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DYRK1A mutations in two unrelated patients
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European Journal of Medical Genetics journal homepage: http://www.elsevier.com/locate/ejmg
Clinical report
DYRK1A mutations in two unrelated patients Q2
Lyse Ruaud a, Cyril Mignot b, Agnès Guët c, Christelle Ohl d, Caroline Nava b, Delphine Héron b, Boris Keren b, Christel Depienne b, Valérie Benoit e, Isabelle Maystadt e, Damien Lederer e, Daniel Amsallem f, Juliette Piard a, * a
Centre de Génétique Humaine, CHU Besançon, France Département de Génétique, APHP, GH Pitié-Salpêtrière, Centre de Référence des Déficiences Intellectuelles de Causes Rares, Paris, France c Service de Pédiatrie, APHP, Hôpital Louis Mourier, Colombes, France d Centre d’Action Médico-Sociale Précoce, Courbevoie, France e Institut de pathologie et de Génétique, Gosselies, Belgique f Service de Pédiatrie, CHU Besançon, France b
a r t i c l e i n f o
a b s t r a c t
Article history: Received 6 December 2014 Accepted 26 December 2014 Available online xxx
The Dual-specify tyrosine phosphorylation-regulated kinase 1A (DYRK1A) gene has been extensively studied for its role in the pathophysiology of intellectual disability (ID) in Down syndrome. The rise of next generation sequencing (NGS) and array-CGH (aCGH) in diagnostic settings for the evaluation of patients with ID allowed the identification of 17 patients carrying heterozygous genetic aberrations involving DYRK1A to date. The rate of DYRK1A mutations in this population reaches >1% in published NGS studies. The current report aims at further defining the phenotype of this encephalopathy with the detailed report of two unrelated patients. Both patients were boys with developmental delay, febrile seizures, facial dysmorphism and brain atrophy on MRI. Patient #1 had autistic behaviors and micropenis and Patient #2 had stereotypies and microcephaly. NGS analyses identified heterozygous de novo variants in DYRK1A: the c.613C >T (p.Arg205*) nonsense mutation in Patient #1 and the c.932C >T (p.Ser311Phe) missense mutation in Patient #2. Together with previously reported cases, patients with DYRK1A mutations share many clinical features and may have a recognizable phenotype that includes, by decreasing order of frequency: developmental delay or ID with behaviors suggesting autism spectrum disorder, microcephaly, epileptic seizures, facial dysmorphism including ear anomalies (large ears, hypoplastic lobes), thin lips, short philtrum and frontal bossing. Delineation of the phenotype/genotype correlation is not feasible at the moment and will be a challenge for the coming years. Ó 2015 Elsevier Masson SAS. All rights reserved.
Keywords: DYRK1A mutation Dysmorphism Intellectual disability Autism spectrum disorder
1. Introduction The Dual-specify tyrosine phosphorylation-regulated kinase 1A (DYRK1A) gene is located on chromosome 21q22.2 and the role of its product, the DYRK1A protein, in neuronal progenitor proliferation and neuronal differentiation is a growing field of research. It has been involved in the pathogenesis of Alzheimer’s disease [Ryoo et al., 2007], Parkinson’s disease [Kim et al., 2006] and Huntington disease [Kang et al., 2005], as well as in the intellectual disability
* Corresponding author. Centre de Génétique Humaine, Centre-hospitalouniversitaire, Pavillon Saint-Paul, 2 place Saint-Jacques, 25000 Besançon, France. Tel.: þ33 381218187; fax: þ33 381218643. E-mail address: [email protected] (J. Piard).
(ID) of patients with Down syndrome [García-Cerro et al., 2014]. The crucial role of DYRK1A in cerebral development has been further highlighted by the report of patients with cognitive impairment since infancy related to deletions or mutations within the gene (MIM #614104). To date, 11 patients with developmental delay/ID or autism spectrum disorder (ASD) and de novo heterozygous genetic aberrations in DYRK1A have been clinically well described (Table 1). Two of them harbored partial deletions of DYRK1A identified by arrayCGH (aCGH) [Courcet et al., 2012; Van Bon et al., 2011], two carried a balanced translocation inducing DYRK1A truncation [Møller et al., 2008] and seven carried frameshift, nonsense or splice site mutations identified by targeted [Courcet et al., 2012] or next generation sequencing (NGS) [Okamoto et al., 2014; O’Roak et al., 2012; Redin et al., 2014]. Six more patients with a
http://dx.doi.org/10.1016/j.ejmg.2014.12.014 1769-7212/Ó 2015 Elsevier Masson SAS. All rights reserved.
Please cite this article in press as: Ruaud L, et al., DYRK1A mutations in two unrelated patients, European Journal of Medical Genetics (2015), http://dx.doi.org/10.1016/j.ejmg.2014.12.014
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Table 1 Description of the 13 patients with DYRK1A-related encephalopathy. References
Present report
Patients Sex/age Genetic anomalies
Patient #1 M5y de novo nonsense c.613C >T, p.Arg205a
Prenatal findings
Okamoto et al. Clin Genet 2014
Redin et al. J Med Genet 2014
Patient #2 M3y de novo missense c.932C >T p.Ser311Phe
Patient 2 F7y Nonsense c.1699C >T p.Q567a
APN-58 M 16 y de novo nonsense c.613C >T p.Arg205a
APN-87 M5y de novo del/ins + fs c.621_624delinsGAA p.Glu208Asnfs*3
IUGR, single umbilical artery
NA
NA
NA
Moderately delayed
Severe ID
Moderate ID
Moderate ID
Unable to walk Absent + Self-injuries, temper tantrums, vocal tics NA +0.6 No Low-set ears
NA Speech delay +
NA NA +
NA 2 No “Dysmorphic traits”
NA 4 No “Dysmorphic traits”
Down-slanting palpebral fissures, epicanthal folds, hypertelorism Downturned mouth, short philtrum, micrognathia, thin upper lip e
e
e
e
e
Arachnodactyly
e
Psychom. dev.
Global development
Born at 33 WG with normal growth parameters Mildly delayed
Behavioral disturbance
Age of walking Language ASD/autistic behavior ADHD/attention deficit Other
25 m First words 3 y +
3y Two syllable words 3 y +/ (see text)
At birth Last exam Ears
Mean 2 + F (2.5 y) Large ears
1.5 <4 + F (16 m), AED+ Large ears
Eyes
Enophthalmia
e
Mouth
Microretrognathia, smooth philtrum, thin upper lip
Small mouth, long philtrum
Extremities
Syndactyly
e
Other
Frontal bossing, low columella, micropenis, microdontia + Bilateral inguinal hernias ELV + CA (2 y 8 m)
e
Frontal bossing, flat nasal bridge
Pectus excavatum
No Asthma, numerous otitis media ELV + CA (15 m)
No Short stature -3DS, severe amblyopia Normal
+ e
NA Sleep disturbance
NA
ELV
OFC (SD) Epilepsy/seizures (age) Morphologic features
Feeding difficultiesa Other features Brain MRI (age)
M: male. F: female. del: deletion. fs: frameshift. WG: weeks of gestation. IUGR: intra uterine growth retardation. m: months. y: years. Psychom. dev.: psychomotor development. ID: intellectual disability. VIQ: verbal intellectual quotient. NVIQ: non verbal intellectual quotient. ASD: autism spectrum disorder. ADHD: attention deficit hyperactivity disorder. OFC: occipitofrontal circumference. GTCS: generalized tonic clonic seizures. Epilepsy/seizures: + F means that the patient had febrile seizures. epil.: “confirmed” epilepsy. AED: anti-epileptic drug. ELV: enlarged lateral ventricles. CA: cortical atrophy. NA: not available. a Feeding difficulties during the first months of life.
