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Posters MG-101 to MG-119

Mutations preventing regulated exon skipping of a receptor tyrosine kinase cause a developmental disorder of osteogenesis
Peter Kannu, Hospital for Sick Children, Toronto ON Canada

          Background. Osteofibrous dysplasia (OFD) is a proliferative fibro-osseous condition affecting bone with an onset in early childhood. Affected individuals develop tibial bowing and spontaneous non-healing fractures.
          Objectives. The purpose of this study was to identify the genetic cause of OFD and clarify how the mutation causes the phenotype.
          Design/Methods. Exome sequencing was performed on individuals diagnosed with bilateral OFD. Cell culture and mouse studies were utilized to study disease mechanism.
          Results. We identified mutations in a gene encoding a receptor tyrosine kinase, in three families segregating an autosomal dominant form of OFD and in a fourth sporadic case. All mutations abolished the splice inclusion of an exon in transcripts resulting in receptors lacking a juxtamembrane cytoplasmic domain. Our mouse data indicate that exclusion of this domain is a physiologically regulated event in the periosteum, which is spatially partitioned during development.  Abnormal fibro-osseous bone cells from the patient tibial fracture site were compared to the cells obtained from unaffected proximal tibial bone. When cultured in osteogenic differentiation media, fracture site-derived osteoblasts displayed considerably higher levels of mineralization compared to the cells from the unaffected tibial bone. Moreover, the fracture site osteoblasts proliferated more rapidly compared to the control cells. Mutant osteoclasts showed decreased bone resorption.
          Conclusions. Together these data indicate that the production of this mutant receptor isoform is a developmentally regulated event during mammalian embryonic development and mutations which render this alternative splice event constitutional subvert core functions of this receptor that regulate osteogenic functions within the periosteum.

Coffin-Siris syndrome caused by a missense mutation in ARID1A
Andrea Guerin MDa, Sohnee Ahmed CGCb, Matthew Deardorff MD PhDc, Melissa Carter MDb
aDivision of Medical Genetics, Kingston General Hospital, Kingston ON, Canada
bDivision of Clinical and Metabolic Genetics, The Hospital for Sick Children, Toronto ON, Canada
cDivision of Human Genetics, Children’s Hospital of Philadelphia, Philadelphia PA, USA

          Coffin-Siris syndrome (CSS; OMIM #135900) is caused by mutations in genes encoding components of the BRMassociated factor (BAF) complex, also known as the mammalian SWItch/sucrose nonfermentable (mSWI/SNF)-like complex. Clinical features are variable and may include hypoplasia of the distal phalanx of the fifth digit, developmental delay, dysmorphic or coarse facial features, growth restriction, and malformations of the brain, heart, and other organs. No widely accepted diagnostic criteria yet exist. Mutations in the ARID1A gene account for approximately 13% of CSS cases with eight patients reported to date (Am J Med Genet C Semin Med Genet. 2014;166C(3):262-75).
Here we describe a 20-month-old boy with clinical features consistent with CSS, including feeding difficulties, hypotonia, speech delay, CNS anomalies, hypoplastic 5th digits, thick lower lip vermillion, and hypertrichosis.  Our patient was found to have a de novo missense mutation (p.Leu1713Pro) in a highly conserved amino acid residue located in one of the leucine-rich motifs of ARID1A.  All reported mutations of ARID1A associated with CSS thus far have been truncating, and are believed to be mosaic in lymphocytes. ARID1A heterozygous knock out mice are embryonic lethal, suggesting a vital role in embryonic development. Notably, while mutations in ARID1A noted in cancer are typically nonsense or frameshift, missense mutations in this same leucine-rich motif have been reported.  These findings suggest several possibilities including: 1. This mutation resides in a critical position key to the function of ARID1A in human development; 2. This effect may be mild or specific enough to be tolerated and not cause lethality; or 3. This mutation causes moderate instability of the protein leading to a milder clinical presentation. Studies are underway to determine the functional consequences of this missense mutation on ARID1A mRNA and protein production.

Determining Genetics referral eligibility for hereditary breast/ovarian cancer risk assessment: an electronic solution
Wendy Meschino, Joanne Honeyford, Tianhua Huang, Ingrid Ambus, Michael Misinai, Stephanie Robinson, Maria Muraca, Saint Doreen
North York General Hospital, Toronto Ontario Canada

          Objectives: To develop and validate an electronic tool to enhance referrals to the Familial Breast/Ovarian Cancer Clinic.
          Design/Methods: Patients attending the Breast Diagnostic Clinic were recruited for this non-randomized 3-phase study, where paper questionnaires (PQ) and surgeon assessments were traditionally used to determine Genetics referral eligibility. 
          Phase 1: Patients completed PQ (N=201).
          Phase 2: Electronic tool (ET) developed; tested for usability, readability, design, interface. Tool accuracy assessed by comparing results for patients completing both PQ & ET (N=100).
          Phase 3: Patients completed ET only (N=200). Health records reviewed to determine data accuracy.  Number of study patients eligible for referral compared with referrals received across all 3 phases. Patient/provider satisfaction assessed.
          Results: Gender, education, number of patients with breast/ ovarian cancer, age at diagnosis, family history, proportion of patients meeting referral criteria were similar across all study phases. No statistically significant difference in either number of patients eligible for referral or overall referral rates (Phase 1: 16.9%; Phase 3: 18.0%) was found. 67% of patients preferred ET over PQ. Patients found ET questions easier to complete and understand. Physician satisfaction was higher with ET in reviewing family history, identifying eligible patients, making timely referrals.
          Conclusions: The electronic tool was accurate and useful in determining referral eligibility. Similar referral rates were seen across study phases. Patients and physicians had positive experiences with the ET.  It is feasible to use this tool to identify patients eligible for Genetics referral. Further studies are in progress to investigate why some eligible patients were not referred.

Fetal jugular lymph sacs – what is the significance?
Marjan M. Nezaratia,b,c, H. Melanie Bedforda, Karen Chongb,c, Ants Toid, David Chitayatb,c
aGenetics Program, North York General Hospital, Toronto ON Canada
bDepartment of Obstetrics and GynecologyMount Sinai Hospital, Toronto ON Canada
cThe Prenatal Diagnosis and Medical Genetics Program, Mount Sinai Hospital, Toronto ON Canada
dDepartment of Medical Imaging, Mount Sinai Hospital, Toronto ON Canada

Fetal jugular lymph sacs are accumulations of lymphatic fluid in the anterolateral region of the fetal neck.  They may be isolated or occur in association with other structural anomalies and in some cases are presenting with an increased nuchal translucency or cystic hygroma.  . Their significance and the guidelines for the prenatal investigation when identified in the course of routine ultrasonography in pregnancy are still controversial.  Our protocol for the investigation of these cases includes microarray analysis and DNA analysis for the Noonan syndrome panel.  We report our experience with ten cases of prenatally diagnosed isolated fetal jugular neck cysts identified in the last 5 years (Table 1).

In 10 cases identified by us in the last 5 years, 50% had a pathogenic mutation in a Noonan syndrome Panel,  one had trisomy 21 and none had abnormality on microarray analysis (in the case with T21 we stopped the analysis following the QF-PCR results).  However, all cases with abnormal results had NT > 3.5 mm or cystic hygroma.  This study suggests that the identification of fetal jugular lymph sacs with an increased NT/cystic hygroma is associated with an increased incidence of Noonan syndrome and its related disorders and should instigate mutation analysis of the Noonan syndrome panel.

Delineating the phenotypes associated with the 15q11.2 BP1-BP2 deletion: Preliminary trends in psychometric evaluation
Melissa Cartera,, Bonnie MacKinnonb, Sanne Jilderdab, Jhoan Falconb, Pooja Swaroopb, Breanne Daleb, Abdul Noora, Anne Bassettc, D. James Stavropoulosd, Irene Drmica
aThe Hospital for Sick Children, Toronto ON Canada
bAutism Research Unit, The Hospital for Sick Children, Toronto ON Canada
cCAMH, Toronto ON Canada
dGenome Diagnostics, The Hospital for Sick Children, Toronto ON Canada

          Background. Microdeletion of the BP1-BP2 region on chromosome 15q11.2 is one of the most commonly reported copy number variants (CNVs) in the clinical population. It has been suggested that this deletion may impact cognitive, behavioral, and neurological function, as it has been observed with a higher frequency in patients being investigated for neurodevelopmental disorders (NDDs) than in controls. Retrospective case reports suffer from ascertainment bias and provide limited information about specific neurobehavioural deficits.
          Objectives. Our study aims to collect detailed phenotypic data on proband and non-proband carriers of the 15q11.2 BP1-BP2 deletion, in particular the intellectual and behavioural profiles of these individuals.
          Design/Method. Families with at least one member with 15q11.2 BP1-2 deletion were recruited from clinicians. Participants were examined by a clinical geneticist, and a structured medical and family history was obtained. Psychometric and behavioural testing protocol included standardized measures of intelligence, language ability, autistic symptomatology, attention, and mental health.
          Results. Results from the first eight participants are reported. Four participants fell within the Intellectually Disabled range (IQ<70). None of the participants met criteria for autism spectrum disorder. However, attention deficit-hyperactivity disorder (ADHD) was present in all children.
          Conclusions. Our preliminary data shows that individuals with 15q11.2 deletion may be more likely to have ADHD, mild intellectual disability and articulation difficulties. Further study of additional probands and non-proband carriers is ongoing.

