Brief Report: Syndromes in Autistic Children in a Finnish Birth Cohort.

We studied the association between specific congenital syndromes and autism spectrum disorders (ASD) in the large Finnish Register material. Our data include all children diagnosed with ASD (n = 4441) according to Finnish Hospital Discharge Register in 1987-2000. Four controls per each case were matched to sex, birthplace, date of birth (±30 days) and residence in Finland (n = 17,695). The prevalence of specific congenital syndromes in the Finnish Register of Congenital Malformations was evaluated among the ASD group and the controls by sex. The results of this study suggest that there is an association between several etiologically different syndromes and ASD when compared to controls without ASD. Statistically significant associations were observed with 47,XYY, Sotos syndrome, neurofibromatosis I, and syndrome not otherwise specified. Syndromes were more common among males with ASD compared to controls. These results support the previous studies of etiological heterogeneity of ASD and have importance in clinical examination, management and rehabilitation.

The landscape of copy number variations in Finnish families with autism spectrum disorders.

Rare de novo and inherited copy number variations (CNVs) have been implicated in autism spectrum disorder (ASD) risk. However, the genetic underpinnings of ASD remain unknown in more than 80% of cases. Therefore, identification of novel candidate genes and corroboration of known candidate genes may broaden the horizons of determining genetic risk alleles, and subsequent development of diagnostic testing. Here, using genotyping arrays, we characterized the genetic architecture of rare CNVs (<1% frequency) in a Finnish case-control dataset. Unsurprisingly, ASD cases harbored a significant excess of rare, large (>1 Mb) CNVs and rare, exonic CNVs. The exonic rare de novo CNV rate (∼22.5%) seemed higher compared to previous reports. We identified several CNVs in well-known ASD regions including GSTM1-5, DISC1, FHIT, RBFOX1, CHRNA7, 15q11.2, 15q13.2-q13.3, 17q12, and 22q11.21. Additionally, several novel candidate genes (BDKRB1, BDKRB2, AP2M1, SPTA1, PTH1R, CYP2E1, PLCD3, F2RL1, UQCRC2, LILRB3, RPS9, and COL11A2) were identified through gene prioritization. The majority of these genes belong to neuroactive ligand-receptor interaction pathways, and calcium signaling pathways, thus suggesting that a subset of these novel candidate genes may contribute to ASD risk. Furthermore, several metabolic pathways like caffeine metabolism, drug metabolism, retinol metabolism, and calcium-signaling pathway were found to be affected by the rare exonic ASD CNVs. Additionally, biological processes such as bradykinin receptor activity, endoderm formation and development, and oxidoreductase activity were enriched among the rare exonic ASD CNVs. Overall, our findings may add data about new genes and pathways that contribute to the genetic architecture of ASD.

The association between autism spectrum disorders and congenital anomalies by organ systems in a Finnish national birth cohort.

The aim of this study was to evaluate the association between autism spectrum disorders (ASD) with and without intellectual disability (ID) and congenital anomalies (CAs) by organ system. The sample included all children diagnosed with ASD (n = 4441) from the Finnish Hospital Discharge Register during 1987-2000 and a total of four controls per case (n = 17,695). CAs of the eye, central nervous system, and specific craniofacial anomalies were most strongly associated with ASD. Children with ASD and co-occurring ID were more likely to have CAs compared to ASD children without ID. The results suggest that some cases of ASD may originate during organogenesis, in the early first trimester of pregnancy. The results of this study may be useful for identifying prenatal etiological factors and elucidating the molecular pathogenesis of ASD.

Association and Promoter Analysis of AVPR1A in Finnish Autism Families.

