European Journal of Human Genetics (2019) 27:1235–1243 https://doi.org/10.1038/s41431-019-0383-8 ARTICLE Phenotypic spectrum associated with a CRADD founder variant underlying frontotemporal predominant pachygyria in the Finnish population 1,2 3 4 5 6 1 Daniel L. Polla ● Elisa Rahikkala ● Michaela K. Bode ● Tuomo Määttä ● Teppo Varilo ● Thyrza Loman ● 6 7,8,9 7,8,9,10,11 12 13 14 Anju K. Philips ● Mitja Kurki ● Aarno Palotie ● Jarmo Körkkö ● Päivi Vieira ● Kristiina Avela ● 15 15 15 1 3,9 Valérie Jacquemin ● Isabelle Pirson ● Marc Abramowicz ● Arjan P. M. de Brouwer ● Outi Kuismin ● 1 6 Hans van Bokhoven ● Irma Järvelä Received: 28 September 2018 / Revised: 13 February 2019 / Accepted: 7 March 2019 / Published online: 26 March 2019 © European Society of Human Genetics 2019 Abstract Intellectual disability (ID), megalencephaly, frontal predominant pachygyria, and seizures, previously called “thin” lissencephaly, are reported to be caused by recessive variants in CRADD. Among five families of different ethnicities identified, one homozygous missense variant, c.509G>A p.(Arg170His), was of Finnish ancestry. Here we report on the 1234567890();,: 1234567890();,: phenotypic variability associated for this potential CRADD founder variant in 22 Finnish individuals. Exome sequencing was used to identify candidate genes in Finnish patients presenting with ID. Targeted Sanger sequencing and restriction enzyme analysis were applied to screen for the c.509G>A CRADD variant in cohorts from Finland. Detailed phenotyping and genealogical studies were performed. Twenty two patients were identified with the c.509G>A p.(Arg170His) homozygous variant in CRADD. The majority of the ancestors originated from Northeastern Finland indicating a founder effect. The hallmark of the disease is frontotemporal predominant pachygyria with mild cortical thickening. All patients show ID of variable severity. Aggressive behavior was found in nearly half of the patients, EEG abnormalities in five patients and megalencephaly in three patients. This study provides detailed data about the phenotypic spectrum of patients with lissencephaly due to a CRADD variant that affects function. High inter- and intrafamilial phenotypic heterogeneity was identified in patients with pachygyria caused by the homozygous CRADD founder variant. The phenotype variability suggests that additional genetic and/or environmental factors play a role in the clinical presentation. Since frontotemporal pachygyria is the hallmark of the disease, brain imaging studies are essential to support the molecular diagnosis for individuals with ID and a CRADD variant. Introduction The human brain is a highly complex structure and its normal development and function is critically dependent on the proper and tightly regulated activity of a large number of genes. Consequently, it is estimated that variants that affect These authors contributed equally: Daniel L. Polla, Elisa Rahikkala, Michaela K. Bode function in more than 2000 genes can give rise to intellectual disability (ID) [1]. Of these genes, about half are still These authors jointly supervised this work: Hans van Bokhoven, Irma unknown [1–3]. ID is clinically and genetically heterogeneous Järvelä and comprises several groups of neurodevelopmental dis- Supplementary information The online version of this article (https:// orders that in total affect 2–3% of the general population doi.org/10.1038/s41431-019-0383-8) contains supplementary [4, 5]. One of these groups is characterized by rare congenital material, which is available to authorized users. anomalies of the cerebral cortex caused by neuronal migration * Irma Järvelä defects, called malformations of cortical development irma.jarvela@helsinki.fi (MCDs) [6]. MCDs are classified into three major groups: Extended author information available on the last page of the article. malformations caused by abnormal cell proliferation, 1236 D. L. Polla et al. misdirected neuronal migration, or aberrant post-migrational c.509G>A p.(Arg170His) variant from Northern Finland cortical organization and connectivity [6, 7] and are usually Intellectual Disability (NFID) project [16] and two cases diagnosed by brain imaging such as MRI (magnetic identified directly in the clinic by one of the authors (ER) resonance imaging) [7]. To date, genetic variants that were included into the detailed phenotype analysis. Finally, affect function in more than 100 genes are reported to cause two Finnish siblings with pachygyria [17] belong to a col- MCDs [7]. lection of cases with prenatal ventriculomegaly and/or Lissencephaly (LIS) spectrum of diseases is associated congenital, primary hydrocephalus whose genetic back- with deficient neuronal migration and abnormal formation of ground was studied by the group of MA. cerebral convolutions or gyri [7–9]. The LIS spectrum of The growth and occipitofrontal head circumference diseases encompasses agyria, pachygyria, and subcortical (OFC) of the Finnish patients were compared with the age band heterotopia (SBH). In addition, LIS is found in