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Thesis Submitted in Conformity with the Requirements for the Degree of Doctor of Philosophy Institute of Medical Science University of Toronto

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Thesis Submitted in Conformity with the Requirements for the Degree of Doctor of Philosophy Institute of Medical Science University of Toronto Expression of Parkinson’s disease and associated neurophenotypes in 22q11.2 deletion syndrome by Nancy Jean Butcher A thesis submitted in conformity with the requirements for the degree of Doctor of Philosophy Institute of Medical Science University of Toronto © Copyright by Nancy Butcher 2015 Expression of Parkinson’s disease and associated neurophenotypes in 22q11.2 deletion syndrome Nancy Jean Butcher Doctor of Philosophy Institute of Medical Science University of Toronto 2015 Abstract The etiology of Parkinson’s disease (PD) remains largely unknown with the exception of a few genetic mutations that affect a small proportion of patients. Case reports suggest that individuals with 22q11.2 deletion syndrome (22q11.2DS), a multisystem genomic disorder associated with hemizygous 22q11.2 deletions, may be at increased risk of early-onset PD. The aim of this thesis was to investigate 22q11.2 deletions as a risk factor for early-onset PD. The prevalence of PD was assessed in a well-characterized cohort of adults with 22q11.2DS. Neuropathological studies were performed in cases with available post-mortem tissue. Whole- genome sequencing was used to investigate the possible contribution of genome-wide rare coding mutations to disease penetrance in neuropathologically confirmed PD cases. Assessment of putative pre-diagnostic clinical and neuroimaging markers of PD in a subset of the older adults (30 to 54 years) provided evidence of an elevated prevalence of pre-morbid motor and olfactory deficits, and of nigrostriatal dopaminergic dysfunction assessed using 11C-dihydrotetrabenazine and positron emission tomography. The clinical context of early-onset PD in this adult cohort of individuals with 22q11.2DS was also investigated. Treatment response was excellent to the atypical antipsychotic, clozapine, a ii candidate for use in psychotic patients at risk of, or with, PD. Cognitive level and severe psychiatric disorders were mediators of baseline functional capacity in 22q11.2DS adults. Practical guidelines were developed from a review of the 22q11.2DS literature to help inform the management of 22q11.2DS-associated conditions, including PD. The results of these studies provided evidence that early-onset PD is associated with the 22q11.2 deletion with important implications for PD pathogenesis and for the clinical management of 22q11.2DS. iii Acknowledgments I would first and foremost like to thank my supervisor, mentor, and friend, Dr. Anne Bassett, for her unwavering encouragement, support, and intellectual guidance over the course of my doctoral studies. It has been my honour and privilege to train with her. I thank my program advisory committee members, Drs. Anthony Lang and Lucy Osborne, for their scientific input and support at each stage of my graduate studies. I also thank the other members of my thesis examination committee, including Drs. Mario Masellis, Gary Remington, and William Dauer. I have been fortunate to work with an amazing team at the Clinical Genetics Research Program (Centre for Addiction and Mental Health) and with wonderful collaborators at the University of Toronto. I would particularly like to acknowledge Drs. Erik Boot, Eva Chow, Gregory Costain, W. L. Alan Fung, Connie Marras, Daniele Merico, Antonio Strafella, and the CAMH Research Imaging Centre staff for their input, support, and contributions. Research funding supporting my doctoral studies was provided from the Canadian Institutes of Health Research (CIHR) and Brain Canada in the form of both personal salary and travel awards. Operating grant support was provided through CIHR grants awarded to Dr. Bassett and her colleagues, and the Canada Research Chairs (Tier 1) program to Dr. Bassett. Thank you to my friends and family, both near and far, for their support throughout this journey. I would like to thank Joey for his encouragement, patience, and love. Special thanks to my brother, Dr. Clifford Butcher, and to my “science sister,” graduate student Chelsea Lowther. I would like to dedicate this thesis to my mother, Betty Ann Devoe, whose constant love and support has brought me to where I am today. iv Contributions Chapter 1: Introduction & Chapter 2: Research aims and hypotheses Contribution statement and acknowledgements: The content of these chapters was conceived, drafted, and revised by me, with the following qualifiers. Anne Bassett provided helpful scientific comments and revisions. Table 1-1 presents the United Kingdom Parkinson’s Disease Society Brain Bank Clinical diagnostic criteria.1 Table 1-3 describes the Diagnostic and Statistical Manual of Mental Disorders diagnostic criteria for schizophrenia.2 Figure 1-1 was reproduced with permission from Fusar-Poli et al. (2012).3 Figure 1-2 was reproduced with permission from Berg et al. (2011).4 Figures 1-35 and Figures 1-46 were prepared under our direction by Sean Bekeschus. Chapter 3: Association between early-onset Parkinson’s disease and hemizygous 22q11.2 deletions Citation: Butcher NJ*, Kiehl T-R*, Hazrati L-N, Chow EWC, Rogaeva E, Lang AE, Bassett AS. 2013. Individuals with 22q11.2 deletion syndrome are at increased risk of early-onset Parkinson disease: Identification of a novel genetic form of Parkinson disease and its clinical implications. JAMA Neurology. 