The arginine vasopressin receptor 1A gene (AVPR1A) is known to affect social communication and has been reported to associate with autism in several studies. Given that the microsatellite RS1 and a few SNPs in the promoter region of the AVPR1A have repeatedly associated with several traits, including autism it is rather surprising that the molecular explanation for these associations has remained unknown, although it has been reported that the allele length of the AVPR1A microsatellites might affect disease risk. Here we carried out an extended association analysis of three microsatellites and 12 tag single nucleotide polymorphisms (SNPs) in and around the AVPR1A gene in 205 Finnish families followed by promoter analysis. FBAT version v2.0.3 was used for family-based genetic association analyses of AVPR1A microsatellites and SNPs. The nearby microsatellite RS1 was found to harbor the best association. Interestingly, there are two potentially relevant transcription factor (TF) binding sites at RS1: for MEF2C and PBX, predicted with the Match algorithm in the TRANSFAC database. Sequence variations changing the affinity of these TFs might partly explain the AVPR1A promoter region associations shown in autism.

The association between congenital anomalies and autism spectrum disorders in a Finnish national birth cohort.

AIM: The first aim of this study was to evaluate the association between different subgroups of autism spectrum disorders (ASDs) (childhood autism, Asperger syndrome, and pervasive developmental disorder/pervasive developmental disorder - not otherwise specified [PDD/PDD-NOS]) and congenital anomalies. Second, we assessed the association among intellectually disabled children with ASDs in the subgroups of childhood autism and PDD/PDD-NOS. METHOD: Nationwide population-based register data for children with a diagnosis of ASD (n=4449; 3548 males, 901 females) were collected during years 1987-2000 from the Finnish Hospital Discharge Register. Data on congenital anomalies were derived from the National Register of Congenital Malformations. Conditional logistic regression models were used as a statistical method. The association between ASD subgroups and congenital anomalies was stratified by the presence or absence of intellectual disability. RESULTS: Congenital anomalies occurred more frequently in all subgroups of ASD than in comparison participants (adjusted odds ratio [OR] for major congenital anomalies 1.8, 95% confidence interval [CI] 1.5-2.2, p<0.001). The association between congenital anomalies and childhood autism (OR 2.4, 95% CI 1.6-3.6, p<0.001) and between congenital anomalies and PDD/PDD-NOS (OR 3.7, 95% CI 2.4-5.7, p<0.001) among children with an intellectual disability was strong but remained significant also without intellectual disability (childhood autism: OR 1.7, 95% CI 1.3-2.3, p<0.001; PDD/PDD-NOS: OR 2.3, 95% CI 1.9-2.8, p<0.001). INTERPRETATION: The results suggest a significant association between ASDs and congenital anomalies regardless of the ASD subgroup. The association between childhood autism and PDD/PDD-NOS and congenital anomalies is stronger among children with intellectual disability is stronger than among those without intellectual disability. These results may have relevance in examining early risk factors in autism during fetal neurodevelopment.

The incidence of diagnosed autism spectrum disorders in Finland.

BACKGROUND: Previous reports indicate an increase in incidence of autism spectrum disorders (ASD). AIMS: First, to assess the incidence of diagnosed ASD in children born between 1996 and 1998, based on nationwide inpatient and outpatient register information. Second, to investigate the incidence rate over time of diagnosed ASD and specifically childhood autism, Asperger's syndrome and pervasive developmental disorder (PDD-NOS) in children born between 1987 and 1998. METHODS: This is population-based cohort study with children born in Finland between 1 January 1987 and 31 December 2005; a total of more than 1.2 million children. Children were identified in the Finnish Hospital Discharge Register, and the reported diagnoses were based on the International Statistical Classification of Diseases (ICD-10, ICD-9). RESULTS: The annual incidence rate of diagnosed ASD based on inpatient and outpatient register data was 53.7 per 10,000 (95% CI 50.4-57.2). Incidence was 82.6 per 10,000 in boys and 23.6 per 10,000 in girls, yielding a sex ratio (boys:girls) of 3.5:1. We report an eightfold increase in the incidence rates in children of diagnosed ASD and specifically in childhood autism, Asperger's syndrome and PDD-NOS and born between 1987 and 1992 based on inpatient register information. CONCLUSIONS: Increased awareness of ASD, more precise diagnostic criteria and changes in practice for diagnosing autism may have had a substantial effect on the increased incidence of inpatient treated ASD cases from 1987 to 1992. Between 1992 and 1998, the incidence rate based on inpatient and outpatient service use remained rather stable.