“neurological phenotype” were reported but not described [Yang et al., 2014]. We report on two patients with de novo DYRK1A mutations, including the first described patient harboring a missense mutation, to delineate the phenotype of the DYRK1Aassociated developmental disorder Table 2. 2. Description of patients Patient #1 was the first child of healthy nonconsanguineous French parents. His mother had a miscarriage. His paternal grandfather has retinitis pigmentosa. He was born after an uneventful pregnancy at 33 weeks by vacuum-assisted delivery after a premature rupture of membranes. Birth weight was 1700 g (1 SD), birth length 41.8 cm (1 SD) and occipito-frontal circumference (OFC) 30.5 cm (mean). During the first months of life, he had congenital torticollis, sucking-swallowing difficulties requiring a feeding tube, episodes of bradycardia associated with gastroesophageal reflux. He underwent repair of bilateral inguinal hernias at 3 months old. A micropenis with a blind dimple on the scrotum was noticed without endocrine anomalies. He was efficiently treated with two injections of testosterone enanthate. One episode of febrile seizures occurred at 2.5 years old. The psychomotor development of Patient #1 was delayed: he sat at 10 months old, walked without assistance at 25 months and said his first words at 3 years old. Daytime toilet training was acquired at 4.5 years old. He had hand stereotypies, repetitive and sexual self-stimulation behaviors.
At first examination at 3 years, an unsteady gait and facial dysmorphism comprising large ears, microretrognatia, frontal bossing, thin upper lip, long eyelashes, smooth philtrum, plagiocephaly, low columella, microdontia with dental diastasis and enophtalmia (Fig. 1A and B), as well as bilateral 2e3 toe syndactyly (Fig. 1C), were noticed. At the age of 5 years, Patient #1 had normal growth parameters and an OFC of 49 cm (2 SD). He was treated by stretching plaster for bilateral equinus foot deformity. Brain MRI performed at 2 years and 8 months old showed diffuse cerebral atrophy (Fig. 2A and B). Standard blood chemistry and metabolic screening were negative as well as search for expansion in FMR1, ZEB2 sequencing and methylation at the SNRPN locus. Patient #2 was the first child of healthy nonconsanguineous parents of Asian and Caucasian descent. His paternal uncle had congenital deafness. During pregnancy, intrauterine growth retardation and a single umbilical artery were noticed. Karyotype was 46,XY on cultured amniocytes. Birth weight was 3170 g (25th centile), length 49 cm (1.5 SD) and OFC 33.5 cm (1.5 SD). Multiple episodes of bronchiolitis and otitis media occurred during infancy. He had a series of short generalized febrile seizures at 16 months old that did not recur under valproate therapy. When examined at that time, his psychomotor development was moderately delayed: he was not able to stand, could catch objects but did not manipulate them, babbled but did not say any word. His OFC was 42 cm (<4 SD) with marked slowing of head growth between 3 and 5 months old. He had Asian
Please cite this article in press as: Ruaud L, et al., DYRK1A mutations in two unrelated patients, European Journal of Medical Genetics (2015), http://dx.doi.org/10.1016/j.ejmg.2014.12.014
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Courcet et al. J Med Genet 2012
O’Roak et al. Science 2012
D12 F 14 y del. 2bp + fs c.290_291delCT p.Ser97Cysfs*98
Index case F4y de novo 69 kb del. arr21q22.13(37 644 501e37 713 641) 1 (hg18) IUGR
13890.p1 F 13.5 y de novo splice site mutation c.1098+1G >A
13552.p1 M6y de novo fs p.Ala498Profs*94
12099.p1 M8y de novo fs p.Ile48Lysfs*2
F 33 y Intragenic del.
Patient #1 Patient #2 M2y F 13 y de novo balanced translocation, breakpoint interrupting DYRK1A
Oligoamnios
NA
NA
NA
IUGR
Severe ID
Moderate ID
Slight retardation
Severe ID
3y NA + Inappropriate laughters 4,5 6 + F (12 m), AED+ Large ears with thick helix Mild hypotelorism
Severe ID, VIQ 26 NVIQ No obvious ID, VIQ 91 42 NVIQ 66 NA NA Single words Speech delay + + +
Slight ID: VIQ 66 PIQ 55 Severe ID
NA Few words 14 y + +
NA Speech delay + +
NA Absent +
15 m First words 2 y +
2y Absent NA NA
NA 1.5 NA NA
NA 2.7 + F, epil. NA
NA 3.8 +F NA
<2 3 + F (infancy) Large simple ears
3 3 +F Large dysplastic ears
3 <3 + F (12 m), epil., AED+ Large ears
NA
NA
NA
Enophthalmia
e
e
Thick lower lip
Micrognathia, thick NA lips, prominentincisors
NA
NA
e
Micrognathia, long Smoothphiltrum, thin philtrum, thin upper lip upper lip
e
e
NA
NA
Polydactyly
Hallux valgus of feet
e
e
Bulbous nose
NA
NA
NA
Bitemporal narrowing, pointed nasal tip
e
+ e
+ e
e Sleep disturbance
e NA
+ Neonatal hypotonia
ELV + CA
Normal (3 y)
Normal
NA
NA
+ + Bilateral inguinal e hernia Mild atrophic brain and Normal (3 m) medulla (24 y)
IUGR
4 6 + F (18 m), epil., AED+ Hypoplastic ear lobes Mild hypotelorism
face traits related to ancestry but also large ears, a small mouth and a long philtrum (Fig. 1D). His adaptative behavior profile assessed with the Vineland Adaptative Behavior Scale at 2.5 years old showed an equivalent-age of 11 months for daily living, 8e9 months for socialization and 8e9 months for motor skills. Further evaluation using the Childhood Autism Rating Scale (CARS) resulted in a score of 29.5, which is under the usual threshold of 30 for ASD. It must be stressed that i) we could not completely rule out a diagnosis of ASD; ii) some deficient features of the CARS score could be explained by global developmental delay (see Discussion section). At 3 years old, the skills and behavior of Patient #2 had progressed: he was able to say two syllable words, started to walk independently, paid attention to his peers though he still had hand stereotypies. Brain MRI performed at 7 months old because of microcephaly had revealed enlarged ventricles with large pericerebral spaces (Fig. 2C and D). Further investigations, including routine blood and urine chemistry, fragile X screening, metabolic workup, and aCGH, were normal.
3. Methods and results In Patient #1, a panel of 150 genes involved in epilepsy with or without ID has been sequenced by NGS (Life Tech technology: Ampli-seqÒ and Ion protonÒ) and showed the heterozygous nonsense c.613C >T (p.Arg205*) DYRK1A mutation (NM_001396.3).
van Bon et al. Clin Genet 2011
3
Møller et al. AJHG 2008
Polyhydramnios, microcephaly, IUGR
Shortened terminal phalanges Pectus excavatum, kyphosis, slight scoliosis + e ELV
The presence of the mutation was further confirmed by Sanger sequencing but was not found in the patient’s parents. This mutation has already been reported [Redin et al., 2014]. In Patient #2, NGS analysis using the TruSight One panel (Illumina) disclosed the c.932C >T (p.Ser311Phe) missense mutation in DYRK1A. This mutation is not reported in public databases (dbSNP, 1000 Genomes, EVS, ExAC). It alters highly conserved nucleotide and aminoacid residue (phyloP: 6.26 [14.1; 6.4]) and is predicted to be probably damaging in Polyphen-2 (score: 1.0) and to be deleterious in SIFT (score: 0). This mutation was confirmed by Sanger sequencing and absent from the parental genomes, thereby proving its de novo occurrence.