syn- and sex matched Finnish standard reference curves [18]. dromic MCDs, such as Miller-Dieker syndrome [10]. Clinical The new Finnish reference for OFC was used for patients manifestations of LIS include developmental delay, ID, early between 0–7 years [19]. As the new reference was not hypotonia with subsequent spastic quadriplegia, and seizures standardized for patients older than 7 years, the old Finnish [11, 12]. To date, a total of 19 genes have been identified to reference was used for patients older than 7 years [20]. OFC underlie the LIS spectrum of brain malformations of which was compared to height and relative megalencephaly was the most common mutated genes are LIS1 and DCX [11]. defined as OFC more than 2 standard deviations (SD) above Based on the combination of brain imaging and molecular height centile for age. genetics data from several patients, a classification of LIS and The CRADD variant data were submitted to LOVD data- SBH described 21 different patterns of the disease, including base (www.LOVD.nl/CRADD) hosted at Leiden University common patterns such as partial agyria-pachygyria and pos- Medical Center, the Netherlands. The parents or legal guar- terior mild pachygyria [9]. Among the rare types of LIS is dians of all patients in this study provided written informed frontal predominant pachygyria with mild cortical thickening, consent. The study was approved by the ethics committees at also called thin pachygyria or thin lissencephaly (TLIS) [9]. the Helsinki University Central Hospital, Northern Ostro- Recently, four homozygous variants that affect function bothnia Hospital District, Hospital District of Helsinki and of the CRADD gene have been reported to cause TLIS Uusimaa, and the Radboud University Medical Center. (OMIM #614499) [13–15]. The phenotype was character- ized by TLIS, relative or absolute megalencephaly, and mild Exome sequencing (ES) to moderate intellectual disability with no other congenital anomalies. Three out of 15 (20%) patients presented with High-throughput sequencing was performed for sibling seizures [13, 14]. One of the variants, c.509G>A p. pairs in FIN-1 and FIN-2 families (Fig. 1) at BGI (BGI- (Arg170His) (GenBank: NM_003805.3), was identified in a Shenzhen, Shenzhen, China). Exome capturing was carried homozygous state in a female of Finnish ancestry [13]. out by using Agilent SureSelect Target Enrichment V5 In search for genes underlying intellectual disability (Agilent Technologies, Santa Clara, CA, USA). Sequencing in Finnish families, the homozygous c.509G>A p. was performed by using the Illumina HiSeq 2000 platform (Arg170His) variant that affects function in CRADD in 22 (Illumina, Inc. San Diego, CA). Illumina base calling soft- patients out of 15 Finnish families with ID was found. This ware v1.7 was employed to analyze the raw image files with suggests a founder variant in the isolated Finnish popula- default parameters. Data analysis were performed using the tion. The presence of an identical variant in a large number Roche Newbler software (v.2.3) using human genome build of individuals presents a unique opportunity to analyze the hg19/GRCh37. phenotypic variation that can be observed for a single Exome sequencing and targeted Sanger sequencing were monogenic condition. We present detailed clinical pheno- also used to analyze the data of 757 patients with ID of type and brain imaging findings of the families, showing unknown etiology belonging to the NFID cohort to identify the that pachygyria is the hallmark of the disease in patients CRADD founder variant in six additional cases with pachy- with the Finnish founder variant. gyria, see detailed description of the project in Kurki et al. [16]. Filtering and annotation Methods Selection of the variants for sibling pairs in FIN-1 and FIN-2 Patients were referred by physicians working in disability was performed by using a seven tier filtering strategy: (i) services and clinical geneticists. In total, 211 patients with inclusion of variants present in ≥2 reads and present in ≥70% ID of unknown cause were collected from Northeastern of all reads (ii) exclusion of variants present with a frequency Finland for the study. In addition, six cases homozygous for ≥1% in unaffected controls (Dept of Human Genetics, Phenotypic spectrum associated with a CRADD founder variant underlying frontotemporal predominant. 1237 I I I II II II III III III IV IV IV V V VI V VI 1 2534 67 8 VII VII FIN-3 VI VIII VIII 1 234 5 1 23 456 IX IX VII 1 23465 1234 5 6 789 10 1 FIN-1 FIN-5 FIN-10 FIN-2 FIN-4 FIN-6 FIN-12 I I I I II II II 1 23 4 1 2 II III III III III 1 1 234 123 IV FIN-8 FIN-9 FIN-11 V VI I I VII DNA available 1 2 3 4 Heterozygous II 1 234 II Homozygous VIII 123 FIN-7 1342 5 678910 FIN-15 ID, not investigated FIN-13 FIN-14 Fig. 1 Pedigrees of the families. Pedigree of the family indicating genotype of the individuals who were available for testing. The arrow indicates the proband in each pedigree where ES was performed Nijmegen, 12,000 controls), (iii) exclusion of variants within (MKB).