Genetic and functional analyses of SHANK2 mutations suggest a multiple hit model of autism spectrum disorders.

Autism spectrum disorders (ASD) are a heterogeneous group of neurodevelopmental disorders with a complex inheritance pattern. While many rare variants in synaptic proteins have been identified in patients with ASD, little is known about their effects at the synapse and their interactions with other genetic variations. Here, following the discovery of two de novo SHANK2 deletions by the Autism Genome Project, we identified a novel 421 kb de novo SHANK2 deletion in a patient with autism. We then sequenced SHANK2 in 455 patients with ASD and 431 controls and integrated these results with those reported by Berkel et al. 2010 (n = 396 patients and n = 659 controls). We observed a significant enrichment of variants affecting conserved amino acids in 29 of 851 (3.4%) patients and in 16 of 1,090 (1.5%) controls (P = 0.004, OR = 2.37, 95% CI = 1.23-4.70). In neuronal cell cultures, the variants identified in patients were associated with a reduced synaptic density at dendrites compared to the variants only detected in controls (P = 0.0013). Interestingly, the three patients with de novo SHANK2 deletions also carried inherited CNVs at 15q11-q13 previously associated with neuropsychiatric disorders. In two cases, the nicotinic receptor CHRNA7 was duplicated and in one case the synaptic translation repressor CYFIP1 was deleted. These results strengthen the role of synaptic gene dysfunction in ASD but also highlight the presence of putative modifier genes, which is in keeping with the "multiple hit model" for ASD. A better knowledge of these genetic interactions will be necessary to understand the complex inheritance pattern of ASD.

Fine mapping of Xq11.1-q21.33 and mutation screening of RPS6KA6, ZNF711, ACSL4, DLG3, and IL1RAPL2 for autism spectrum disorders (ASD).

About 80% of cases with autism express intellectual disability. Both in autism and in mental retardation without autism the majority of the cases are males, suggesting a X-chromosomal effect. In fact, some molecular evidence has been obtained for a common genetic background for autism spectrum disorders (ASD) and X-linked mental retardation (XLMR). In several genome-wide scans (GWS), evidence for linkage at X-chromosome has been reported including the GWS of Finnish ASD families with the highest multipoint lod score (MLS) of 2.75 obtained close to DXS7132 at Xq11.1. To further dissect the relationship between autism and genes implicated in XLMR, we have fine-mapped Xq11.1-q21.33 and analyzed five candidate genes in the region. We refined the region using 26 microsatellite markers and linkage analysis in 99 Finnish families with ASD. The most significant evidence for linkage was observed at DXS1225 on Xq21.1 with a nonparametric multipoint NPL(all) value of 3.43 (P = 0.0004). We sequenced the coding regions and splice sites of RPS6KA6 and ZNF711 residing at the peak region in 42 male patients from families contributing to the linkage. We also analyzed ACSL4 and DLG3, which have previously been known to cause XLMR and IL1RAPL2, a homologous gene for IL1RAPL1 that is mutated in autism and XLMR. A total of six novel and 11 known single nucleotide polymorphisms were identified. Further studies are warranted to analyze the candidate genes at Xq11.1-q21.33.

Analysis of 9p24 and 11p12-13 regions in autism spectrum disorders: rs1340513 in the JMJD2C gene is associated with ASDs in Finnish sample.

OBJECTIVE: Autism spectrum disorders (ASD) often show obsessive repetitive symptoms that are characteristic to obsessive-compulsive disorder (OCD). Aberrant glutamate function has been suggested to a risk for both ASDs and OCD. Considering the common metabolic pathway and recent results from association studies both in OCD and ASDs, a question, whether there is common molecular background in ASDs and OCD, was raised. METHODS: Ten single nucleotide polymorphisms (SNPs) at 9p24 and 11p12-p13 containing glutamate transporter genes SLC1A1 and SLC1A2 and their neighboring regions in 175 patients with ASDs and 216 controls of Finnish origin were analyzed using real-time-PCR or direct sequencing. RESULTS: The strongest association was detected with rs1340513 in the JMJD2C gene at 9p24.1 (P=0.007; corrected P=0.011) that is the same SNP associated with infantile autism (P=0.0007) in the autism genome project consortium (2007). No association was detected at 11p12-p13 with ASD. Interestingly, the strongest association in OCD has been found at rs301443 (P=0.000067) residing between SLC1A1 and JMJD2C at 9p24. CONCLUSION: In summary, our results give evidence for a possible common locus for OCD and ASDs at 9p24. We speculate that the area may represent a special candidate region for obsessive repetitive symptoms in ASDs.