4. Discussion The involvement of DYRK1A in the ID of patients with Down syndrome brought the attention of many researchers on its role in brain development [Song et al., 1996]. The products of DYRK1A and its orthologs are expressed during different neurodevelopmental phases in invertebrates and vertebrates and appear as key elements in neuronal development [Dierssen and de Lagrán, 2006; Hämmerle et al., 2008]. The phenotype of Dyrk1a-deficient mice includes a decreased neonatal viability, significant body size reduction from birth, preweaning developmental delay, specific behavioral alteration and reduction of brain size in a region-specific manner [Fotaki et al., 2002]. In drosophila melanogaster, mutations
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Table 2 Summary of features reported in 13 DYRK1A patients. Features
Number of patients
Sex
6 7 3 1 4 1 2 2 4 1 1 5 4 1 2 1 10 5 2 9 2 9 3 7 2 2 4 4 4 8 4 6
Females Males Genetic anomalies Nonsense Missense Frameshift Splice-site Partial deletion Translocation + gene breakage Prenatal features IUGR Polyhydramnios Oligomanios Psychomotor development Severe Moderate ID/delay Mild delay Slight ID/delay No ID Behavioral disturbance Autistic traits Attention deficit OFC Normal Microcephaly Undecided (2 SD) Seizures Febrile seizures Confirmed epilepsy Morphologic features Large/dysplastic ears Enophthalmia Hypotelorism Micrognathia Long or smooth philtrum Thin upper lip Feeding difficulties during the first months of life Brain images Normal Cerebral atrophy
IUGR: intrauterine growth retardation. ID: intellectual disability. OFC: occipitofrontal circumference.
in minibrain (Mnb), ortholog of the human DYRK1A, cause a specific and marked size reduction of the optic lobes and central brain hemispheres [Tejedor et al., 1995]. Recent findings proved that DYRK1A has a crucial role in human brain development as well. The first clinical description of the DYRK1A-related phenotype traces back to 2008 with two patients with developmental delay harboring balanced translocations and a breakpoint interrupting DYRK1A [Møller et al., 2008]. Several patients with a large deletion encompassing DYRK1A [Fujita et al., 2010; Valetto et al., 2012; Yamamoto et al., 2011] were reported, which allowed to incriminate it in their developmental phenotype. Eleven patients with genetic aberrations involving the DYRK1A gene only have been described since then (Table 1). Developmental disorder and/ or ID with varying degrees of severity were reported in all of these 11 patients, with some of them being too young to make a definitive diagnosis. Five patients had severe ID (patient D12 in [Courcet et al., 2012], patient 13890.p1 in [O’Roak et al., 2012], patient #2 in [Møller et al., 2008], and [Okamoto et al., 2014; Van Bon et al., 2011]), five others had mild/moderate ID or mildly/ moderately delayed psychomotor development (the two patients reported here, [Redin et al., 2014], index case in [Courcet et al., 2012]), whereas two patients had slight psychomotor delay (patient #1 in [Møller et al., 2008], patient 12099.p1 in [O’Roak et al., 2012]) and one had no ID (according to reported intellectual quotient scores; patient 13552.p1 in [O’Roak et al., 2012]). Like the two patients reported here, the behavior of reported patients with DYRK1A mutations/deletions includes manifestations of the autism spectrum (mainly speech delay/ absence of language, impaired social interactions, stereotypies), sometimes associated with hyperactivity and impulsiveness
Fig. 1. Photographs of the patients at 3 years. AeC: Morphologic features in Patient #1: large ears, microretrognatism, frontal bossing, thin upper lip, long eyelashes, smooth philtrum, plagiocephaly, low columella, microdontia with dental diastasis (A and B); 2e3 toe syndactyly (C). D: Morphologic features in Patient #2 with Asian ancestry: small mouth, long philtrum, large ears and (likely ethnic) epicanthal folds.
[Courcet et al., 2012; O’Roak et al., 2012; Redin et al., 2014] or self-injuries [Okamoto et al., 2014]. Five patients were diagnosed with ASD or autism. As previously outlined, the diagnosis of ASD based on commonly used scales, such as the CARS, is negatively correlated with the level of cognitive and adaptative functioning [Perry et al., 2005]. It seems that most patients with DYRK1A mutations have autistic behaviors in a context of ID rather than typical autism according to canonical criteria. However, it is worth mentioning that ASD with attention deficit/hyperactivity disorder was diagnosed in the only patient with a preserved verbal intellectual quotient known to date (patient 13552.p1 in [O’Roak et al., 2012]). Thus, DYRK1A is one of the numerous genes recently implicated in developmental disorder and/or ID with different occurrence rates according to the studied populations: 1.9% (2/106) of unselected individuals with ID (negative for aCGH, fragile-X and other specific genetic analyses) [Redin et al., 2014], 1.1% (6/526) of individuals with ID or autism [Yang et al., 2014], 0.95% (1/105) of individuals with ID among which 74% had microcephaly [Courcet et al., 2012], but only 0.12% (3/2446) of individuals with ASD [O’Roak et al., 2012]. Consequently, the overall incidence of DYRK1A mutations in patients with ID (excluding ASD) selected for NGS analyses is close to 1.2%. Together with previous reports, our patients’ description highlights clinical data that may help recognize the phenotype. Prenatal findings were reported in six patients but were nonspecific:
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Fig. 2. Brain MRI of the two patients with DYRK1A mutations. A and B: Brain MRI of Patient #1 at 2 years 8 months. Coronal (A) and sagittal (B) T2-weighted images showing enlarged lateral ventricles and large pericerebral spaces. C and D: Brain MRI of Patient #2 at 7 months. T2-weighted coronal (C) and T1-weighted sagittal section showing enlarged lateral ventricles and large cerebral spaces.
intrauterine growth retardation in five (patient #2 and all patients in [Courcet et al., 2012; Møller et al., 2008], oligoamnios and polyhydramnios in one patient each (patient 13890.p1 in [O’Roak et al., 2012] and patient #2 in [Møller et al., 2008], respectively). Eight patients, including Patient #1 reported here, had significant feeding difficulties (due at least to gastroesophageal reflux) during the first months of life [Courcet et al., 2012; Møller et al., 2008; O’Roak et al., 2012; Redin et al., 2014; Van Bon et al., 2011]. Nutritional assistance was necessary for our patient but not mentioned in other reports. Most patients (9/12) experienced epileptic seizures. It seems that all of them first had short febrile seizures during infancy
Fig. 3. Schematic representation of the DYRK1A protein. Boxes represent the different elements of the structure of the protein: the nuclear localization signal (NLS), the kinase domain, the PEST domain, the His repeat and the S/T-rich region. Nonsense, missense and frameshift mutations so far reported are indicated. Large deletions and gene interruptions are not represented.
