Phenotypic categorization of putative pathogenic CNVs in a population of Autism Spectrum Disorder patients Autism Spectrum Disorders (ASD) have a strong genetic component, with an estimated heritability of over 90% 1 . Recent studies carried out by the Autism Genome Project (AGP) consortium and others suggest that rare Copy Number Variants (CNVs), characterized by submicroscopic chromosomal deletions and duplications, are more frequent in ASD cases compared to controls, and may play an important role in susceptibility to this disorder 2 . However, to adequately assess pathogenicity, a detailed characterization of each patients’ CNVs and clinical phenotype is required. The goal of this study was to establish the clinical and etiological relevance for ASD of potentially pathogenic CNVs identified in a Portuguese population sample by whole genome CNV analysis, through the detailed characterization of CNVs and correlation with clinical phenotypes. A genome-wide CNV screening of 342 ASD patients, carried out by the AGP using 1M SNP microarrays 2 , identified a total of 14218 CNVs in this sample. We selected for further characterization 149 genic CNVs (CNVs containing genes or parts of genes) present in 127 individuals, using the following criteria: 1) CNVs that contained implicated/candidate genes for ASD; 2) CNVs in genomic regions known to be implicated/candidate for ASD; 3) CNVs in regions associated with syndromes presenting ASD symptoms; and 4) high confidence CNVs that did not overlap more than 20% with controls in available databases. We explored recurrence rates, genic content, regulatory elements, inheritance patterns and phenotypic correlations. Inês C. Conceição 1,2,3 , Catarina Correia 1,2,3 , Bárbara Oliveira 1,2,3 , Maria M. Rama 1 , Cátia Café 4 , Joana Almeida 4 , Susana Mouga 4,5 , Frederico Duque 4,6 , Guiomar Oliveira 4,5,6,7 , Astrid M. Vicente 1,2,3 1 Instituto Nacional de Saúde Dr. Ricardo Jorge, 1600 Lisboa, Portugal; 2 Center for Biodiversity, Functional & Integrative Genomics, Lisboa, Portugal; 3 Instituto Gulbenkian de Ciência, Oeiras, Portugal; 4 Unidade Neurodesenvolvimento e Autismo, Centro de Desenvolvimento, Hospital Pediátrico (HP); Centro Hospitalar e Universitário de Coimbra (CHUC), Coimbra, Portugal; 5 Instituto Biomédico de Investigação em Luz e Imagem, Faculdade de Medicina da Universidade de Coimbra, Portugal; 6 Faculdade de Medicina da Universidade de Coimbra, Portugal; 7 Centro de Investigação e Formação Clinica do HP-CHUC, Portugal CNV characterization and distribution Phenotypic correlations REFERENCES: 1. Betancur C. (2010) Brain Res 1380:42-77; 2. Pinto D. et al. (2010) Nature 466:368-72; 3. Franceschini A. et al. (2013) Nucleic Acids Res. 41:D808-15. ACKNOWLEDGMENTS: This work was supported by the fellowships SFRH/BPD/74739/2010 to ICC, SFRH/BPD/64281/2009 to CC and SFRH/BD/79081/2011 to BO from Fundação para a Ciência e a Tecnologia (Portugal). All individuals in this study signed an informed consent. The 149 genic CNVs selected were identified in 127 individuals (12 females and 115 males), ranged from 5 Kb to 3 Mb, 58% were deletions, and included from one gene (67% of all genic CNVs) to 25 genes in a single CNV. Large CNVs (>500 Kb) were more frequently duplications. Although most CNVs (82%) were present in a single individual, 26 CNVs (17.2%), distributed across 14 genomic regions, were present in more than one individual, and some encompassed regions/genes implicated in autism 1,2 , such as 16p13.11 (N=3), PARK2 (N=5), and VPS13B (N=3), as well as putative novel genes for ASD (eg UBC, SLC25A4, TERT). Network analysis of the 309 genes mapping to genic CNVs, using the String software 3 , yielded a network including 67 genes (Fig.1). This network is enriched in the following Biological Processes: regulation of apoptotic processes (14 genes; p-value=0.05), and PPAR and ErbB signaling pathways (5 genes; p-value=0.02 in both cases). This data adds novel genes for autism to the list of potential candidates, and identifies biological pathways not previously associated with ASD. Figure 1. Protein network using the genes present in CNVs of the Portuguese dataset This patient sample consisted of individuals with no severe intellectual disability (IQ>35) and no gross chromosomal aberrations or obvious dysmorphisms, but included some subjects with minor dysmorphisms or macrocephaly. We explored differences in the type, size and number of the genic CNVs and in the number of genes implicated, in relation to different phenotypic categories (Fig. 2): 1) intellectual disability (ID): no (N=79) and yes (IQ=35-69; N=59); 2) minor dysmorphisms: no (N=122) and yes (N=17); and 3) family history of neuropsychiatric disorders: no (N=61) and yes (N=78). As observed in previous studies, there were more deletions than duplications, regardless of the phenotype. In general, individuals with ID, dysmorphisms and/or family history, had a higher burden of genic CNVs, higher number of genes affected (both per CNV and per individual) and a larger average CNV size. At least 7 individuals presented CNVs disrupting genes frequently identified in ASD patients, which constitute an etiological diagnosis. Most CNVs contained only one gene, and were present in a single individual, regardless of phenotypic category, thus reinforcing the role of rare variants in this disease and the large heterogeneity in ASD etiology. CNV inheritance and parental personality traits We further evaluated correlations between data for autistic traits in the parents and CNV inheritance, using the Broad Autism Phenotype Questionnaire (BAPQ) and the Social Responsiveness Scale (SRS) (Fig. 3). A significant excess of autistic traits was observed in the fathers that transmitted a CNV, mainly in the “aloof” personality, which is defined as lacking interest in social interaction. Paternal inheritance does not explain all changes, indicating a putative maternal contribution. Using SRS questionnaire results from parents and probands, we also calculated familial correlations for de novo or inherited CNVs and all parent-offspring and parental pair types (data not shown). While there were no SRS correlations between any parent-offspring types for de novo or inherited CNVs, a significant correlation between the SRS results from both parents supports the idea of assortative mating in ASD. Figure 3. Average parental BAPQ and SRS results vs type of inheritance of the CNVs, using Mann-Whitney U test. Comparisons were made between: 1) UT vs T; 2) UT vs Tmat; and 3) UT vs Tpat. UT, untransmitting parents; T, transmitting parents; Tmat, transmitting mothers; Tpat, transmitting fathers. Aloof personality, lack of social interest; pragmatic (prs) personality, communication deficits; rigid personality, lack of ability to adjust. Two-hundred and fifty parents were used for these tests. 0 5 10 15 20 25 30 35 40 0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 BAPQ total BAPQ aloof BAPQ rigid BAPQ prs SRS SRS results BAPQ results UT T Tmat Tpat * Figure 2. Phenotypic correlation with the different types of genic CNVs. 0 50 100 150 200 250 0 10 20 30 40 50 60 70 deletion duplication Average size of CNVs (Kb) Type of CNV (%) 0 0.5 1 1.5 2 2.5 3 3.5 4 1.15 1.20 1.25 1.30 1.35 1.40 1.45 1.50 1.55 per individual per CNV per individual # genes # genic CNVs ID no ID yes dysmorphisms no dysmorphisms yes family history no family history yes Copy Number Variants in Autism Spectrum Disorder brought to you by CORE View metadata, citation and similar papers at core.ac.uk provided by Repositório Científico do Instituto Nacional de Saúde