Allelic variants in HTR3C show association with autism.

Autism spectrum disorders (ASDs) are severe neurodevelopmental disorders with a strong genetic component. Only a few predisposing genes have been identified so far. We have previously performed a genome-wide linkage screen for ASDs in Finnish families where the most significant linkage peak was identified at 3q25-27. Here, 11 positional and functionally relevant candidate genes at 3q25-27 were tested for association with autistic disorder. Genotypes of 125 single nucleotide polymorphisms (SNPs) were determined in 97 families with at least one individual affected with autistic disorder. The most significant association was observed using two non-synonymous SNPs in HTR3C, rs6766410 and rs6807362, both resulting in P = 0.0012 in family-based association analysis. In addition, the haplotype C-C corresponding to amino acids N163-A405 was overtransmitted to affected individuals (P = 0.006). Sequencing revealed no other variants in the coding region or splice sites of HTR3C. Based on the association analysis results in a previously identified linkage region, we propose that HTR3C represents a novel candidate locus for ASDs and should be tested in other populations.

Sleep in children with Asperger syndrome.

The prevalence of sleep disturbances in 52 children with Asperger syndrome (AS) as compared with 61 healthy controls (all subjects aged 5-17 years) was investigated. Problems with sleep onset and maintenance, sleep-related fears, negative attitudes toward sleeping, and daytime somnolence were more frequent among children with AS than among controls. Short sleep duration (<9 h) was almost twofold (59% vs. 32%), and the risk for sleep onset problems more than fivefold (53% vs. 10%) more common in the AS group than in the control group. Child-reported sleeping problems were also more prevalent in the AS group than in controls (58% vs. 7%). The results suggest that sleep disturbances should be routinely evaluated in children with AS.

No association between common variants in glyoxalase 1 and autism spectrum disorders.

The autism spectrum disorders (ASDs) are complex diseases with a strong genetic component. Numerous candidate gene studies have tested association between various functional and positional candidate genes and autism, but no common variation predisposing for autism has been identified to date. It has been previously proposed, that glyoxalase 1 (GLO1) might be involved in the pathogenesis of autism as GLO1 protein polarity was significantly changed in the brains of autism patients compared to controls. GLO1 harbors a functional polymorphism that affects the polarity and the enzymatic activity of the protein. In the same study, this polymorphism showed a suggestive association to autism. To investigate whether common variants in GLO1 predispose to autism in the Finnish population, we have genotyped six polymorphisms in GLO1 in families with more than 230 individuals affected with ASDs and carried out both linkage and association analyses. We did not observe significant linkage or association between any SNP and ASDs. Therefore, we suggest that common variants in GLO1 are not significant susceptibility factors for ASDs in the Finnish population.

Cerebrospinal fluid insulin-like growth factors IGF-1 and IGF-2 in infantile autism.

There has been little exploration of major biologic regulators of cerebral development in autism. We measured insulin-like growth factors (IGF) -1 and -2 from cerebrospinal fluid (CSF) by radio immunoassay in 25 children with autism (median age 5y 5mo; range 1y 11mo-15y 10mo; 20 males, 5 females), and in 16 age-matched comparison children without disability (median age 7y 4mo; range 1y 1mo-15y 2mo; eight males, eight females). IGF-1 and -2 concentrations were further correlated with age of patients and head size. CSF IGF-1 concentration was significantly lower in patients with autism than in the comparison group. The CSF concentrations of children with autism under 5 years of age were significantly lower than their age-matched comparisons. The head circumferences correlated with CSF IGF-1 in children with autism but no such correlation was found in the comparison group. There was no difference between the two groups in CSF IGF-2 concentrations. No patients with autism had macrocephaly. We conclude that low concentrations of CSF IGF-1 at an early age might be linked with the pathogenesis in autism because IGF-1 is important for the survival of Purkinje cells of the cerebellum. The head growth might be explained by the actions of IGF-1 and -2 reflected in CSF concentrations.