(present report, [Courcet et al., 2012; Møller et al., 2008; Van Bon et al., 2011]), and that three of them had afebrile seizures afterwards (patient #2 in Møller et al., 2008; patient 13552.p1 in O’Roak et al., 2012, patient D12 in Courcet et al., 2012). Since the epilepsy was not described in most cases, it is not yet possible to specify seizure types, whether it was drug-responsive or not, etc. Like in Patient #2, microcephaly (OFC <3 SD) is usual (8/13), but not universal, in DYRK1A-mutated patients. Microcephaly may be present at birth [Courcet et al., 2012; Møller et al., 2008; Van Bon et al., 2011] and tends to worsen with time [Courcet et al., 2012; Van Bon et al., 2011], or it may appear during the first months of life (patient #2). Brain MRI was reportedly normal in 4/9 patients [Okamoto et al., 2014]; index case in [Courcet et al., 2012], patient 13890.p1 in [O’Roak et al., 2012], patient #1 in [Møller et al., 2008] has thin corpus callosum but he is only 3 month-old) and showed enlargement of lateral ventricles with or without cortical atrophy suggesting cerebral atrophy in 5/9 (our two patients, patient APN87 in [Redin et al., 2014]; patient D12 in [Courcet et al., 2012], and [Møller et al., 2008]). The establishment of a relationship between cerebral atrophy and the severity of microcephaly is not obvious from available data and needs further reports. Dysmorphic features were found in our two patients and reported in six others. Noticeable features of the facial morphology include hypoplastic/low-set or large ear lobes (n ¼ 7), thin (sometimes thick) lips or thin upper lip (n ¼ 4), frontal bossing, epicanthal folds, flat nasal bridge, low columella, down-slanting palpebral fissures, long eyelashes, enophtalmia, short philtrum, downturned mouth, micrognathia, microdontia and high arched
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palate. Anomalies of limb extremities were found in some patients, including syndactyly (Patient #1), polydactyly [O’Roak et al., 2012], hallux valgus of feet [Van Bon et al., 2011] and arachnodactyly [Redin et al., 2014]. We report for the first time a patient with micropenis. DYRK1A is a complex proline-directed protein kinase that catalyzes the phosphorylation of multiple protein targets [Aranda et al., 2011; Dierssen and de Lagrán, 2006]. It contains several domains, including the catalytic domain (kinase domain), a nuclear localization signal at its N-terminus end, a PEST domain for protein degradation, a 13-consecutive-histidine repeat, and a S/Trich region with unknown function (Fig. 3). It undergoes selfactivation through autophosphorylation at Tyr in position 321 (located in the kinase domain), which is crucial for its intracellular distribution and compartment-specific functions [Kaczmarski et al., 2014]. Most DYRK1A mutations reported to date induce the production of a truncated mRNA with a predicted truncated protein or no protein if the nonsense mediated decay is elicited. These mutations, reported in Table 1, are five nonsense (patient #1 reported here, patient APN-58 in [Redin et al., 2014], 2 patients in [Yang et al., 2014] and [Okamoto et al., 2014]), five small insertions/deletions with a frameshift (patient APN-87 in [Redin et al., 2014], patient 13552 p.1 and 12099.p1 in [O’Roak et al., 2012] and [Courcet et al., 2012]), two larger deletions removing the 50 part of the gene (index case in [Courcet et al., 2012]) or part of the coding sequence [Van Bon et al., 2011], two splice site mutations (patient 13890.p1 in [O’Roak et al., 2012] and [Yang et al., 2014]) and two genic interruptions due to chromosome translocations [Møller et al., 2008]. In two cases [Yang et al., 2014], the mutation was inherited, once with proven parental mosaicism, but no information is available on the clinical status of the transmitter. Significantly, the developmental phenotype of patients with a large deletion encompassing DYRK1A is close to that of patients with the above-mentioned mutations [Fujita et al., 2010; Valetto et al., 2012; Yamamoto et al., 2011]. Furthermore, animal models showed that DYRK1A influences neuronal proliferation and differentiation with consequences on brain size in a dosagedependant manner [Dierssen and de Lagrán, 2006]. Thus, the human phenotype is likely due to haploinsufficiency, with the putative DYRK1A product from the mutated allele being either absent or enzymatically inactive. To date, phenotypical variations do not appear to be related to the location of the frameshift/nonsense mutation. Indeed, patients carrying the premature frameshift p.Ile48Lysfs*2 (patient 12099.p1 in [O’Roak et al., 2012]) and p.Ser97Cysfs*98 (patient D12 in [Courcet et al., 2012]) mutations relatively close to the 50 end of the coding sequence have slight ID and severe ID, respectively. On the other hand, the two patients with the most 30 nonsense mutations, i.e. p.Q567* [Okamoto et al., 2014] and p.Ala498Profs*94 (patient 13552.p1 in [O’Roak et al., 2012]), have severe ID without microcephaly and ASD with microcephaly but without ID, respectively. However, the effect of the mutation may differ depending on whether the protein is truncated (produced) or null (not produced) due to NMD. It is also possible that some mutations have different effects on the different isoforms of DYRK1A. The only known recurrent mutation is the p.Arg205* (c.613C >T) nonsense mutation found in Patient #1 reported here and in patient ANP-58 [Redin et al., 2014]. Both had feeding difficulties during infancy and have low-normal OFC (2 SD) and mild/moderate developmental delay or ID. The two previously reported aminoacid substitutions were not described [Yang et al., 2014] and cannot be discussed. The phenotype of Patient #2 with the p.Ser311Phe missense mutation is not less severe than others since it comprises
seizures, microcephaly and developmental delay. Interestingly, the Ser311 residue lies within the kinase domain 10 residues upstream of the crucial Tyr321 and is conserved across different DYRK proteins in mammals [Aranda et al., 2011]. It belongs to the cation binding site that chelates Mg2þ, the metal cation that bridges the ATP beta- and gamma-phosphates and aids in the orientation of the transferable phosphate. We speculate that the p.Ser311Phe substitution may induce the loss of DYRK1A kinase activity due to its inability to transfer the available phosphate mioety for the phosphorylation of target proteins. Obviously, more data are needed to understand the effect of DYRK1A mutations on brain development and to correlate mutations and phenotypes. References Aranda S, Laguna A, de la Luna S. DYRK family of protein kinases: evolutionary relationships, biochemical properties, and functional roles. FASEB J 2011;25: 449e62. http://dx.doi.org/10.1096/fj.10-165837. Courcet J-B, Faivre L, Malzac P, Masurel-Paulet A, Lopez E, Callier P, et al. The DYRK1A gene is a cause of syndromic intellectual disability with severe microcephaly and epilepsy. J Med Genet 2012;49:731e6. http://dx.doi.org/ 10.1136/jmedgenet-2012-101251. Dierssen M, de Lagrán MM. DYRK1A (dual-specificity tyrosine-phosphorylated and -regulated kinase 1A): a gene with dosage effect during development and neurogenesis. Sci World J 2006;6:1911e22. http://dx.doi.org/10.1100/ tsw.2006.319. Fotaki V, Dierssen M, Alcántara S, Martínez S, Martí E, Casas C, et al. Dyrk1A haploinsufficiency affects viability and causes developmental delay and abnormal brain morphology in mice. Mol Cell Biol 2002;22:6636e47. Fujita H, Torii C, Kosaki R, Yamaguchi S, Kudoh J, Hayashi K, et al. Microdeletion of the Down syndrome critical region at 21q22. Am J Med Genet Part A 2010;152A: 950e3. http://dx.doi.org/10.1002/ajmg.a.33228. García-Cerro S, Martínez P, Vidal V, Corrales A, Flórez J, Vidal R, et al. Overexpression of Dyrk1A is implicated in several cognitive, electrophysiological and neuromorphological alterations found in a mouse model of Down syndrome. PloS One 2014:9ee106572. http://dx.doi.org/10.1371/ journal.pone.0106572. Hämmerle B, Elizalde C, Tejedor FJ. The spatio-temporal and subcellular expression of the candidate Down syndrome gene Mnb/Dyrk1A in the developing mouse brain suggests distinct sequential roles in neuronal development. Eur J Neurosci 2008;27:1061e74. http://dx.doi.org/10.1111/j.1460-9568.2008.06092.x. Kaczmarski W, Barua M, Mazur-Kolecka B, Frackowiak J, Dowjat W, Mehta P, et al. Intracellular distribution of differentially phosphorylated dual-specificity tyrosine phosphorylation-regulated kinase 1A (DYRK1A). J Neurosci Res 2014;92:162e73. http://dx.doi.org/10.1002/jnr.23279. Kang JE, Choi SA, Park JB, Chung KC. Regulation of the proapoptotic activity of huntingtin interacting protein 1 by Dyrk1 and caspase-3 in hippocampal neuroprogenitor cells. 