RET gene analysis in families with medullary thyroid carcinoma in special populations of north-east Iran
Parastoo Kheiroddina, Fatemeh Mousavib, Babak Mahmudianc, Monireh Halimid, Seyed Ziaedin Rasihashemie, Ebrahim Sakhiniaa,b
aDepartment of Clinical Bioshimi, Division of Medical Genetics, Faculty of Medicine, Tabriz University of Medical Sciences,  Tabriz, Iran
bTabriz Genetic Analysis Centre (TGAC), Tabriz University of Medical Sciences, Tabriz, Iran.
cDivision of Nuclear Medicine, Department of Radiology. Tabriz University of Medical Sciences, Tabriz, Iran
dDepartment of Pathology, Tabriz University of Medical Sciences, Tabriz, Iran
eDepartment of Thorax Surgery, Imam Reza Hospital, Tabriz University of Medical Sciences, Tabriz, Iran.

          Objectives: Medullary thyroid carcinoma (MTC) is a neuroendocrine tumor from the para follicular C cells of the thyroid gland. It occurs either sporadically or part of an inherited syndrome. It causes by autosomal dominant mutation in the RET (Rearranged during Transfection) proto-oncogene.
          Design and Methods: The study population consisted of 20 (4 families) patients diagnosed with MTC in specific population of North-East Iran and their 3 children. The mean age at diagnosis was 29.55 years. Blood samples were collected from all subjects, genomic DNA was extracted and RET exons 10, 11, 13, 14, 15 and 16 were analyzed using PCR and sanger sequencing.
          Results: All cases had normal 10, 14, 15 and 16 exons. In all patients exon 11 had mutation in codon 634; C634R (6 of 20), C634Y (5of 20), C634G (5 of 20) and C634W (4 of 20) in each family. 11 out of 16 patients in three families (55%) had also C691S polymorphism in exon 11. Two of these three families (11patients 55%) had L769L polymorphism in exon 13 and one patient also had S765T in exon 13. 2out of 3 children had the same changes as their parents (C634R &L769L and C634G & G691S) one child had T675T change in exon 11.
          Conclusions:  It was the first study of RET proto-oncogene in this population.  Among these nucleotide substitutions, there were 2 new changes (S765T) and (T675T) in one family. These results underlined the importance of the genetic background of family members of any patient with MTC.

Multiple pathogenic variants identified by next-generation sequencing hereditary cancer panel testing – a case report
Christopher A. Tana, Marina Rabideaua, Stephanie Cohenb, Shan Yanga, Karen Vikstroma, Federico A. Monzona.
aInvitae, San Francisco, CA, USA
bSt. Vincent Hospital, Indianapolis, IN, US

          Background.  The utilization of next-generation sequencing technology to interrogate multiple genes simultaneously is being utilized more frequently in hereditary cancer testing. While this has benefits of reducing cost and allowing clinicians to cast a wide net in the elucidation of their patient’s cancer, panel testing has the potential to reveal unexpected information. We report on a proband with pathogenic variants in two different hereditary colon cancer syndromes.
          Case Report. A 39 year-old male with history of colon cancer diagnosed at 38, with normal IHC, and >20 colon polyps presented for genetic counseling. Family history was significant for a paternal aunt and paternal uncle with colon cancer in their early 50s. Both parents reportedly had colon polyps requiring frequent colonoscopy; his mother had a TAH-BSO at 40 for unknown reasons.  Testing with a 7-gene high-risk hereditary colon cancer panel identified a homozygous pathogenic variant, c.1187G>A (p.Gly396Asp), in MUTYH and a likely pathogenic duplication of exon 7 in MSH2. Due to this finding, his parents were referred for genetic counseling and testing; his mother, who was diagnosed with colon cancer in the interim, was found to carry the MSH2 duplication. Both parents were obligate carriers of the MUTYH variant.
          Conclusion. This case example demonstrates the impact of identifying multiple hereditary syndromes. Being aware of all conditions has significant impact on at-risk family members particularly those who test negative for a known familial pathogenic variant, yet could still be at risk for cancer due to a second familial pathogenic variant.

Beyond the ACMG 56: Parental choices and initial results from a comprehensive whole genome sequencing-based search for predictive genomic variants in children
M.Stephen Meyna,b,c,d,e, Nasim Monfaredb, Christian R. Marshallf,g, Daniele Mericoa,f, Dmitri J. Stavropoulosg,h, Robin Z. Hayeemsb,i, Michael Szegof,j,k, Rebekah Joblingd, Marta Girdeaa,l, Gary D. Baderc,m, Michael Brudnoa,l, Ronald D. Cohna,b,c,d, , Stephen W. Scherera,b,c,f,n, Randi Zlotnik Shaule,j,o, Cheryl Shumanc,d, Peter N. Raya,b,c,f,g, Sarah C. Bowdinb,d,e
aProgram in Genetics and Genome Biology, Hospital for Sick Children, Toronto, Ontario, Canada
bCentre for Genetic Medicine, The Hospital for Sick Children, Toronto, ON, Canada;
cDepartment of Molecular Genetics, University of Toronto, Toronto, ON, Canada;
dDivision of Clinical and Metabolic Genetics, Dept of Paediatrics, The Hospital for Sick Children, Toronto, ON, Canada;
eDepartment of Paediatrics, University of Toronto, Toronto, ON, Canada;
fThe Centre for Applied Genomics, The Hospital for Sick Children, Toronto, ON, Canada;
gDepartment of Paediatric Laboratory Medicine, The Hospital for Sick Children, Toronto, ON, Canada;
hDepartment of Laboratory Medicine and Pathology, University of Toronto, Toronto, ON, Canada;
IProgram in Child Health Evaluative Sciences, Hospital for Sick Children, Toronto, ON, Canada;
jJoint Centre for Bioethics, University of Toronto, Toronto, ON, Canada;
kCentre for Clinical Ethics, St. Joseph's Health Centre, Toronto, ON, Canada;
lDepartment of Computer Science, University of Toronto, Toronto, ON, Canada;
mThe Donnelly Centre, University of Toronto, Toronto, ON, Canada;
nMcLaughlin Centre, University of Toronto, Toronto, ON, Canada;
oDepartment of Bioethics, The Hospital for Sick Children, Toronto, ON, Canada

          Objective: The overall goal of the SickKids Genome Clinic is to pilot paediatric genomic medicine. To this end we have assessed parental interest in predictive secondary medically-actionable variants (MAVs) and used whole genome sequencing (WGS) to determine the frequency and nature of these MAVs in children.
          Design: The Genome Clinic conducts diagnostic WGS for 150+ children/year who are undergoing genetic evaluations. With parents’ permission, we search children’s genomes for predictive MAVs in 2800+ disease genes listed in the NIH Clinical Genomic Database.
          Results: Of 373 families approached to date, 56% agreed to participate. 58% of participants chose to learn their child's secondary adult-onset MAVs. Among these parents, 79% decided to learn their own status for these variants.  Bioinformatics analysis of the first 100 patient genomes yielded 2957 candidate variants in 1132 genes (~30 variants/genome.)  ~70% of candidates were listed in HGMD. However, subsequent manual assessment rejected >90% of variants for dominant diseases listed in HGMD as disease causing due to inadequate evidence of pathogenicity. After manual assessment, 33/100 children had at least one reportable predictive MAV. 9 MAVs occurred in a 2013 ACMG-guideline reportable gene. Expanding our search 50 fold to include 2800+ disease genes yielded 29 additional reportable predictive MAVs. Return of predictive MAVs and assessment of their penetrance is underway.
          Conclusions: Parental opinions vary widely regarding return of predictive MAVs and comprehensive genomic analysis can yield predictive MAVs in 1/3 of children, with the number of reportable predictive MAVs constrained by disease prevalence and imperfect variant interpretation.

Revisiting a clinical diagnosis 15 years later with the aid of whole exome sequencing: Osteopetrosis versus Harderophorphyria
Roberto Mendozaa, Lucie Dupuisb, Peter Kannub, Taila Hartleyc, Eric Barekec, Kym Boycotta
aDivision of Clinical & Metabolic Genetics, The Hospital for Sick Children, Toronto ON Canada
bThe Hospital for Sick Children, Toronto ON Canada
cCare for Rare, Children’s Hospital of Eastern Ontario, Ottawa ON Canada

          The advent of whole exome sequencing (WES) has revolutionized gene discovery and led to the identification of atypical phenotypes for well-known syndromes. Our case illustrates the phenotypic overlap between very different syndromes, harderoporphyria and infantile osteopetrosis, which was only resolved by WES.
We report two siblings, both born with severe hepatosplenomegaly, jaundice, thrombocytopenia and anemia. The eldest child had increased bone density on radiographs and was diagnosed with infantile osteopetrosis. Both children responded well to bone marrow transplant (BMT).  Molecular testing was negative for genes involved in infantile osteopetrosis:  CLCN7, TCIRG1, OSTM1, TNFSF11A and PLEKHMI1. Many years later, the family was offered whole exome sequencing to clarify the underlying molecular etiology. The siblings were found to harbor a homozygous mutation (p.K404E) in the CPOX gene confirming a diagnosis harderoporphyria. No mutations were found in any known genes causing infantile osteopetrosis.
Defects of heme biosynthesis enzymes result in porphyrias. Harderoporphyria is caused by homozygous mutations in the CPOX gene which encodes coproporphyrinogen oxidase. Individuals with harderoporphyria exhibit neonatal hyperbilirubinemia, hemolytic anemia, hepatosplenomegaly and photosensitivity. Infantile osteopetrosis presents with increased bone density, reduction of bone marrow spaces leading to anemia, hepatosplenomegaly, cranial nerves compression and severe growth failure. It is a lethal condition, but responds well to BMT.
Given that both children had undergone BMT, their own hematopoietic precursors would have been replaced by those of the donor. Thus, they would not be expected to manifest a harderoporphyria phenotype. To our knowledge, there have been no reports of increased bone mineral density in patients with harderoporphyria.
An initial clinical presentation suggesting a diagnosis infantile osteopetrosis was only challenged after the WES data returned. Since the siblings received BMT in the first year of life, it remains unknown whether increased bone density was a persistent finding. This case report illustrates the clinical utility of WES and the importance of establishing a definitive molecular diagnosis for appropriate genetic counselling and medical management.

