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Contribution of Rare Copy Number Variants to the Development of Autism Spectrum Disorder in High-Risk Siblings

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Contribution of Rare Copy Number Variants to the Development of Autism Spectrum Disorder in High-Risk Siblings Contribution of Rare Copy Number Variants to the Development of Autism Spectrum Disorder in High-Risk Siblings by Lia D’Abate A thesis submitted in conformity with the requirements for the degree of Master of Science Department of Molecular Genetics University of Toronto © Copyright by Lia D’Abate 2016 Contribution of Rare Copy Number Variants to the Development of Autism Spectrum Disorder in High-Risk Siblings Lia D’Abate Master of Science Department of Molecular Genetics University of Toronto 2016 Abstract As microarrays are commonly used to identify the causes of autism spectrum disorder (ASD) we are investigating whether genetic testing in young children can predict ASD status prior to the typical age of onset of symptoms. A total of 441 individuals (190 probands, 88 siblings with ASD, 163 unaffected siblings) from the Infant Siblings Research Consortium were genotyped on the high-resolution Affymetrix CytoScan HD platform. De novo and inherited presumed pathogenic CNVs were identified in 23/199 families (11.4%). In 6 of these families, the index case and one or more siblings carried the presumed pathogenic CNV; in every such case the carrier individual either had ASD (4) or had a subclinical form of ASD (2). There were 6 additional families in which CNV data would have been informative for early ASD diagnosis or detection of developmental delay. Our results suggest CNVs may serve as biological markers for ASD. ii Acknowledgements The completion of this thesis could not have been possible without the unwavering support of my supervisor Dr. Stephen Scherer. One of the greatest privileges of working under the supervision of Dr. Scherer was to not only see his commitment to scientific excellence, but his genuine desire to share that with the public and improve the quality of life of children with neurodevelopmental disorders. I now strive to bring that level of humanity and reverence for basic science to all of my projects. I am very grateful to have had access to first-class technologies that make answering these scientific questions possible. I would also like to thank my committee members Dr. Lucy Osborne and Dr. Freda Miller for always giving me excellent guidance and supporting me on my path to becoming a better graduate student. My ability to make use of multitude of resources was not only contingent on the mentorship and support of my supervisor, but that of all members of The Centre for Applied Genomics (TCAG). In particular, I would like to thank my lab mates Dr. Ryan Yuen, Dr. Susan Walker, Dr. Mohammed Uddin, Dr.Mehdi Zarrei, Dr. Kristiina Tammimies and Dr. Eric Denault for their willingness to share their expertise and sound advice as I worked towards completing this project. Additionally, I would especially like to thank Dr. Richard Wintle, Dr. Daniele Merico, Dr. Liz Li, Dr. Chao Lu, Dr. Giovanna Pellecchia, Mrs. Bhooma Thiruvahindrapuram and Mr. John Wei for guiding me so carefully along the very technical aspects of my analysis. Lastly, my time in the lab would not have been complete without my fellow graduate students Ada Chan, Matthew Gazzellone and Dr. Anath Lionel, whom are more like family than lab mates. A student’s pursuit of graduate studies not only requires a fertile academic environment, but a tremendously supportive home one as well. I would like to deeply thank my parents for their unwavering love and support and all of the wonderful friends I have made both in and out of school. You anchored me during my most difficult days, and I could only hope that I have given you as much friendship and compassion as you have given me. iii Table of Contents Acknowledgements ........................................................................................................................ iii Table of contents ........................................................................................................................... iv List of Tables ................................................................................................................................ vi List of Figures .............................................................................................................................. vii List of Appendices ...................................................................................................................... viii Introduction ......................................................................................................................................1 1.1 Clinical Presentation of ASD ................................................................................................1 1.2 Genetic Etiology of ASD ......................................................................................................3 1.2.1. Evidence from Family-Based Studies ...........................................................................4 1.2.2 The Broader Autism Phenotype ....................................................................................5 1.3 Genetic Models of ASD .........................................................................................................6 1.3.1. Common Disease-Common Variant Hypothesis ...........................................................6 1.3.2 Common Disease-Rare Variant Hypothesis ..................................................................8 1.4 Copy Number Variation in ASD ............................................................................................9 1.5 Developmental Neurobiology of ASD .................................................................................11 1.6 Project Rationale .................................................................................................................12 1.6.1. Hypothesis ....................................................................................................................12 1.6.2 Objectives ....................................................................................................................13 Methods..........................................................................................................................................15 2.1 Sample Ascertainment and Diagnostic Criteria ..................................................................15 2.1.1 Autism Diagnostic Observation Schedule, Second Edition ..........................................17 2.1.2 Mullen Scales of Early Learning ...................................................................................17 iv 2.1.3 Vineland Adaptive Behavioral Scales, Second Edition ...............................................17 2.2 DNA Collection and Genotyping .........................................................................................18 2.3 CNV Calling ........................................................................................................................18 2.4 Variant Prioritization and Molecular Validation .................................................................23 2.5 Critical Exon Analysis ........................................................................................................24 Results ............................................................................................................................................26 3.1 De Novo CNV Analysis .....................................................................................................26 3.1.1 De Novo CNVs in Individuals with ASD .....................................................................26 3.1.2 De Novo CNVs in Individuals without ASD ...............................................................28 3.1.3 Critical Exon Analysis of De Novo CNVs ...................................................................29 3.1.4 Distribution of De Novo and Inherited Presumed Pathogenic CNVs ..........................29 3.2 Predictive Value of Chromosomal Microarray ....................................................................35 Discussion ......................................................................................................................................38 4.1 Current Study Limitations ....................................................................................................41 Conclusion and Significance..........................................................................................................43 Future Directions ...........................................................................................................................44 6.1 WGS of Infant Siblings Cohort .....................................................................................45 6.2 Investigating the Role of PTCHD1-AS in ASD ............................................................46 Appendices ....................................................................................................................................49 7.1 Rare, Exonic CNVs .............................................................................................................50 References ......................................................................................................................................63 v List of Tables Table 1: de novo CNVs in Probands (n=189) and Infant Siblings (n=119) .................................27 Table 2: Identification of Exonic Presumed Pathogenic CNVs in Infant Siblings of ASD Probands ........................................................................................................................................33 Table 3: Predictive Value
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