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Perry A, Condillac RA, Freeman NL, Dunn-Geier J, Belair J. Multi-site study of the Childhood Autism Rating Scale (CARS) in five clinical groups of young children. J Autism Dev Disord 2005;35:625e34. http://dx.doi.org/10.1007/s10803-0050006-9. Redin C, Gérard B, Lauer J, Herenger Y, Muller J, Quartier A, et al. Efficient strategy for the molecular diagnosis of intellectual disability using targeted highthroughput sequencing. J Med Genet 2014;51:724e36. http://dx.doi.org/ 10.1136/jmedgenet-2014-102554. Ryoo S-R, Jeong HK, Radnaabazar C, Yoo J-J, Cho H-J, Lee H-W, et al. DYRK1Amediated hyperphosphorylation of Tau. A functional link between Down syndrome and Alzheimer disease. J Biological Chem 2007;282:34850e7. http:// dx.doi.org/10.1074/jbc.M707358200. Song WJ, Sternberg LR, Kasten-Sportès C, Keuren ML, Chung SH, Slack AC, et al. Isolation of human and murine homologues of the Drosophila minibrain gene: human homologue maps to 21q22.2 in the Down syndrome “critical region”. 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L. Ruaud et al. / European Journal of Medical Genetics xxx (2015) 1e7 Tejedor F, Zhu XR, Kaltenbach E, Ackermann A, Baumann A, Canal I, et al. Minibrain: a new protein kinase family involved in postembryonic neurogenesis in Drosophila. Neuron 1995;14:287e301. Valetto A, Orsini A, Bertini V, Toschi B, Bonuccelli A, Simi F, et al. Molecular cytogenetic characterization of an interstitial deletion of chromosome 21 (21q22.13q22.3) in a patient with dysmorphic features, intellectual disability and severe generalized epilepsy. Eur J Med Genet 2012;55:362e6. http:// dx.doi.org/10.1016/j.ejmg.2012.03.011. Van Bon BWM, Hoischen A, Hehir-Kwa J, de Brouwer APM, Ruivenkamp C, Gijsbers ACJ, et al. Intragenic deletion in DYRK1A leads to mental retardation
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and primary microcephaly. Clin Genet 2011;79:296e9. http://dx.doi.org/ 10.1111/j.1399-0004.2010.01544.x. Yamamoto T, Shimojima K, Nishizawa T, Matsuo M, Ito M, Imai K. Clinical manifestations of the deletion of Down syndrome critical region including DYRK1A and KCNJ6. Am J Med Genet Part A 2011;155A:113e9. http://dx.doi.org/10.1002/ ajmg.a.33735. Yang Y, Muzny DM, Xia F, Niu Z, Person R, Ding Y, et al. Molecular findings among patients referred for clinical whole-exome sequencing. JAMA 2014. http:// dx.doi.org/10.1001/jama.2014.14601.
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Contents lists available at ScienceDirect
European Journal of Medical Genetics journal homepage: http://www.elsevier.com/locate/ejmg
Clinical report
DYRK1A mutations in two unrelated patients Q2
Lyse Ruaud a, Cyril Mignot b, Agnès Guët c, Christelle Ohl d, Caroline Nava b, Delphine Héron b, Boris Keren b, Christel Depienne b, Valérie Benoit e, Isabelle Maystadt e, Damien Lederer e, Daniel Amsallem f, Juliette Piard a, * a
Centre de Génétique Humaine, CHU Besançon, France Département de Génétique, APHP, GH Pitié-Salpêtrière, Centre de Référence des Déficiences Intellectuelles de Causes Rares, Paris, France c Service de Pédiatrie, APHP, Hôpital Louis Mourier, Colombes, France d Centre d’Action Médico-Sociale Précoce, Courbevoie, France e Institut de pathologie et de Génétique, Gosselies, Belgique f Service de Pédiatrie, CHU Besançon, France b
a r t i c l e i n f o
a b s t r a c t
Article history: Received 6 December 2014 Accepted 26 December 2014 Available online xxx
The Dual-specify tyrosine phosphorylation-regulated kinase 1A (DYRK1A) gene has been extensively studied for its role in the pathophysiology of intellectual disability (ID) in Down syndrome. The rise of next generation sequencing (NGS) and array-CGH (aCGH) in diagnostic settings for the evaluation of patients with ID allowed the identification of 17 patients carrying heterozygous genetic aberrations involving DYRK1A to date. The rate of DYRK1A mutations in this population reaches >1% in published NGS studies. The current report aims at further defining the phenotype of this encephalopathy with the detailed report of two unrelated patients. Both patients were boys with developmental delay, febrile seizures, facial dysmorphism and brain atrophy on MRI. Patient #1 had autistic behaviors and micropenis and Patient #2 had stereotypies and microcephaly. NGS analyses identified heterozygous de novo variants in DYRK1A: the c.613C >T (p.Arg205*) nonsense mutation in Patient #1 and the c.932C >T (p.Ser311Phe) missense mutation in Patient #2. Together with previously reported cases, patients with DYRK1A mutations share many clinical features and may have a recognizable phenotype that includes, by decreasing order of frequency: developmental delay or ID with behaviors suggesting autism spectrum disorder, microcephaly, epileptic seizures, facial dysmorphism including ear anomalies (large ears, hypoplastic lobes), thin lips, short philtrum and frontal bossing. Delineation of the phenotype/genotype correlation is not feasible at the moment and will be a challenge for the coming years. Ó 2015 Elsevier Masson SAS. All rights reserved.
Keywords: DYRK1A mutation Dysmorphism Intellectual disability Autism spectrum disorder
1. Introduction The Dual-specify tyrosine phosphorylation-regulated kinase 1A (DYRK1A) gene is located on chromosome 21q22.2 and the role of its product, the DYRK1A protein, in neuronal progenitor proliferation and neuronal differentiation is a growing field of research. It has been involved in the pathogenesis of Alzheimer’s disease [Ryoo et al., 2007], Parkinson’s disease [Kim et al., 2006] and Huntington disease [Kang et al., 2005], as well as in the intellectual disability
* Corresponding author. Centre de Génétique Humaine, Centre-hospitalouniversitaire, Pavillon Saint-Paul, 2 place Saint-Jacques, 25000 Besançon, France. Tel.: þ33 381218187; fax: þ33 381218643. E-mail address: [email protected] (J. Piard).
(ID) of patients with Down syndrome [García-Cerro et al., 2014]. The crucial role of DYRK1A in cerebral development has been further highlighted by the report of patients with cognitive impairment since infancy related to deletions or mutations within the gene (MIM #614104). To date, 11 patients with developmental delay/ID or autism spectrum disorder (ASD) and de novo heterozygous genetic aberrations in DYRK1A have been clinically well described (Table 1). Two of them harbored partial deletions of DYRK1A identified by arrayCGH (aCGH) [Courcet et al., 2012; Van Bon et al., 2011], two carried a balanced translocation inducing DYRK1A truncation [Møller et al., 2008] and seven carried frameshift, nonsense or splice site mutations identified by targeted [Courcet et al., 2012] or next generation sequencing (NGS) [Okamoto et al., 2014; O’Roak et al., 2012; Redin et al., 2014]. Six more patients with a
http://dx.doi.org/10.1016/j.ejmg.2014.12.014 1769-7212/Ó 2015 Elsevier Masson SAS. All rights reserved.
Please cite this article in press as: Ruaud L, et al., DYRK1A mutations in two unrelated patients, European Journal of Medical Genetics (2015), http://dx.doi.org/10.1016/j.ejmg.2014.12.014
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Table 1 Description of the 13 patients with DYRK1A-related encephalopathy. References
Present report
Patients Sex/age Genetic anomalies
Patient #1 M5y de novo nonsense c.613C >T, p.Arg205a
Prenatal findings
Okamoto et al. Clin Genet 2014
Redin et al. J Med Genet 2014
Patient #2 M3y de novo missense c.932C >T p.Ser311Phe
Patient 2 F7y Nonsense c.1699C >T p.Q567a
APN-58 M 16 y de novo nonsense c.613C >T p.Arg205a
APN-87 M5y de novo del/ins + fs c.621_624delinsGAA p.Glu208Asnfs*3
IUGR, single umbilical artery
NA
NA
NA
Moderately delayed
Severe ID
Moderate ID
Moderate ID
Unable to walk Absent + Self-injuries, temper tantrums, vocal tics NA +0.6 No Low-set ears
NA Speech delay +
NA NA +
NA 2 No “Dysmorphic traits”
NA 4 No “Dysmorphic traits”
Down-slanting palpebral fissures, epicanthal folds, hypertelorism Downturned mouth, short philtrum, micrognathia, thin upper lip e
e
e
e
e
Arachnodactyly
e
Psychom. dev.