Intravenous neonatal gene therapy corrects GM2 gangliosidoses in sandhoff mice for 'long-term', by using an AAV expressing a new hexosaminidase variant
Karlaina JL. Osmona, Evan Woodleyb, Patrick Thompsonc, Katalina Ongc, Subha Karumuthil-Melethild, Brian Marke, Don Mahuranf,g, Steven J. Grayd,h. Jagdeep S. Waliaa,b,c.
aCentre for Neuroscience Research, Queen's University, Kingston, Ontario, Canada, K7L 3N6
bDepartment of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario, Canada, K7L 3N6
cMedical Genetics/Departments of Pediatrics and Pathology and Molecular Medicine, Queen's University, Kingston, Ontario, Canada, K7L 2V7
dGene Therapy Centre, University of North Carolina, Chapel Hill, North Carolina, United States
eDepartment of Microbiology, University of Manitoba, Winnepeg, Manitoba, Canada, R3T 2N2
fGenetics and Genome Biology, Sick Kids, Toronto, Ontario, Canada, M5G 0A4
gDepartment of Laboratory Medicine and Pathology, University of Toronto, Toronto, Ontario, Canada, M5S 1A8
hDepartment of Ophthalmology, University of North Carolina, Chapel Hill, North Carolina, United States

          Background: GM2gangliosidosis is a group of neurodegenerative disorders, characterized by the malfunctioning HexosaminidaseA(HexA) enzyme, for which there is no treatment. HexA is composed of two similar, but non-identical subunits, alpha and beta, which interact to hydrolyze GM2gangliosides. Mutations in either subunit result in the development of GM2gangliosidosis. The malfunctioning HexA is unable to cleaving GM2ganglioside, whose accumulation within the neurons of the central nervous system (CNS) is neurotoxic. The resulting neuronal death induces the primary symptoms of the disease; motor impairment, seizures, and sensory impairments.
          Objectives: The aim of this study is to observe the long-term in vivo affects of a novel Hex isoenzyme, HexM treatment in a Sandhoff (beta-deficient) mouse model.
          Design/Method: Our methods include intravenous injections of neonatal mice with self-complementary vector expressing HexM at day 0-1. We monitored one cohort for 8 weeks and another cohort long-term for biochemical and behavioural analyses.
          Results: Through the enzymatic and GM2ganglioside lipid analyses, we see that with a slight increase in enzyme activity, there is a significant increase in the clearance of GM2gangliosides. On behavioural tests, the treated mice outperform their knockout age matched controls. While the untreated controls die by 15 weeks, treated animals survived to x̄=41.77 weeks. The molecular analyses reveal a uniform distribution of the vector in the CNS.
          Conclusions: The neonatal delivery of our newly synthesized viral vector expressing HexM to the Sandhoff mice provided long-term correction of the disease. This study will have implications not only for treatment of Sandhoff, but also Tay-Sachs disease (alpha-deficiency).

Clinical genetics education: Building foundations using e-modules for pediatric residents
Jennifer J. MacKenziea,b, Amy Ackera,b, Theresa Suartb, Andrea Guerina,b
aDepartment of Pediatrics, Queen’s University, Kingston, ON
bSchool of Medicine, Queen’s University, Kingston, ON

          Objectives: Clinical Genetics is rapidly evolving so it is essential that future physicians are equipped to practice in a genetics literate world. The goal of this project is to provide non-genetics trainees with an accessible resource to enhance their genetics education and to complement traditional teaching modalities. We are undertaking a pilot study of Pediatric residents’ genetics knowledge and perceptions before and after completing the e-modules.
          Design/Methods: We have created two e-modules highlighting common situations encountered in practice, a positive newborn screen and developmental disability/autism. The e-modules lead students through gathering information, interpreting findings, and management strategies. Basic genetic concepts, indications and limitations of testing are highlighted.
The modules are designed for trainees prior to exposure to Clinical Genetics, to provide a foundation to build upon with clinical experience. The modules are available through the Queen’s School of Medicine technology platform.
A questionnaire assessing knowledge and comfort with genetics will be administered before, immediately after, and 6 months after the modules. Focus groups will be conducted to determine residents’ perception of the modules and thematically analysed through a lens of constructivist grounded theory.
          Results/Conclusions: The e-modules have been designed and are in the process of being implemented. By sharing our preliminary experience we hope to expand genetics education opportunities for non-genetics residents.

A KMT2D mutation segregating in a family presenting with autosomal dominant choanal atresia reinforces the Kabuki/CHARGE connection
Lauren Badalatoa, Sali M. Farhanb, Allison Dilliottb, Robert Hegelec, Sharan Goobied
aChildren’s Hospital of Eastern Ontario, Ottawa ON Canada
bDepartment of Biochemistry and Medicine, Robarts Research Institute, Schulich School of Medicine and Dentistry, Western University
cRobarts Research Institute, London ON Canada
dMedical Genetics Program of Southwestern Ontario, London Health Sciences Centre, London ON Canada

          Background.  Choanal atresia has only been rarely reported in Kabuki syndrome, but is a common feature of CHARGE syndrome.  Otherwise, the two conditions have a number of overlapping features, and molecular links between them have recently been elucidated.
          Objectives.  We report a mother and her two children who presented with congenital choanal atresia.  All three were found to have a mutation in the KMT2D gene, suggesting that the family has an unusual presentation of Kabuki syndrome.
          Design and Methods.  Phenotypic and molecular data on the affected patients was collected.  Microarray and CHD7 sequencing were previously normal in the children.  Samples from the mother and her unaffected parents were sent as a trio for whole exome sequencing.
          Results. The facial features in the family were not typical for Kabuki syndrome, but were highly conserved among affected individuals; they included short palpebral fissures, anteverted nares, tall forehead, and prognathism.  All three had bilateral choanal atresia, high arched palate, hypodontia, hearing loss, fetal fingertip pads, and speech delay.  One or more had mild learning difficulties, bifid uvula, atrialseptal defect, and scoliosis.  A de novo dominant missense mutation in KMT2D was found in the mother (p.Q3575H) and confirmed in the two children.  This mutation has not been previously reported in the literature or in ExAC Browser.
          Conclusions. We report a unique phenotype associated with a novel mutation in the KMT2D gene.  Our findings further demonstrate the overlap of features of Kabuki and CHARGE syndromes, and reinforce the potential association between these conditions.

A Case ‘Out of the Blue’
Divya Pachata, Sudha Krishnanunnib
aConsultant Clinical Geneticist,
bHead-Department of Paediatrics, Malabar Institute of Medical Sciences- Calicut, Kerala, India

          Objective: To delineate a retrospective diagnosis in a case of severe fixed encephalopathy.
          Case Description: Propositus was a male child, first born to 3rd degree consanguineous South Asian parents. Pregnancy, delivery and clinical examination at birth were normal. During the first months of life, he had generalised hypotonia with spontaneous dystonic leg extension spasm and frequent vomiting. All the developmental milestones were delayed, he never sat alone or never attained speech. From his available photographs head size appeared to be small, and esotropia and adducted thumbs were noted. Magnetic resonance imaging of his brain showed delayed myelination. He also had gradeII/III gastro oesophageal reflux evidenced by GER Scintigraphy. Other investigations including routine biochemical and haematological parameters, neurometabolic screening and electrophysiological analysis were within normal limits. He was treated as spastic cerebral palsy from another center without much improvement to the symptoms. Child expired at the age of 2 years followed by an attack of pneumonia; clinical exome test was performed later from his DNA.
          Results: Long read sequencing revealed a homozygous, pathogenic novel variation p.Val112ProFSTer8 resulting from the deletion of exons 5-8 of CYB5R3 gene, associated with recessive hereditary methemoglobinemia (RHM) type II, which could explain the clinical profile. This large deletion was validated by polymerase chain reaction and carrier status of parents was established.
          Conclusion: Type II RHM is a rare condition, with less than 100 cases reported worldwide.

  1. Cyanosis is an invariable presenting feature and important clinical clue to methemoglobinemia. There was no history or documentation of cyanosis in our case. In the absence/failure to elicit the sign in children with severe encephalopathy, possibility of RHM might get overlooked.
  2. Majority of the reported mutations in RHM type II are point mutations with two prior documentations of single exonic deletion. Ours is the first family with the largest deletion of multiple exons of CYB5R3 gene, thus expanding the genotype spectrum.