Search for autism loci by combined analysis of Autism Genetic Resource Exchange and Finnish families.

OBJECTIVE: Several genome-wide screens have been performed in autism spectrum disorders resulting in the identification of numerous putative susceptibility loci. Analyses of pooled primary data should result in an increased sample size and the different study samples have a potential to strengthen the evidence for some earlier identified loci, reveal novel loci, and even to provide information of the general significance of the locus. The objective of this study was to search for potential susceptibility loci for autism, which are supported by two independent samples. METHODS: We performed a combined analysis of the primary genome scan data of the Autism Genetic Resource Exchange (AGRE) and Finnish autism samples to reveal susceptibility loci potentially shared by these study samples. RESULTS: In the initial combined data analysis, the best loci (p < 0.05) were observed at 1p12-q25, 3p24-26, 4q21-31, 5p15-q12, 6q14-21, 7q33-36, 8q22-24, 17p12-q21, and 19p13-q13. The combined analysis of Finnish and AGRE families showed the most promising shared locus on 3p24-26 with nonparametric logarithm of odds (NPL) score of 2.20 (p = 0.011). The combined data analysis did not provide increased linkage evidence for the earlier identified loci on 3q25-27 or 17p12-q21. However, the 17p12-q21 locus remained promising also in the combined sample (NPL(all) =2.38, p = 0.0076). INTERPRETATION: Our study of 314 autism families highlights the importance of further analyses on 3p24-26 locus involving comprehensive molecular genetic analyses of oxytocin receptor gene (OXTR), a positional and functional candidate gene for autism.

The discrimination of and orienting to speech and non-speech sounds in children with autism.

The present study aimed to find out how different stages of cortical auditory processing (sound encoding, discrimination, and orienting) are affected in children with autism. To this end, auditory event-related potentials (ERP) were studied in 15 children with autism and their controls. Their responses were recorded for pitch, duration, and vowel changes in speech stimuli, and for corresponding changes in the non-speech counterparts of the stimuli, while the children watched silent videos and ignored the stimuli. The responses to sound repetition were diminished in amplitude in the children with autism, reflecting impaired sound encoding. The mismatch negativity (MMN), an ERP indexing sound discrimination, was enhanced in the children with autism as far as pitch changes were concerned. This is consistent with earlier studies reporting auditory hypersensitivity and good pitch-processing abilities, as well as with theories proposing enhanced perception of local stimulus features in individuals with autism. The discrimination of duration changes was impaired in these children, however. Finally, involuntary orienting to sound changes, as reflected by the P3a ERP, was more impaired for speech than non-speech sounds in the children with autism, suggesting deficits particularly in social orienting. This has been proposed to be one of the earliest symptoms to emerge, with pervasive effects on later development.

Analysis of four neuroligin genes as candidates for autism.

Neuroligins are cell-adhesion molecules located at the postsynaptic side of the synapse. Neuroligins interact with beta-neurexins and this interaction is involved in the formation of functional synapses. Mutations in two X-linked neuroligin genes, NLGN3 and NLGN4, have recently been implicated in pathogenesis of autism. The neuroligin gene family consists of five members (NLGN1 at 3q26, NLGN2 at 17p13, NLGN3 at Xq13, NLGN4 at Xp22, and NLGN4Y at Yq11), of which NLGN1 and NLGN3 are located within the best loci observed in our previous genome-wide scan for autism in the Finnish sample. Here, we report a detailed molecular genetic analysis of NLGN1, NLGN3, NLGN4, and NLNG4Y in the Finnish autism sample. Mutation analysis of 30 probands selected from families producing linkage evidence for Xq13 and/or 3q26 loci revealed several polymorphisms, but none of these seemed to be functional. Family-based association analysis in 100 families with autism spectrum disorders yielded only modest associations at NLGN1 (rs1488545, P=0.002), NLGN3 (DXS7132, P=0.014), and NLGN4 (DXS996, P=0.031). We conclude that neuroligin mutations most probably represent rare causes of autism and that it is unlikely that the allelic variants in these genes would be major risk factors for autism.