Global development
Born at 33 WG with normal growth parameters Mildly delayed
Behavioral disturbance
Age of walking Language ASD/autistic behavior ADHD/attention deficit Other
25 m First words 3 y +
3y Two syllable words 3 y +/ (see text)
At birth Last exam Ears
Mean 2 + F (2.5 y) Large ears
1.5 <4 + F (16 m), AED+ Large ears
Eyes
Enophthalmia
e
Mouth
Microretrognathia, smooth philtrum, thin upper lip
Small mouth, long philtrum
Extremities
Syndactyly
e
Other
Frontal bossing, low columella, micropenis, microdontia + Bilateral inguinal hernias ELV + CA (2 y 8 m)
e
Frontal bossing, flat nasal bridge
Pectus excavatum
No Asthma, numerous otitis media ELV + CA (15 m)
No Short stature -3DS, severe amblyopia Normal
+ e
NA Sleep disturbance
NA
ELV
OFC (SD) Epilepsy/seizures (age) Morphologic features
Feeding difficultiesa Other features Brain MRI (age)
M: male. F: female. del: deletion. fs: frameshift. WG: weeks of gestation. IUGR: intra uterine growth retardation. m: months. y: years. Psychom. dev.: psychomotor development. ID: intellectual disability. VIQ: verbal intellectual quotient. NVIQ: non verbal intellectual quotient. ASD: autism spectrum disorder. ADHD: attention deficit hyperactivity disorder. OFC: occipitofrontal circumference. GTCS: generalized tonic clonic seizures. Epilepsy/seizures: + F means that the patient had febrile seizures. epil.: “confirmed” epilepsy. AED: anti-epileptic drug. ELV: enlarged lateral ventricles. CA: cortical atrophy. NA: not available. a Feeding difficulties during the first months of life.
“neurological phenotype” were reported but not described [Yang et al., 2014]. We report on two patients with de novo DYRK1A mutations, including the first described patient harboring a missense mutation, to delineate the phenotype of the DYRK1Aassociated developmental disorder Table 2. 2. Description of patients Patient #1 was the first child of healthy nonconsanguineous French parents. His mother had a miscarriage. His paternal grandfather has retinitis pigmentosa. He was born after an uneventful pregnancy at 33 weeks by vacuum-assisted delivery after a premature rupture of membranes. Birth weight was 1700 g (1 SD), birth length 41.8 cm (1 SD) and occipito-frontal circumference (OFC) 30.5 cm (mean). During the first months of life, he had congenital torticollis, sucking-swallowing difficulties requiring a feeding tube, episodes of bradycardia associated with gastroesophageal reflux. He underwent repair of bilateral inguinal hernias at 3 months old. A micropenis with a blind dimple on the scrotum was noticed without endocrine anomalies. He was efficiently treated with two injections of testosterone enanthate. One episode of febrile seizures occurred at 2.5 years old. The psychomotor development of Patient #1 was delayed: he sat at 10 months old, walked without assistance at 25 months and said his first words at 3 years old. Daytime toilet training was acquired at 4.5 years old. He had hand stereotypies, repetitive and sexual self-stimulation behaviors.
At first examination at 3 years, an unsteady gait and facial dysmorphism comprising large ears, microretrognatia, frontal bossing, thin upper lip, long eyelashes, smooth philtrum, plagiocephaly, low columella, microdontia with dental diastasis and enophtalmia (Fig. 1A and B), as well as bilateral 2e3 toe syndactyly (Fig. 1C), were noticed. At the age of 5 years, Patient #1 had normal growth parameters and an OFC of 49 cm (2 SD). He was treated by stretching plaster for bilateral equinus foot deformity. Brain MRI performed at 2 years and 8 months old showed diffuse cerebral atrophy (Fig. 2A and B). Standard blood chemistry and metabolic screening were negative as well as search for expansion in FMR1, ZEB2 sequencing and methylation at the SNRPN locus. Patient #2 was the first child of healthy nonconsanguineous parents of Asian and Caucasian descent. His paternal uncle had congenital deafness. During pregnancy, intrauterine growth retardation and a single umbilical artery were noticed. Karyotype was 46,XY on cultured amniocytes. Birth weight was 3170 g (25th centile), length 49 cm (1.5 SD) and OFC 33.5 cm (1.5 SD). Multiple episodes of bronchiolitis and otitis media occurred during infancy. He had a series of short generalized febrile seizures at 16 months old that did not recur under valproate therapy. When examined at that time, his psychomotor development was moderately delayed: he was not able to stand, could catch objects but did not manipulate them, babbled but did not say any word. His OFC was 42 cm (<4 SD) with marked slowing of head growth between 3 and 5 months old. He had Asian
Please cite this article in press as: Ruaud L, et al., DYRK1A mutations in two unrelated patients, European Journal of Medical Genetics (2015), http://dx.doi.org/10.1016/j.ejmg.2014.12.014
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Courcet et al. J Med Genet 2012
O’Roak et al. Science 2012
D12 F 14 y del. 2bp + fs c.290_291delCT p.Ser97Cysfs*98
Index case F4y de novo 69 kb del. arr21q22.13(37 644 501e37 713 641) 1 (hg18) IUGR
13890.p1 F 13.5 y de novo splice site mutation c.1098+1G >A
13552.p1 M6y de novo fs p.Ala498Profs*94
12099.p1 M8y de novo fs p.Ile48Lysfs*2
F 33 y Intragenic del.
Patient #1 Patient #2 M2y F 13 y de novo balanced translocation, breakpoint interrupting DYRK1A
Oligoamnios
NA
NA
NA
IUGR
Severe ID
Moderate ID
Slight retardation
Severe ID
3y NA + Inappropriate laughters 4,5 6 + F (12 m), AED+ Large ears with thick helix Mild hypotelorism
Severe ID, VIQ 26 NVIQ No obvious ID, VIQ 91 42 NVIQ 66 NA NA Single words Speech delay + + +
Slight ID: VIQ 66 PIQ 55 Severe ID
NA Few words 14 y + +
NA Speech delay + +
NA Absent +
15 m First words 2 y +
2y Absent NA NA
NA 1.5 NA NA
NA 2.7 + F, epil. NA
NA 3.8 +F NA
<2 3 + F (infancy) Large simple ears
3 3 +F Large dysplastic ears
3 <3 + F (12 m), epil., AED+ Large ears
NA
NA
NA
Enophthalmia
e
e
Thick lower lip
Micrognathia, thick NA lips, prominentincisors
NA
NA
e
Micrognathia, long Smoothphiltrum, thin philtrum, thin upper lip upper lip
e
e
NA
NA
Polydactyly
Hallux valgus of feet
e
e
Bulbous nose
NA
NA
NA
Bitemporal narrowing, pointed nasal tip
e
+ e
+ e
e Sleep disturbance
e NA
+ Neonatal hypotonia
ELV + CA
Normal (3 y)
Normal
NA
NA
+ + Bilateral inguinal e hernia Mild atrophic brain and Normal (3 m) medulla (24 y)
IUGR
4 6 + F (18 m), epil., AED+ Hypoplastic ear lobes Mild hypotelorism
face traits related to ancestry but also large ears, a small mouth and a long philtrum (Fig. 1D). His adaptative behavior profile assessed with the Vineland Adaptative Behavior Scale at 2.5 years old showed an equivalent-age of 11 months for daily living, 8e9 months for socialization and 8e9 months for motor skills. Further evaluation using the Childhood Autism Rating Scale (CARS) resulted in a score of 29.5, which is under the usual threshold of 30 for ASD. It must be stressed that i) we could not completely rule out a diagnosis of ASD; ii) some deficient features of the CARS score could be explained by global developmental delay (see Discussion section). At 3 years old, the skills and behavior of Patient #2 had progressed: he was able to say two syllable words, started to walk independently, paid attention to his peers though he still had hand stereotypies. Brain MRI performed at 7 months old because of microcephaly had revealed enlarged ventricles with large pericerebral spaces (Fig. 2C and D). Further investigations, including routine blood and urine chemistry, fragile X screening, metabolic workup, and aCGH, were normal.