Clinical actionability of multi-gene panel tests for hereditary breast and ovarian cancer (HBOC)
Leif W. Ellisena, Stephen E. Lincolnb, Allison W. Kurianc, Andrea J. Desmonda, Shan Yangb, Meredith A. Millsc, Yuya Kobayashib, Federico A. Monzonb, and James M. Fordc
aMassachusetts General Hospital Cancer Center, Boston NA USA
bInvitae, San Francisco CA USA
cStanford University Cancer Institute, Palo Alto CA USA

          Background: The practice of genetic testing is rapidly evolving with the recent introduction of multi-gene panels. While the prevalence of non-BRCA1/2 mutations in HBOC patients is now well documented, the clinical impact of these findings is not yet fully understood.
          Objectives: We sought to measure how often and in which ways non-BRCA1/2 findings might change patient management recommendations in a representative clinical cohort.
          Design/Method: We used similar 25 or 29 gene panels to test over 1000 BRCA1/2-negative patients, all of whom were enrolled prospectively at three academic medical centers and all met NCCN guidelines for HBOC evaluation. We established a uniform algorithm based on current practice guidelines to recommend management actions for the non-BRCA1/2 positive individuals, and we evaluated which of these recommendations would represent changes in management above and beyond any recommendations based on personal and family history alone.
          Results: 63 patients were identified with pathogenic or likely pathogenic mutations in non-BRCA1/2 genes. We found that the majority of these findings (52%) would result in consideration of additional screening and/or prevention measures for the patient. Moreover, we found that genetic testing of 70% of first-degree family members would also be warranted given the potential management changes in these individuals if found to be mutation positive.
          Conclusions: In appropriately referred patients, multi-gene panel testing yields clinically relevant findings with potential management impact for more patients than does BRCA1/2 testing alone.

Colorectal cancer patients with BRCA1 and BRCA2 mutations: Preparing for unexpected results
Karen Vikstroma, Shan Yanga, Raluca Kurza, Stephen E. Lincolna, Edward D. Esplina
aInvitae, San Francisco, CA, USA

          Background. A new paradigm in genetic panel testing for hereditary colorectal cancer (CRC) has emerged. CRC association with BRCA1/2 has been suggested, but guidelines do not include CRC in Hereditary Breast and Ovarian Cancer syndrome (HBOC). 
          Objectives. We describe 6 patients with CRC and germline mutations in BRCA1 or BRCA2, detected by multi-gene panels, to highlight actionable findings that would have been missed by traditional CRC genetic testing.
          Design/Method. 585 patients with a personal history of CRC and/or gastrointestinal (GI) polyps were tested. Variants were identified using an NGS-based cancer gene panel with CRC genes and BRCA1/2. Germline variants were classified using a point-based system based on ACMG guidelines. Clinical histories from test request forms were de-identified for analysis.
          Results. Hereditary cancer panel testing found Pathogenic (P) or Likely Pathogenic (LP) variants in 92 of 585 (15%) patients. Of the 92 mutation carriers, 69 (75%) had a P/LP variant in a CRC gene, while 6 (6%) had a P/LP variant in BRCA1/2. None of the patients with BRCA1/2 mutations reported Ashkenazi Jewish ancestry. The 4 male patients did not meet HBOC testing guidelines.
          Conclusions. In this series, a substantial minority of P/LP variants were in non-canonical CRC genes. BRCA1/2 pathogenic variants’ prevalence in the general population is insufficiently elevated to account for these findings. More research is needed to link CRC and BRCA1/2, and clinicians need to prepare themselves and their patients to deal with unexpected, potentially actionable results.

Determining a Genetic Cause for Familial Intracranial Aneurysms
Emma Hitchcocka, Jillian Diamonda, Katelin Townsendb Brian Chungb, William Gibsonb
aMcMaster University, Hamilton ON Canada
bUniversity of British Columbia, Vancouver BC Canada

          Intracranial berry aneurysms (IA) can develop in arterial walls where the endothelial layer has weakened. Subarachnoid hemorrhage (SAH) occurs when an IA bursts causing blood to flow into the brain space. SAH leads to death in 35-50% of patients, and to brain damage in 25-50% of survivors. Familial IA (FIA) is suspected when a patient has two or more first-degree relatives with IA or SAH. The risk for a ruptured aneurysm in an individual with FIA is 17 times greater compared to the general population. Currently, there is no genetic test to assess the risk of developing an IA, and repeated brain imaging tests are required to screen those at risk as affected individuals are often asymptomatic. We have identified a family with a history of FIA spanning over three-generations, with three affected and four unaffected siblings. The inheritance pattern of FIA in this family appears to be autosomal dominant, strongly suggestive of a true Mendelian disorder. Our hypothesis is that there will be a single rare pathogenic variant that leads to the development of FIA in this family. We have collected DNA samples from this family, and carried out whole-genome SNP microarray on DNA from one affected and four unaffected siblings. We also performed whole-exome sequencing (WES) on one of the affected siblings. We will further filter our list of candidate genes created from genomic areas of interest that were identified from the microarray data using our list of rare, predicted damaging, variants generated from the WES data. These areas of interest will include regions shared between affected individuals, and exclude those shared with unaffected individuals. This family is unique to our knowledge, and our research could lead to the first identification of a monogenic cause for IA.

Clinical Features of PRC2 Complex-Related Overgrowth Due to Mutations in EED
Ana Cohena, Yaoqing Shena, Steven JM Jonesa, William Gibsonb
aMichael Smith Genome Sciences Centre, Vancouver BC Canada
bWilliam Gibson, University of British Columbia, Vancouver BC Canada

          Our lab was the first to publish that constitutional mutations in the epigenetic regulator EZH2 cause Weaver Syndrome (WS). WS is characterized by overgrowth, increased height, large head, intellectual disability and susceptibility to various cancers. We found pathogenic mutations in EZH2 among 7 out of 45 individuals with Weaver-like features, of which two had developed malignancies prior to referral. Our efforts focus on determining characteristics that will help us predict the likelihood of WS patients developing cancer, through phenotype/genotype correlations and functional studies. As of yet, it is not clear whether screening for haematological malignancies in patients with Weaver syndrome would increase the patients' chances of survival.
Given that EZH2 is a histone-modifying enzyme known to be mutated in various somatic cancers, we hypothesized that constitutional mutations in other epigenetic regulators could explain the overgrowth features seen in our undiagnosed patients. To investigate this, we carried out whole exome sequencing in patients with pediatric syndromes similar to WS who did not have coding mutations in EZH2.
We have identified novel de novo mutations in EZH2’s partner protein EED in two patients to date.  Both proteins are members of the Polycomb Repressive Complex 2 that maintains gene silencing. Based on the similarities of the patients' phenotypes to WS, and on other lines of evidence including mouse Eed hypomorphs and an epigenetic signature that clustered with Weaver syndrome patients, we characterized these mutations as pathogenic. Here we describe additional features of overgrowth associated with constitutional mutations in human EED, which is also mutated somatically in various cancers. Samples from additional families are required to make a definitive link between mutations in these genes, Weaver-like syndromes, and cancer development. We continue to perform exome sequencing in patients with Weaver-like syndromes who have normal microarray studies and no mutations in EZH2 or NSD1.

Developing a multidisciplinary Fragile X and Related Conditions clinic in Victoria, BC
Molly Perrya, Gudrun Aubertinb, Jonathan Downc
aFragile X and Related Conditions Clinic, Queen Alexandra Centre for Children’s Health, Victoria, BC, Canada
bMedical Genetics, Victoria General Hospital, Victoria, BC, Canada
cQueen Alexandra Centre for Children’s Health, Victoria, BC, Canada

          Background:  Fragile-X syndrome (FXS), being the most common form of inherited intellectual disability, remains a rare disorder. Families living with FXS and related conditions in British Columbia (BC) are believed to lack healthcare involvement with clinical expertise.  Access to appropriate services, diagnostic assessments and specialized management recommendations is essential to optimal care for these patients.
          Objectives: Develop a clinical-model of multidisciplinary, family-centred care for FXS families; improved patient care through coordinated, specialist assessments; increased clinical expertise for healthcare professionals, and increased public and professional awareness of clinical services for FXS and related conditions.
          Design/Method: The Fragile X and Related Conditions (FXRC) clinic was established January 2015 with Doctors of BC, Specialist Services Committee (DOBC/SSC) funding.  A clinic coordinator was hired to implement objectives.  Public and professional awareness of services is tracked through social media and referrals to the clinic. An FXRC conference is planned with leading FXS specialists providing relevant information to parents and professionals. 
          Results: FXRC achievements include establishing a resource-based website, developing an active social media presence, and multiple presentations to differing medical disciplines including pediatrics, psychiatry and perinatal services.  A retrospective chart review from medical genetics identified 65 local individuals with a personal or family history of FXS.
          Conclusions: FXRC implementation has been challenging despite DOBC/SSC support.  Although early systemic barriers hindered clinic development, current feedback has been positive from a wide range of health and administrative professionals.  Coordination of specialist care, together with identifying the spectrum nature of FMR1 disorders, has been key to our success.