MECP2 mutation analysis in patients with mental retardation.

Mutations in the methyl-CpG-binding protein 2 (MECP2) gene are known to underlie Rett' syndrome, the most common cause of mental retardation (MR) in girls. Since the original report, phenotypes resulting from MECP2 mutations have been shown to extend, for example, to several Rett variants, autism, atypical Angelman syndrome, and nonspecific MR. It was earlier proposed that MECP2 mutations might account for approximately 2% of the male cases with nonspecific MR. Thereby, the frequency of MECP2 mutations in the mentally retarded population would be comparable to that of Fragile-X syndrome. The aim of this study was to analyze well-characterized cases with MR and to clarify the role of the MECP2 gene in the etiology of MR and atypical Angelman syndrome. The coding sequence of the MECP2 gene was analyzed in a sample of 118 patients (103 males, 15 females) by direct sequencing. Two coding sequence variants, 602C > T (A201V) and 1189G > A (E397K), were identified. In addition, we identified four variants in the intronic or 3'UTR regions. None of these variants is likely to be causal. We conclude that the evidence across all the mutation screening studies implies that MECP2 mutations do not represent a major cause of nonspecific MR.

Family-based association study of DYX1C1 variants in autism.

DYX1C1: was recently identified as a candidate gene for developmental dyslexia, which is characterized by an unexpected difficulty in learning to read and write despite adequate intelligence, motivation, and education. It will be important to clarify, whether the phenotype caused by DYX1C1 extends to other language-related or comorbid disorders. Impaired language development is one of the essential features in autism. Therefore, we analyzed the allelic distribution of the DYX1C1 gene by family-based association method in 100 Finnish autism families. No evidence for association was observed with any intragenic marker or with haplotypes constructed from alleles of several adjacent markers. No evidence for deviated allelic diversity was either observed: the frequency of expected dyslexia risk haplotype was comparable to its frequency in Finnish controls. Thus it seems unlikely that DYX1C1 gene would be involved in the genetic etiology of autism in Finnish patients.

Increased presynaptic dopamine function in Asperger syndrome.

The etiology of Asperger syndrome is essentially unknown, but abnormality of the dopamine system has been shown in clinically overlapping disorders. The present study was designed to investigate the presynaptic dopamine function in Asperger syndrome. Eight healthy, drug-free males with Asperger syndrome and five healthy male controls were examined with positron emission tomography using 6-[18F]fluoro-L-DOPA ([18F]FDOPA) as a tracer. In the Asperger syndrome group, the [18F]FDOPA influx (Ki) values were increased in the striatum, i.e. in the putamen and caudate nucleus and in the frontal cortex. The results indicate that the dopamine system is affected in subjects with Asperger syndrome. Partially similar results have also been obtained in schizophrenia, suggesting an overlap not only of the clinical features but also of pathogenesis.

Identification of two AGTR2 mutations in male patients with non-syndromic mental retardation.

Mutations in the coding region of the angiotensin II type 2 receptor gene (AGTR2) were recently identified to cause X-linked recessive mental retardation. We report a mutation screening of the AGTR2 gene in 57 Finnish male patients with non-syndromic mental retardation. We identified two mutations, a 62G-->T transversion, which leads to a substitution of glycine for valine (G21V) and a 157A-->T transversion, which causes a substitution of isoleucine for phenylalanine (I53F). The patients with AGTR2 sequence variants had severe/profound mental retardation, epileptic seizures, restlessness, hyperactivity, and disturbed development of speech.