3. Methods and results In Patient #1, a panel of 150 genes involved in epilepsy with or without ID has been sequenced by NGS (Life Tech technology: Ampli-seqÒ and Ion protonÒ) and showed the heterozygous nonsense c.613C >T (p.Arg205*) DYRK1A mutation (NM_001396.3).
van Bon et al. Clin Genet 2011
3
Møller et al. AJHG 2008
Polyhydramnios, microcephaly, IUGR
Shortened terminal phalanges Pectus excavatum, kyphosis, slight scoliosis + e ELV
The presence of the mutation was further confirmed by Sanger sequencing but was not found in the patient’s parents. This mutation has already been reported [Redin et al., 2014]. In Patient #2, NGS analysis using the TruSight One panel (Illumina) disclosed the c.932C >T (p.Ser311Phe) missense mutation in DYRK1A. This mutation is not reported in public databases (dbSNP, 1000 Genomes, EVS, ExAC). It alters highly conserved nucleotide and aminoacid residue (phyloP: 6.26 [14.1; 6.4]) and is predicted to be probably damaging in Polyphen-2 (score: 1.0) and to be deleterious in SIFT (score: 0). This mutation was confirmed by Sanger sequencing and absent from the parental genomes, thereby proving its de novo occurrence.
4. Discussion The involvement of DYRK1A in the ID of patients with Down syndrome brought the attention of many researchers on its role in brain development [Song et al., 1996]. The products of DYRK1A and its orthologs are expressed during different neurodevelopmental phases in invertebrates and vertebrates and appear as key elements in neuronal development [Dierssen and de Lagrán, 2006; Hämmerle et al., 2008]. The phenotype of Dyrk1a-deficient mice includes a decreased neonatal viability, significant body size reduction from birth, preweaning developmental delay, specific behavioral alteration and reduction of brain size in a region-specific manner [Fotaki et al., 2002]. In drosophila melanogaster, mutations
Please cite this article in press as: Ruaud L, et al., DYRK1A mutations in two unrelated patients, European Journal of Medical Genetics (2015), http://dx.doi.org/10.1016/j.ejmg.2014.12.014
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Table 2 Summary of features reported in 13 DYRK1A patients. Features
Number of patients
Sex
6 7 3 1 4 1 2 2 4 1 1 5 4 1 2 1 10 5 2 9 2 9 3 7 2 2 4 4 4 8 4 6
Females Males Genetic anomalies Nonsense Missense Frameshift Splice-site Partial deletion Translocation + gene breakage Prenatal features IUGR Polyhydramnios Oligomanios Psychomotor development Severe Moderate ID/delay Mild delay Slight ID/delay No ID Behavioral disturbance Autistic traits Attention deficit OFC Normal Microcephaly Undecided (2 SD) Seizures Febrile seizures Confirmed epilepsy Morphologic features Large/dysplastic ears Enophthalmia Hypotelorism Micrognathia Long or smooth philtrum Thin upper lip Feeding difficulties during the first months of life Brain images Normal Cerebral atrophy
IUGR: intrauterine growth retardation. ID: intellectual disability. OFC: occipitofrontal circumference.
in minibrain (Mnb), ortholog of the human DYRK1A, cause a specific and marked size reduction of the optic lobes and central brain hemispheres [Tejedor et al., 1995]. Recent findings proved that DYRK1A has a crucial role in human brain development as well. The first clinical description of the DYRK1A-related phenotype traces back to 2008 with two patients with developmental delay harboring balanced translocations and a breakpoint interrupting DYRK1A [Møller et al., 2008]. Several patients with a large deletion encompassing DYRK1A [Fujita et al., 2010; Valetto et al., 2012; Yamamoto et al., 2011] were reported, which allowed to incriminate it in their developmental phenotype. Eleven patients with genetic aberrations involving the DYRK1A gene only have been described since then (Table 1). Developmental disorder and/ or ID with varying degrees of severity were reported in all of these 11 patients, with some of them being too young to make a definitive diagnosis. Five patients had severe ID (patient D12 in [Courcet et al., 2012], patient 13890.p1 in [O’Roak et al., 2012], patient #2 in [Møller et al., 2008], and [Okamoto et al., 2014; Van Bon et al., 2011]), five others had mild/moderate ID or mildly/ moderately delayed psychomotor development (the two patients reported here, [Redin et al., 2014], index case in [Courcet et al., 2012]), whereas two patients had slight psychomotor delay (patient #1 in [Møller et al., 2008], patient 12099.p1 in [O’Roak et al., 2012]) and one had no ID (according to reported intellectual quotient scores; patient 13552.p1 in [O’Roak et al., 2012]). Like the two patients reported here, the behavior of reported patients with DYRK1A mutations/deletions includes manifestations of the autism spectrum (mainly speech delay/ absence of language, impaired social interactions, stereotypies), sometimes associated with hyperactivity and impulsiveness
Fig. 1. Photographs of the patients at 3 years. AeC: Morphologic features in Patient #1: large ears, microretrognatism, frontal bossing, thin upper lip, long eyelashes, smooth philtrum, plagiocephaly, low columella, microdontia with dental diastasis (A and B); 2e3 toe syndactyly (C). D: Morphologic features in Patient #2 with Asian ancestry: small mouth, long philtrum, large ears and (likely ethnic) epicanthal folds.
[Courcet et al., 2012; O’Roak et al., 2012; Redin et al., 2014] or self-injuries [Okamoto et al., 2014]. Five patients were diagnosed with ASD or autism. As previously outlined, the diagnosis of ASD based on commonly used scales, such as the CARS, is negatively correlated with the level of cognitive and adaptative functioning [Perry et al., 2005]. It seems that most patients with DYRK1A mutations have autistic behaviors in a context of ID rather than typical autism according to canonical criteria. However, it is worth mentioning that ASD with attention deficit/hyperactivity disorder was diagnosed in the only patient with a preserved verbal intellectual quotient known to date (patient 13552.p1 in [O’Roak et al., 2012]). Thus, DYRK1A is one of the numerous genes recently implicated in developmental disorder and/or ID with different occurrence rates according to the studied populations: 1.9% (2/106) of unselected individuals with ID (negative for aCGH, fragile-X and other specific genetic analyses) [Redin et al., 2014], 1.1% (6/526) of individuals with ID or autism [Yang et al., 2014], 0.95% (1/105) of individuals with ID among which 74% had microcephaly [Courcet et al., 2012], but only 0.12% (3/2446) of individuals with ASD [O’Roak et al., 2012]. Consequently, the overall incidence of DYRK1A mutations in patients with ID (excluding ASD) selected for NGS analyses is close to 1.2%. Together with previous reports, our patients’ description highlights clinical data that may help recognize the phenotype. Prenatal findings were reported in six patients but were nonspecific:
Please cite this article in press as: Ruaud L, et al., DYRK1A mutations in two unrelated patients, European Journal of Medical Genetics (2015), http://dx.doi.org/10.1016/j.ejmg.2014.12.014
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Fig. 2. Brain MRI of the two patients with DYRK1A mutations. A and B: Brain MRI of Patient #1 at 2 years 8 months. Coronal (A) and sagittal (B) T2-weighted images showing enlarged lateral ventricles and large pericerebral spaces. C and D: Brain MRI of Patient #2 at 7 months. T2-weighted coronal (C) and T1-weighted sagittal section showing enlarged lateral ventricles and large cerebral spaces.