Genetic polymorphism in the vitamin D receptor gene and 25-hydroxyvitamin D serum levels in East Indian women with polycystic ovary syndrome
Dipanshu Sura, Ratnabali Chakravorty
aDept. of Obs/Gynae, ILS Hospital, Saltlake, Kolkata West Bengal, India

          Background: Polycystic ovary syndrome (PCOS) is the most common metabolic abnormality such as changes in lipid profile, diabetes, hypertension and metabolic syndrome occurring in young women of reproductive age. Low vitamin D levels were found to be associated with the development of obesity and insulin resistance in women with PCOS. Variants on vitamin D receptor (VDR) gene have also been related to metabolic comorbidities in general population.
          Aim: The aim of this case-control study was to investigate whether the VDR gene polymorphisms are associated with susceptibility to PCOS.
          Methods: Women with PCOS and a control group, all aged 16-40 years, were enrolled. Genotyping of VDR Fok-I (rs2228570), VDR Apa-I (rs7975232) as well as GC (rs2282679), DHCR7 (rs12785878) SNPs between groups were determined by using direct sequencing. Serum 25-hydroxyvitamin D [25(OH)] levels were measured by ELISA.
          Results: Mean serum 25(OH)D in the PCOS and control samples were 19.08±7 and 23.27±6.03 (p=0.048) which were significantly lower in PCOS patients compared with controls.  CC genotype of the VDR Apa-I SNP was same frequent in PCOS (25.6%) and controls (25.6%) (OR: 0.9995; 95%CI: 0.528 to 1.8921; p= 0.9987). The CC genotype was also significantly associated with both lower E2 (p=0.031) and Androstenedione levels (p=0.062). We observed a significant association of GC polymorphism with 25(OH)D levels. PCOS women carrying the GG genotype (in GC genes) had significantly higher risk for vitamin D deficiency than women carrying the TT genotype.
          Conclusions: In conclusion, data from this study indicate that vitamin D levels are lower, and vitamin D deficiency more frequent, in PCOS than in controls. The present findings suggest that the Apa-I, Fok-I polymorphism of the VDR gene is associated with PCOS and seems to modulate ovarian steroid secretion. Further studies are needed to better clarify the biological mechanisms by which the polymorphism influences PCOS risk.

Posters MG-120 to MG-123

Chorionic villus sampling: 3 cases with discrepancies between iFISH, array-CGH and karyotype
France Léveilléa, Sonia Nizarda, Géraldine Mathonneta, Emmanuelle Lemyrea et Frédérique Tihya
aService de Génétique Médicale, Laboratoire de Cytogénétique, CHU Sainte-Justine, Montréal, Québec

          Background. Chorionic villus sampling (CVS) is the method of choice for prenatal diagnosis (PND) in first-trimester pregnancies (10-13th weeks and 6 days), particularly for high risk of fetal abnormalities. This procedure allows earlier diagnosis in order to enable effective pregnancy management. Microarray analysis is now performed on CVS.
          Objectives/Method. We review 3 cases using fluorescence in situ hybridization on uncultivated nuclei (iFISH), CGH array results on DNA extraction from the entire villi and karyotype analyses on cultured CVS.
          Results. Two cases showed normal iFISH profile for chromosomes 13, 18, 21, X and Y, while the CGH array revealed a weak mosaicism for monosomy X in one CVS case and for trisomy 18 in the other. Karyotypes showed 45,X and 47,XY,+18 in more than 95% of the cells. In the third case, the iFISH and the CGH array were normal, but the karyotype revealed a trisomy 9 in 76% of the cells.
          Conclusions. CVS mosaicism and feto-placental discrepancies are rare phenomena occurring in ~1–2% of the PND. Cytogenetic analysis of CVS may give different results since CVS is from various placental cell lineages. Therefore, a cultured CVS (mesenchyme) analysis should be completed in order to reduce the incidence of false-positive or false-negative findings. In those cases, amniocentesis follow-up could be offered to complete the prenatal diagnosis.

Complexity of phenotypes of females with unbalanced X-autosomal translocations exemplified by a case with 46,X,der(X)t(X;16)(p11.2;p13.2) karyotype
Daria Grafodatskayaa , Edith Dellb, Chumei Lib, Elizabeth McCreadya
aPathology and Molecular Medicine, McMaster University, Hamilton, ON, Canada
bClinical Genetics Program, McMaster Children's Hospital and McMaster University, Canada

          Background: Females with unbalanced X-autosomal translocations frequently exhibit skewed X-chromosome inactivation, as  cells with the least functional imbalance have selective advantage. However, predicting the inactivation status of the translocated autosomal segment and its impact on phenotype is challenging. Active vs inactive state can depend on translocation breakpoint and sequence features of the translocated segment.
          Objective: Report a case with 46,X,der(X)t(X;16)(p11.2;p13.2) karyotype and discuss karyotype/phenotype correlations.
          Results: We report a 18 month old girl with intrauterine growth restriction, failure to thrive, cardiac anomalies and dysmorphic features.  Oligonucleotide microarray has revealed a terminal ~9 Mb gain of chromosome 16p and terminal loss of most of Xp. A de novo unbalanced translocation t(X;16)(p11.2;p13.2) was confirmed by karyotype analysis and family studies. Androgen receptor assay further showed complete skewing of X-inactivation, suggesting that derivative X-chromosome was inactive. Phenotype of our patient was not fully consistent with either Xp deletion or 16p13.3 duplication, leaving a question about inactivation status of the translocated 16p material. A case with 46, X,der(X) t(X;16)(q28;p12) karyotype and phenotype consistent with  16p13.3 duplication syndrome was previously  reported . Replication studies have shown skewed inactivation of the der(X), not spreading into 16p translocated segment Am J Med Genet 1996; 61:117-21).
          Conclusions: Our patient phenotype suggests that while Androgen Receptor assay provides valuable information in regard of X-inactivation skewing, it has limitations for karyotype/phenotype correlations for patients with unbalanced X-autosomal translocation. Additional analysis of inactivation status of the genes of the translocated segment is underway to understand the phenotype of our patient.

Atypical FISH Pattern Consistent with Insertion of 5’ CBFB into 16p13 in a Case of Therapy-related Acute Myeloid Leukemia
Adeel Mohammad Razaa, Jessica Sanchezb, Daniel Snowerc, Adewale Adeyinkad
aHouston Methodist Hospital, Houston TX USA
bHenry Ford Health System, Detroit MI USA
cSt. John Hospital and Medical Center, Detroit MI USA
dQueen’s University/Kingston General Hospital, Kingston ON Canada

          Background: The inv(16) and related t(16;16) resulting in fusion of CBFB gene (16q22) to MYH11 (16p13) are not commonly observed in therapy-related AML (t-AML).
          Objective: To present a case of t-AML with CBFB/MYH11 fusion and rearrangement of chromosome 16 other than the commonly described inv(16) and related t(16;16).
          Case report: A 46-year-old woman presented with t-AML 1.25 years after adjuvant radiation and chemotherapy for rectal carcinoma. Peripheral blood showed marked reduction in neutrophils, 24% blasts and increased lymphocytes and monocytes. Bone marrow aspirates revealed large blasts with abundant cytoplasm, fine nuclear chromatin, occasional deep nuclear folds and prominent nucleoli. Bone core biopsy mainly consisted of blasts and was remarkable for numerous eosinophilic myelocytes and metamyelocytes. Cytogenetic analysis of bone marrow aspirate was interpreted as 46,XX,inv(16)(p13.3q22)[6]/47,idem,+8[14]/48,idem,+6,+8[2]. Interphase FISH analysis using the CBFB break-apart probe showed an atypical abnormal signal pattern of two CBFB fusion signals and one 5’ CBFB probe signal. Metaphase FISH analysis showed an abnormal chromosome 16 with a CBFB fusion signal on one arm and a 5’ CBFB signal on the other arm. The normal 16 homologue showed the expected one CBFB fusion signal. The CBFB/MYH11 fusion transcript type A was detected by RT-PCR.
          Conclusion: The abnormal CBFB pattern observed in this case differs from what is expected for an inv(16) or t(16:16), but can be explained by an insertion of 5’ CBFB into 16p13. This case demonstrates that insertion of 5’ CBFB into 16p13 is another mechanism of producing a CBFB /MYH11 fusion.

Exonic and intronic NRXN1 deletions: novel genotype-phenotype correlations
Chelsea Lowther1, Marsha Speevak2, Christine Armour3, Elaine Goh2, Gail Graham3, Chumei Li4, Susan Zeesman4, Malgorzata J.M. Nowaczyk7, Lee-Anne Schultz6, Antonella Morra2, Robert Nicolson7, Manjulata Rajguru8, Sharon Goobie9, Mark A. Tarnopolsky4, Chitra Prasad9, Paul T. Dick10, Asmaa S. Hussain11, Matthew Gazzellone12, Anath C. Lionel12, Christian R. Marshall12, Stephen W. Scherer12, Dimitri J. Stavropoulos13, Elizabeth McCready6, Anne S. Bassett1,14
1Institute of Medical Science, University of Toronto, Toronto, ON, Canada
2Trillium Health Partners Credit Valley Site, Toronto, ON, Canada
3Department of Pediatrics, Faculty of Medicine, University of Ottawa, Ottawa, ON, Canada
4Department of Pediatrics, McMaster University, Hamilton, ON, Canada
5Hamilton Health Sciences, Clinical Genetics Program, Hamilton, ON, Canada
6Department of Pathology and Molecular Medicine, McMaster University, Hamilton, ON, Canada
7Departments of Psychiatry, Pediatrics, Psychology and Medical Biophysics, Schulich School of Medicine and Dentistry, Western University, London, ON, Canada
8Cambridge Memorial Hospital, Cambridge, ON, Canada
9Department of Pediatrics, Schulich School of Medicine and Dentistry, London, ON, Canada
10Grey Bruce Health Services, Owen Sound, ON, Canada
11London Health Sciences Centre, Children’s Hospital of Western Ontario, London, ON, Canada
12The Centre for Applied Genomics, The Hospital for Sick Children, Toronto, ON, Canada
13Cytogeneitcs Laboratory, Department of Pediatric Laboratory Medicine, The Hospital for Sick Children, Toronto, ON, Canada
14Department of Psychiatry, Faculty of Medicine, University of Toronto, Toronto, ON, Canada