70:1359-1366. *Co-first authors Contribution statement and acknowledgements: My roles in this study included the following: 1) conceiving and designing the study, 2) Identifying and characterizing cases of PD in the existing 22q11.2DS cohort, 3) Checking and correcting demographic data in existing 22q11.2DS database, 4) Statistical analyses, 5) Coordinating and participating in one of the three autopsies of a 22q11.2DS patient with PD, 6) Providing input on immunohistochemistry methods, interpretation, and presentation of results, 7) Coordinating and interpreting genetic test results, 8) Writing the manuscript, 9) Submitting and revising the manuscript for publication, and 10) Communicating results through conference presentations and press interviews. I thus played a primary role in every aspect of this study with the exception of performing wet lab work (immunohistochemistry, genetic testing, and autopsies completed prior to the beginning of my doctoral studies). v The roles of the co-first author, Rasmus Kiehl, were as follows: 1) Initiating neuropathological studies of adults with 22q11.2DS, 2) Coordinating and/or performing autopsies, 3) Immunohistochemistry experiments and interpretation, and 4) Critically revising the manuscript. Anne Bassett supervised all aspects of my involvement in this study, and made critical contributions to the study design, data interpretation, and writing the manuscript. Adults with 22q11.2DS were recruited and assessed over a period of many years as part of a longitudinal study of 22q11.2DS, directed by Anne Bassett and Eva Chow, which made this study possible. Lili-Naz Hazrati conducted one of the three autopsies, and performed immunohistochemistry experiments including interpretation and quantification (Table 3-3). Eva Chow contributed to statistical analyses. Anthony Lang provided critical input on data interpretation and contributed the early-onset PD cohort for testing for the 22q11.2 deletion, together with Ekaterina Rogaeva, who directed the genetic testing of known Parkinson’s disease genes in our four 22q11.2DS-PD cases. All authors provided critical revision of the manuscript for important intellectual content. We thank the generosity of the patients’ families that made this study possible, the pathologists who performed the general autopsies, and colleagues for referring patients. This work was supported by The Canadian Institutes of Health Research (N.J.B, A.S.B), Canada Research Chairs Program (A.S.B), the Ontario Research Fund (E.R.), and the National Parkinson Foundation (A.E.L.). Chapter 4: Genetic markers of Parkinson’s disease in 22q11.2 deletion syndrome Contribution statement and acknowledgements: My roles in the study were as follows: 1) Conceiving and designing the study, 2) Providing input on the whole-genome sequencing bioinformatics pipeline, 3) Developing, selecting, and testing PD-relevant gene-sets, 4) Statistical analyses, 5) Checking and correcting demographic and clinical data in existing 22q11.2DS database, 6) Interpreting genetic variant results, 7) Writing the manuscript and preparing the manuscript for publication, and 8) Communicating results through conference presentations. I thus played a primary role in every aspect of this study, with the exception of genome sequencing and core bioinformatic processing, performed by Complete Genomics and The Centre for Applied Genomics. vi Anne Bassett supervised all aspects of my involvement in this study, and made critical contributions to the study conception and design, data interpretation, and revising the manuscript. Daniele Merico and Mehdi Zarrei performed bioinformatic processing of the whole-genome sequencing data. Lucas Ogura assisted with data analyses and quality checking. Thomas Nalpathamkalam and Bhooma Thiruvahindrapuram performed variant annotation. Christian Marshall and Stephen Scherer contributed to study conception and design. Eva Chow, Daniele Andrade, and Anthony Lang contributed clinical data. All authors provided critical revision of the manuscript for important intellectual content. This work was supported by the Canadian Institutes of Health Research (A.S.B, MOP #97800, MOP #111238), Canada Research Chairs program (A.S.B), a Brain Canada Mental Health Training Award (N.J.B), a Canadian Institutes
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