intrauterine growth retardation in five (patient #2 and all patients in [Courcet et al., 2012; Møller et al., 2008], oligoamnios and polyhydramnios in one patient each (patient 13890.p1 in [O’Roak et al., 2012] and patient #2 in [Møller et al., 2008], respectively). Eight patients, including Patient #1 reported here, had significant feeding difficulties (due at least to gastroesophageal reflux) during the first months of life [Courcet et al., 2012; Møller et al., 2008; O’Roak et al., 2012; Redin et al., 2014; Van Bon et al., 2011]. Nutritional assistance was necessary for our patient but not mentioned in other reports. Most patients (9/12) experienced epileptic seizures. It seems that all of them first had short febrile seizures during infancy
Fig. 3. Schematic representation of the DYRK1A protein. Boxes represent the different elements of the structure of the protein: the nuclear localization signal (NLS), the kinase domain, the PEST domain, the His repeat and the S/T-rich region. Nonsense, missense and frameshift mutations so far reported are indicated. Large deletions and gene interruptions are not represented.
(present report, [Courcet et al., 2012; Møller et al., 2008; Van Bon et al., 2011]), and that three of them had afebrile seizures afterwards (patient #2 in Møller et al., 2008; patient 13552.p1 in O’Roak et al., 2012, patient D12 in Courcet et al., 2012). Since the epilepsy was not described in most cases, it is not yet possible to specify seizure types, whether it was drug-responsive or not, etc. Like in Patient #2, microcephaly (OFC <3 SD) is usual (8/13), but not universal, in DYRK1A-mutated patients. Microcephaly may be present at birth [Courcet et al., 2012; Møller et al., 2008; Van Bon et al., 2011] and tends to worsen with time [Courcet et al., 2012; Van Bon et al., 2011], or it may appear during the first months of life (patient #2). Brain MRI was reportedly normal in 4/9 patients [Okamoto et al., 2014]; index case in [Courcet et al., 2012], patient 13890.p1 in [O’Roak et al., 2012], patient #1 in [Møller et al., 2008] has thin corpus callosum but he is only 3 month-old) and showed enlargement of lateral ventricles with or without cortical atrophy suggesting cerebral atrophy in 5/9 (our two patients, patient APN87 in [Redin et al., 2014]; patient D12 in [Courcet et al., 2012], and [Møller et al., 2008]). The establishment of a relationship between cerebral atrophy and the severity of microcephaly is not obvious from available data and needs further reports. Dysmorphic features were found in our two patients and reported in six others. Noticeable features of the facial morphology include hypoplastic/low-set or large ear lobes (n ¼ 7), thin (sometimes thick) lips or thin upper lip (n ¼ 4), frontal bossing, epicanthal folds, flat nasal bridge, low columella, down-slanting palpebral fissures, long eyelashes, enophtalmia, short philtrum, downturned mouth, micrognathia, microdontia and high arched
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palate. Anomalies of limb extremities were found in some patients, including syndactyly (Patient #1), polydactyly [O’Roak et al., 2012], hallux valgus of feet [Van Bon et al., 2011] and arachnodactyly [Redin et al., 2014]. We report for the first time a patient with micropenis. DYRK1A is a complex proline-directed protein kinase that catalyzes the phosphorylation of multiple protein targets [Aranda et al., 2011; Dierssen and de Lagrán, 2006]. It contains several domains, including the catalytic domain (kinase domain), a nuclear localization signal at its N-terminus end, a PEST domain for protein degradation, a 13-consecutive-histidine repeat, and a S/Trich region with unknown function (Fig. 3). It undergoes selfactivation through autophosphorylation at Tyr in position 321 (located in the kinase domain), which is crucial for its intracellular distribution and compartment-specific functions [Kaczmarski et al., 2014]. Most DYRK1A mutations reported to date induce the production of a truncated mRNA with a predicted truncated protein or no protein if the nonsense mediated decay is elicited. These mutations, reported in Table 1, are five nonsense (patient #1 reported here, patient APN-58 in [Redin et al., 2014], 2 patients in [Yang et al., 2014] and [Okamoto et al., 2014]), five small insertions/deletions with a frameshift (patient APN-87 in [Redin et al., 2014], patient 13552 p.1 and 12099.p1 in [O’Roak et al., 2012] and [Courcet et al., 2012]), two larger deletions removing the 50 part of the gene (index case in [Courcet et al., 2012]) or part of the coding sequence [Van Bon et al., 2011], two splice site mutations (patient 13890.p1 in [O’Roak et al., 2012] and [Yang et al., 2014]) and two genic interruptions due to chromosome translocations [Møller et al., 2008]. In two cases [Yang et al., 2014], the mutation was inherited, once with proven parental mosaicism, but no information is available on the clinical status of the transmitter. Significantly, the developmental phenotype of patients with a large deletion encompassing DYRK1A is close to that of patients with the above-mentioned mutations [Fujita et al., 2010; Valetto et al., 2012; Yamamoto et al., 2011]. Furthermore, animal models showed that DYRK1A influences neuronal proliferation and differentiation with consequences on brain size in a dosagedependant manner [Dierssen and de Lagrán, 2006]. Thus, the human phenotype is likely due to haploinsufficiency, with the putative DYRK1A product from the mutated allele being either absent or enzymatically inactive. To date, phenotypical variations do not appear to be related to the location of the frameshift/nonsense mutation. Indeed, patients carrying the premature frameshift p.Ile48Lysfs*2 (patient 12099.p1 in [O’Roak et al., 2012]) and p.Ser97Cysfs*98 (patient D12 in [Courcet et al., 2012]) mutations relatively close to the 50 end of the coding sequence have slight ID and severe ID, respectively. On the other hand, the two patients with the most 30 nonsense mutations, i.e. p.Q567* [Okamoto et al., 2014] and p.Ala498Profs*94 (patient 13552.p1 in [O’Roak et al., 2012]), have severe ID without microcephaly and ASD with microcephaly but without ID, respectively. However, the effect of the mutation may differ depending on whether the protein is truncated (produced) or null (not produced) due to NMD. It is also possible that some mutations have different effects on the different isoforms of DYRK1A. The only known recurrent mutation is the p.Arg205* (c.613C >T) nonsense mutation found in Patient #1 reported here and in patient ANP-58 [Redin et al., 2014]. Both had feeding difficulties during infancy and have low-normal OFC (2 SD) and mild/moderate developmental delay or ID. The two previously reported aminoacid substitutions were not described [Yang et al., 2014] and cannot be discussed. The phenotype of Patient #2 with the p.Ser311Phe missense mutation is not less severe than others since it comprises
seizures, microcephaly and developmental delay. Interestingly, the Ser311 residue lies within the kinase domain 10 residues upstream of the crucial Tyr321 and is conserved across different DYRK proteins in mammals [Aranda et al., 2011]. It belongs to the cation binding site that chelates Mg2þ, the metal cation that bridges the ATP beta- and gamma-phosphates and aids in the orientation of the transferable phosphate. We speculate that the p.Ser311Phe substitution may induce the loss of DYRK1A kinase activity due to its inability to transfer the available phosphate mioety for the phosphorylation of target proteins. Obviously, more data are needed to understand the effect of DYRK1A mutations on brain development and to correlate mutations and phenotypes. References Aranda S, Laguna A, de la Luna S. DYRK family of protein kinases: evolutionary relationships, biochemical properties, and functional roles. FASEB J 2011;25: 449e62. http://dx.doi.org/10.1096/fj.10-165837. 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