          Objectives: To 1) further characterize the extended phenotype of exonic NRXN1 deletions and 2) systematically evaluate the pathogenicity of intronic NRXN1 deletions in order to help inform clinical diagnostic practice.
          Methods: We examined clinically ascertained cases from three Canadian cytogenetic laboratories for exonic NRXN1 deletions. Referring physicians completed a clinical checklist to identify major lifetime features and illnesses. One lab’s clinically referred cases were available to investigate the prevalence of intronic NRXN1 deletions in comparison to 15,254 controls. Genome-wide CNV data from high-resolution microarrays were investigated for the presence of additional rare variants.
          Results: We identified 41 (0.21%) exonic NRXN1 deletions among 19,263 clinically referred cases, an over eight-fold increase compared to controls (p<0.0001). Novel phenotypes identified in two or more exonic NRXN1 deletion cases included nine adults with comorbid intellectual disability and a psychiatric illness, movement disorders, automutilation, sleep disorders, and obsessions and preoccupations. The prevalence of congenital anomalies was low. The prevalence of intronic NRXN1 deletions did not differ between clinical cases (19/6,022; 0.32%) and controls (55/15,524; 0.35%). Additional pathogenic rare variants were four times more common in intronic (n=6/19; 31.6%) compared to exonic (n=3/41; 7.3%) NRXN1 deletion cases.
          Conclusions: The expression of exonic NRXN1 deletions is primarily neuropsychiatric and may be associated with comorbid intellectual disability and a psychiatric illness in adulthood. To our knowledge this is the first study to demonstrate that the majority of intronic NRXN1 deletions by themselves are unlikely to cause clinical phenotypes, however further study on their potential functional impact is needed.

Posters MG-124 to MG-136

THE INVESTICATE project: Identification of New Variation, Establishment of Stem cells, and TIssue Collection Advancing Treatment Efforts
Walla Al-Hertania, Naguib Mechawarb, Carl Ernstb
aAlberta Children’s Hospital; University of Calgary, Calgary, AB, Canada
bDouglas Hospital Research Institute and brain bank; McGill University, Montreal, QC, Canada

          Objective: Neurodevelopmental disorders (NDDs) are a large and complex group of disorders with varied etiologies. Recent advances in sequencing, induced stem cells, and small molecule screening technologies provide an opportunity to develop personalized treatment for NDDs.
          Methods: The INVESTICATE project recruits patients with NDDs from Children’s Hospitals in Canada and internationally.  Patients are enrolled if they have a similarly affected sibling and negative genetic tests, or a de novo balanced chromosomal rearrangement (BCR).  We use Next-Generation Sequencing tools to find variation and structural variant breakpoints.  Fibroblasts from patients undergo rapid induced pluripotent stem cell (iPSC) re-programming, neural progenitor cell (NPC) differentiation, CRSIPR/Cas9 mutation correction, and finally cell phenotyping.  Where feasible, we collect brains from cases with reduced life expectancy.  Patient-derived NPCs undergo high-throughput small molecule screening to reverse cell phenotypes associated with disease.
          Results:  INVESTICATE has recruited six families, and we identified mutations in genes not previously associated with NDDs.  We identified a stop codon altering single base deletion in one family, a 51-basepair promoter deletion in another family, and a gene truncating chromosomal translocation implicating chromatin remodelling, netrins, and maintenance of brain pH in NDDs.  Functional assays using iPSC-NPCs of these rare variants support their role in disease.
          Conclusions:  INVESTICATE is a rapid bedside-to-bench and back again pipeline capable of finding variants missed using standard methodology, and complements variant detection with a full battery of cell phenotyping assays, brain collection, and high-throughput screening.  INVESTICATE is well positioned to attempt to provide cost-effective, personalized care to children with NDDs

CYP21A2 mutation spectrum in congenital adrenal hyperplasia identified from molecular genetic testing
Yanwei Xia, Jillian Parboosinghb, Heather Johnsona, Lisa Grahama, Ryan Lamontb
aAlberta Children’s Hospital, Calgary AB Canada
bUniversity of Calgary, Calgary AB Canada

          Background. Mutations in CYP21A2, the gene encoding 21-hydroxylase, are identified in >80% of individuals with congenital adrenal hyperplasia (CAH). CYP21A2 and its highly homologous pseudogene, CYP21A1P, reside in close proximity; as a result >90% of CYP21A2 mutation-containing alleles are caused by gene conversion from CYP21A1P or CYP21A2 deletions arising from non-allelic homologous recombination. Further, CYP21A2 duplications exist and provide a source of false negative and false positive results as carriers of a CYP21A2 deletion may be masked by a CYP21A2 duplication on the opposite allele (2+0 configuration) or carriers of a point mutation may also carry a second non-mutated CYP21A2 in cis.
          Objectives. The purpose of this study was to investigate the distribution of CYP21A2 mutations identified from molecular genetic tested CAH individuals and determine the frequency of 2+0 carriers in a control population.
          Methods. A total of 469 CAH individuals were screened for CYP21A2 mutation by MLPA and common mutation and/or sequencing analysis., A PCR-based assay was developed and to detect 2+0 CYP21A2 deletion carriers.
          Results. The p.V282L, c.293-13A/C>G, and CYP21A2 gene deletion were the most frequent mutations identified in CAH. Cases with two independent mutations in cis and mutation-containing CYP21A2 duplications were observed. No silent 2+0 CYP21A2 deletion carriers were identified in our control population.
          Conclusions. Segregation and copy number analysis are critical in CYP21A2 testing. A PCR-based assay can be used to detect CYP21A2 duplications, although the 2+0 silent carrier frequency is rare.

Data sharing and Variant Classification Consensus Building in the Canadian Open Genetics Repository (COGR)
Marina Wanga, Shana Whitec, Kathleen-Rose Zakoora, Andrew H. Girgisa, Matthew S Lebocd, Jordan Lerner-Ellisa,b, and the Canadian Open Genetics Repository Working Groups (opengenetics.ca)
aLaboratory Medicine and Pathobiology & Pathology and Laboratory Medicine, University of Toronto, Mount Sinai Hospital, Toronto, ON, Canada
bOntario Institute for Cancer Research, Toronto, ON, Canada
cLaboratory for Molecular Medicine, Partners HealthCare, Cambridge, MA, USA
dDepartments of Pathology, Harvard Medical School and Brigham and Woman’s Hospital, MA, USA

          Background:  There is a critical need for collaborative measures between Canadian institutions to better facilitate variant analysis and data sharing.
          Objectives: The Canadian Open Genetics Repository (COGR) is a collaborative effort for the collection, sharing and analysis of variants reported by medical diagnostics laboratories across Canada. The project focuses on reaching consensus agreements on variant classification among clinical laboratories through data sharing and analysis and disseminating such information to a large, public data repository.
          Design/Method: COGR provides laboratories a custom Variant Assessment Tool, and facilitates sharing through GeneInsight®, a database capturing variant interpretations, reference sequence data, and gene-disease associations. Agreements and discrepancies for individual variant interpretations were identified, and a voting system was put in place to attempt to reach consensus on discrepant classifications.
          Results: The COGR network currently contains over 3,000 variants across 23 genes associated with 10 diseases. There are 46 variants seen by at least three laboratories with fully consistent classifications. A total of 96 variants had discrepant classifications across at least two laboratories. When targeting the 5 most discrepant variants for review via a presentation and anonymous voting, only 1 variant reached consensus.
          Conclusions: By sharing data among laboratories, consensus classifications could quickly be reached for a subset of variants. However, despite presenting evidence for variant classification and discussions amongst experts in the field, there is still considerable difficulty reaching consensus for ambiguous variants. This highlights the need for structured and rule-based variant review. The COGR is facilitating collaboration between Canadian laboratories and international efforts through data sharing and consensus building.

Residual disease monitoring in a retinoblastoma patient by PCR of a novel deletion breakpoint
Tim Corson1, Hilary Racher2, Bob Argiropoulos3, Helen Chan4, Renée Perrier3, Donco Matevski2, Diane Rushlow2, Furqan Shaikh4, Heather Trang4, Brenda Gallie4
1Eugene and Marilyn Glick Eye Institute, Indianapolis, IN, USA
2Impact Genetics, Bowmanville, ON, Canada
3Alberta Children’s Hospital, Calgary, AB, Canada
4Hospital for Sick Children, Toronto, ON, Canada

          Background: Allele-specific (AS)-PCR for known point mutations is a high-sensitivity method to monitor for minimal residual disease (MRD) in metastatic retinoblastoma. However, this approach is difficult to apply to large RB1 gene deletions.
          Objective: We describe a novel, high sensitivity molecular assay for MRD monitoring of a large RB1 deletion.
          Method: The proband’s retinoblastoma RB1 deletion was identified using quantitative multiplex (QM)-PCR.  Array comparative genomic hybridization (aCGH) was used to map the boundaries of the tumor RB1 deletion. Inverse PCR was then applied to capture the unique deletion breakpoint, and AS primers were optimized for high sensitivity surveillance for MRD.
          Result: Full RB1 gene testing revealed no mutation in blood of this patient. Using QM-PCR, we found one RB1 mutation in the retinoblastoma tumor (del P->11). aCGH confirmed an ~238 kb hemizygous deletion extending centromeric from RB1. The precise deletion breakpoint was determined using inverse PCR to amplify from the known two-copy flanking sequence to identify a PCR product in tumor DNA that was absent from the patient’s blood DNA.  AS primers based on the breakpoint detected a tumor-specific PCR product with a sensitivity of 1 in 1000 cells.  The original bone marrow (BM) prior to therapy was strongly positive, but all subsequent BM, CSF, and cells harvested for stem cell transplant have been negative for the tumor-specific marker.
          Conclusion: aCGH followed by inverse PCR provides a cost- and time-effective way to map QM-PCR detected RB1 breakpoints for high sensitivity molecular surveillance for MRD.

