaide AC, Dahoun S, et al. Subtelomeric 6p deletion: clinical and arrah-CGH characterization in two patients. Am J Med Genet A 2008;146A:2094-102. 8. Imessaoudene B, Bonnefont JP, Royer G, Cormier-Daire V, Lyonnet S, Lyon G, et al. MECP2 mutation in non-fatal, non-progressive encephalopathy in a male. J Med Genet 2001;38:171-4. 9. Dotti MT, Orrico A, De Stefano N, Battisti C, Siccurelli F, Severi N, et al. A Rett syndrome MECP2 mutation that causes mental retardation in men. Neurology 2002;58:226-30. 10. Charman T, Neilson TC, Mash V, Archer H, Gardiner MT, Knudsen GP, et al. Dimensional phenotypic analysis and functional categorisation of mutations reveal novel genotype-phenotype associations in Rett syndrome. Eur J Hum Genet 2005;13:1121-30.
A Novel MECP2 Mutation in a Boy with Neonatal Encephalopathy and Facial Dysmorphism
143