Variants at Polycomb Repressive Complex 2 (PRC2) interacting non-coding RNAs (ncRNAs) are connected with differential regulation of key cellular functions
Athanasios Zovoilis1, Jeannie T Lee1
1Department of Genetics, Harvard Medical School, Boston, USA

          Background:  Traditionally, research in molecular genetics has focused on the 3% of the genome that encodes for proteins. Accumulating evidence suggests that many of the ncRNAs produced from the rest of the genome may be functional but it remains unclear whether non-coding variants at these loci are associated with predisposition to diseases.
          Objectives: Given the size of the non-coding genome that has to be screened, an efficient integrative genomics approach is needed to identify genomic elements that would be good candidates for further genetic testing. Here we present such an approach.
          Design/Method: We reasoned that searching for ncRNAs that bind chromatin modifiers already connected with human disease would increase the chances for identification of ncRNAs with a possible role in disease. In order to identify such candidates we used RIPsequencing and CLIPsequencing to identify ncRNAs that bind PRC2.
          Results: Surprisingly, a number of these RNAs, in response to cell signaling, are processed further into shorter distinct RNA fragments with different biological activity. In vitro mutations that altered this process led to RNA fragments that bind PRC2 with different efficiencies resulting in apoptosis.  We were able to identify a similar biological activity in the respective human orthologs of these ncRNAs.
          Conclusions: Our findings reveal a novel mode of regulation of cell function by a class of ncRNAs. Non-coding variants at genetic loci producing these RNAs would be good candidates for connection with human disease.

Our experience of in silico gene panel testing for clinically heterogeneous disorders using exome sequencing.
Raveen K. Basrana, Christian R. Marshalla, Adam Shliena, Marianne Elioua, Jennifer Orra , Lynette Laua, Dimitri J. Stavropoulosa, Peter N. Raya
aMolecular Genetics Laboratory, Genome Diagnostics, Department of Paediatric Laboratory Medicine, The Hospital for Sick Children, Toronto, Canada

          Background. The implementation of next generation sequencing technology (NGS) in a molecular diagnostics laboratory has resulted in a major transformation in service delivery.  The ability to generate accurate sequence data from thousands of genes in a single experiment has facilitated the development of comprehensive gene panel tests for genetically heterogeneous disorders. Previous methodology of gene panel testing included Sanger sequencing, which was time-consuming, labor-intensive and costly and therefore restricted the number of genes that were included in a panel.
          Methods/Results. At The Hospital for Sick Children (Toronto, Canada), we have developed and clinically validated in silico gene panels using whole exome sequencing (WES) for several heterogeneous disorders including hereditary spastic paraplegia (HSP), connective tissue and bone disorders (CT), hereditary hearing loss (HL), Noonan syndrome (NS) and autoimmune disorders (AI).  Our laboratory’s current strategy of using WES has several advantages including: (1) standardized workflows that facilitate the development of additional gene panels; (2) expansion of current gene panel as new disease associations are discovered; and (3) building of an internal database of allele frequencies to aid in interpretation of variants. Through our validation studies we have shown high reproducibility and accuracy with >99% sensitivity for detection of single nucleotide variants (SNVs) and >95% for detection of small insertion/deletions (indels).
          Conclusions. NGS technologies have enabled our laboratory to expand our testing menu to include genes that comprise part of the differential diagnoses for several disorders which will lead to improved detection rates, increased genetic diagnoses and ultimately better clinical care.

Utilizing whole exome sequencing to identify causative variants in genetically heterogeneous disorders.
Dimitri J. Stavropoulosa, Christian R. Marshalla, Raveen K. Basrana, Lynette Laua, Marianne Elioua, Jennifer Orra, Eriskay J. Listona, Sarah C. Bowdinb, M. Stephen Meynb,c,d, Melissa T. Carterb, Chris Carewe, Ronald D. Cohnb,c,d,e, Adam Shliena,c, Peter N. Raya,c,d,e,f  
aGenome Diagnostics, Department of Paediatric Laboratory Medicine, The Hospital for Sick Children, Toronto, ON, Canada;
bDivision of Clinical and Metabolic Genetics, The Hospital for Sick Children, Toronto, ON, Canada;
cProgram in Genetics and Genome Biology, Hospital for Sick Children, Toronto, Ontario, Canada;
dDepartment of Molecular Genetics, University of Toronto, Toronto, ON, Canada;
eCentre for Genetic Medicine, The Hospital for Sick Children, Toronto, ON, Canada;
fThe Centre for Applied Genomics, The Hospital for Sick Children, Toronto, ON, Canada;

          Background The rapid pace of innovation within the era of genomic sequencing has provided unprecedented insights into the genetic basis of human disease.  Clinical whole exome sequencing provides a means for clinicians and clinical labs to take advantage of this ongoing expansion of knowledge, to benefit patient care and offer individualized medicine.
          Methods/Results We have developed a workflow and customized informatics pipeline suitable for processing large numbers of clinical exomes to achieve a diagnosis for genetically heterogeneous disorders.  In order to guide genetic variant interpretation, we have implemented the Phenotips software tool within the hospital to collect detailed phenotype information from physicians, through a web-based interface.  We describe our exome variant interpretation algorithm that integrates multiple control and clinical databases, as well as our on-site patient database to achieve a diagnosis.  In order to maximize diagnostic yield, our analysis includes variants affecting all genes known to cause genetic disorders, which are prioritized according to phenotypic information provided by the referring physician.  In addition, we provide patients the option of reporting secondary findings as described in the ACMG guidelines.
          Conclusions This comprehensive approach provides an effective strategy for identifying causative variants in patients with genetically and phenotypically heterogeneous disorders, who would otherwise remain undiagnosed.

ZNF259 is a candidate gene for Alopecia-primordial dwarfism-renal syndrome (APDRS)
Amanda Smitha, Carol Clericuzioa , Afsana Ahmeda, Laura McDonella, Sarah Sawyera, Dennis Bulmanb, Kym Boycottc
aChildren’s Hospital of Eastern Ontario, Ottawa ON Canada
bResearch Institute & Newborn Screening ON, Children’s Hospital of Eastern Ontario, Ottawa ON Canada
cDepartment of Genetics, Children’s Hospital of Eastern Ontario, Ottawa ON Canada

          APDRS is characterized by congenital alopecia, dwarfism and hypoplastic kidneys. This syndrome was observed in 5 Hispanic New Mexican children from four families. All had symmetric intrauterine growth and progressively severe postnatal growth restriction.  Skin color is very pale with a porcelain-like appearance. Affected children have a prominent forehead, deep-set eyes, downturned corners of the mouth and full cheeks. One had cleft palate, two had shunted hydrocephalus and both males had genital hypoplasia. All had severe global delays and sensorineural hearing loss. Four children died within the first two years of life, from uremia and/or sepsis. The surviving patient is a 16 yo girl with renal failure, central fat distribution and type1 diabetes.
Whole exome sequencing performed on the surviving patient and her unaffected parents yielded a homozygous missense mutation at a highly conserved locus of the ZNF259 gene, both parents were carriers. Sanger sequencing of unaffected parents and siblings of a deceased patient demonstrated parents and one sibling were heterozygous carriers of the mutation. The parents of a third affected were found to be heterozygous carriers of the same mutation, patient DNA was unavailable.  These results suggest a founder mutation in these families.
ZNF259 encodes a zinc finger protein that has been demonstrated to play an important role in transcription, DNA synthesis and cell proliferation.  Down-regulation of ZNF259 results in arrest at G1 and G2 phases of cell cycle.  Western blot on patient fibroblasts demonstrated a loss of ZNF259 protein and cell cycle analysis is currently underway.

Diagnostic utility of whole genome sequencing in pediatric medicine
Christian R. Marshall1, Dmitri J. Stavropoulos1, Rebekah Jobling2, Daniele Merico3, Sarah Bowdin2, Nasim Monfared4, M. Stephen Meyn2,5, Michael Szego6, Randi Zlotnik Shaul7, Bhooma Thiruvahindrapuram3, Giovanna Pellecchia3, Thomas Nalpathamkalam3, Michael Brudno5,8, Marta Girdea8, Robin Hayeems9, Chris Carew4, Raith Erickson10, Richard A. Leach10, Cheryl Shuman2,5,11, Peter N. Ray1,5,11, Ronald D. Cohn2,4,5, Stephen W. Scherer3,4,5,11
1Genome Diagnostics, Department of Paediatric Laboratory Medicine, The Hospital for Sick Children, Toronto, ON, Canada;
2Division of Clinical and Metabolic Genetics, The Hospital for Sick Children, Toronto, ON, Canada;
3The Centre for Applied Genomics, The Hospital for Sick Children, Toronto, ON, Canada;
4Centre for Genetic Medicine, The Hospital for Sick Children, Toronto, ON, Canada;
5Program in Genetics and Genome Biology, Hospital for Sick Children, Toronto, Ontario, Canada;
6Joint Centre for Bioethics, University of Toronto, Toronto, ON, Canada;
7Department of Bioethics, The Hospital for Sick Children, Toronto, ON, Canada
8Department of Computer Science, University of Toronto, Toronto, ON, Canada;
9Child Health Evaluative Sciences, The Hospital for Sick Children, Toronto, ON, Canada;
10Complete Genomics Inc, Mountain View, CA, USA
11Department of Molecular Genetics, University of Toronto, Toronto, ON, Canada

          Background: Chromosome microarray analysis (CMA) is the current first-tier clinical investigation for pediatric patients with congenital malformations and/or neurodevelopmental disorders. Whole genome sequencing (WGS) promises to capture all classes of genetic variation in a single experiment, but the diagnostic yield of WGS compared to CMA in the clinical setting has not been established.
Objectives: The purpose of the study was to compare the diagnostic yield of WGS with CMA and other targeted sequence analysis.
Methods: Through the SickKids genome clinic 100 consecutive patients undergoing CMA were recruited and WGS was performed to compare diagnostic yield.
Results: WGS identified variants meeting clinical diagnostic criteria in 32% of cases, representing a 4-fold increase in diagnostic rate over CMA (8%) alone and >2-fold increase in CMA plus targeted sequence testing (15%). WGS identified all reportable rare CNVs that were detected by CMA. In an additional 27 patients, WGS revealed clinically significant SNV and indel mutations presenting in a dominant (72%) or a recessive (28%) manner. Four cases had variants in at least two genes involved in distinct genetic disorders, contributing to a more complex clinical phenotype.
Conclusions: Our data indicate that WGS is highly accurate and efficient, providing a diagnosis in 32% pediatric patients that meet clinical criteria for CMA.  Clinical implementation of WGS as a single and primary molecular test will provide a higher diagnostic yield than conventional testing while decreasing the number of genetic tests and ultimately the time before reaching a genetic diagnosis.

Development and launch of an expanded pan-ethnic carrier screening panel
Ashley H. Bircha, Guiqing Caia, Xiaoqiang Caia, Anastasia Fedicka, Lama Elkhourya, Janelle McCarthya, Maria Delioa, Jinglan Zhangb, Ruth Kornreicha, Lisa Edelmanna
aMount Sinai Genetic Testing Laboratory, Department of Genetic and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, New York, USA
bBaylor Miraca Genetic Laboratories, Baylor College of Medicine, Houston, TX, USA

          Background. Prenatal carrier screening has traditionally focused on well-defined ethnic groups with an increased chance of being a carrier for specific genetic diseases. However, many people are of mixed ethnicity or cannot accurately identify their full ethnic background. With the ability to test for hundreds of diseases at one time, expanded carrier screening panels are an attractive option.
Objectives. To build a comprehensive pan-ethnic carrier screening panel with a rapid turnaround time (TAT).
Design/Method. Following literature review, internal research, and physician input, a panel of 281 autosomal recessive and X-linked diseases were selected. Next generation sequencing (NGS) was performed using Agilent’s SureSelect QXT target enrichment approach followed by Illumina HiSeq 2500 Rapid Run mode sequencing. Additional methodologies include MLPA, Asuragen CGG repeat analysis/Southern blotting, and Sequenom/Luminex genotyping assays to assay variants not amenable to NGS. Variant confirmation is performed simultaneously by genotyping, or subsequently by Sanger sequencing. Over 3,700 recurrent pathogenic variants with a well-established relationship to disease phenotype are guaranteed to be sequenced at a depth

Validation of a customizable next generation sequencing panel for ear and eye diseases
Neal Codya, Maria Delioa, Jinlian Wanga, Geetu Vija, Guiqing Caia, Brynn Webba,b, Amy Yanga,b, Yumi Kasaia, Steven Sperbera, Ruth Kornreicha, Lisa Edelmanna
a Department of Genetic and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, New York, USA
b Department of Pediatrics, Mount Sinai School of Medicine, New York, New York 10029, USA

Background. Hearing and/or vision loss can result from both genetic and non-genetic etiologies. Up to 70% of prelingual hearing loss has a genetic basis, as does up to 60% of congenital blindness among infants. Next generation sequencing (NGS) technology is ideal for diagnostic testing for such disorders due to extreme locus heterogeneity and phenotypic overlap.
Objectives.  The aim is to design and validate a comprehensive, customizable NGS panel for detection of pathogenic variants in genes involved in hearing and/or vision loss.
Design/Method380 genes were selected for inclusion based on literature review and comparison with commercially available assays, including 245 genes for vision loss and 83 genes for hearing loss.  Agilent SureSelect biotinylated RNA baits were designed to capture 1.5 Mb of DNA using SureSelect XT target enrichment followed by Illumina HiSeq 2500 sequencing.  DNA from six HapMap cell lines and five Coriell repository samples with known pathogenic variants were used to assess intra-run and inter-run variability. Additionally, DNA from a family with a known hearing loss etiology was included.
Results.  An average yield of 6 Gb of sequence per sample was obtained, where 94% of bases were covered at a depth >20XGenotype calls from HapMap samples were highly concordant across three runs. Eight known pathogenic variants were confirmed in GJB2, USH1C, MARVELD2, TRIOBP, and OAT.
ConclusionsThis NGS panel targets an extensive list of genes implicated in diseases of the ear and the eye. Customizable testing allows ordering of an ear-specific or eye-specific gene panel, or both.

p300 regulates glucose homeostasis by maintaining islet mass
Chi Kin Wonga, Francis Lynna, Paul Brindlea, William Gibsonb
aSt. Jude’s Children’s Research Hospital, Memphis TN USA
bUniversity of British Columbia, Vancouver BC

          We recently identified a patient with Rubinstein-Taybi syndrome (RTS) who also had early-onset diabetes at the age of 23 years despite a thin body mass index of 23 kg per sq.metre. She has retained endogenous insulin production and is controlled on oral hypoglycemics. Such a phenotype has not been associated with RTS previously. RTS is caused by autosomal dominant mutations in CREBBP or EP300; she had a microdeletion affecting one EP300 allele. p300 is an important transcriptional coactivator with histone acetyltransferase activity. Other studies have suggested that p300 coactivates NeuroD1, HNF4a and PDX1, which are transcription factors important for beta cell function and identity. Based on these lines of evidence, we hypothesized that p300 is required to maintain beta cell function, and that deletion of one allele of the gene may cause the diabetes in our patient. To test this hypothesis, we have generated mice missing p300 in Ngn3+ pancreatic islets by the Cre-loxP system. Mice lacking p300 in their pancreatic islets develop glucose intolerance at the age of eight weeks, which is caused by hypoinsulinemia. These phenotypes recapitulated a key portion of our patient’s phenotype. Immunostaining confirmed reduced total islet mass in the pancreata of p300IsletKO mice, involving the loss of both beta cells and alpha cells. At the cellular level, p300-null islets display normal insulin synthesis and glucose-stimulated insulin secretion. We carried out RNA-seq and qPCR to identify transcriptome-wide changes in p300-null islets. We found that several genes previously associated with islet proliferation are differentially expressed. Our experiments suggest that p300 acts as a critical regulator of islet mass and that its expression in islets is required for maintaining glucose homeostasis.
The project is supported by Canucks for Kids diabetes catalyst grant and NSERC discovery grant (RGPIN 402576-11).

Functional Analysis of the Autism and Intellectual Disability Gene PTCHD1 Reveals Hedgehog Receptor-Like Functions and PDZ-Binding Domain-Specific Regulation of CNTNAP1 and NLGN1
Kirti Mittal1, Bryan Degagne1, Taimoor Sheikh1, John B. Vincent1
1Molecular Neuropsychiatry & Development (MiND) Lab, Campbell Family Mental Health Research Institute, Centre for Addiction and Mental Health, Toronto, ON, M5T 1R8, Canada

          Background  Genomic copy number variants (CNVs) have identified genes associated with autism spectrum disorder (ASD) and intellectual disability (ID). Recently, we reported on 23 individuals in 17 families with loss CNVs or frame-shifting indels disrupting PTCHD1. However, little is known for the function of PTCHD1.
Objectives  PTCHD1, an X-linked gene for ASD and/or (ID), encodes a protein with homology to the sonic hedgehog (Shh) receptors PTCH1 and 2, and has been reported to show similar Gli repression activity. However, little else is known about the function of PTCHD1 or the pathway(s) that are perturbed by PTCHD1 mutations leading to ASD/ID.
Design/ Method We looked at the effect of over-expression of PTCHD1 on transcription levels of neuronal and synapse-related genes. We also included a truncated construct lacking the C-terminal four amino acids, Ile-Thr-Thr-Val (ITTV) of PTCHD1, which is predicted to interact with the PDZ domains of proteins. We tested the transcription of Shh and its putative receptor, Ptchd1, and Smoothened (Smo), from mouse embryonic and postnatal brains. We also analyzed the expression of PTCHD1 and Smo in primary cilia.
Results Transcriptional analysis showed Shh, Ptch1, and Smo expressed in developing mouse embryonic and postnatal hippocampus, but with gradually decreasing levels, whereas Ptchd1 levels steadily increase, suggesting that Ptchd1 may be more relevant in later embryonic and postnatal neurodevelopment. Preliminary results indicate that PTCHD1 is able to localize to the cilia.
A significant increase in level of transcription of NLGN1 and CNTNAP1 that is dependent on the C-terminal four amino acids of PTCHD1 suggests either a regulatory or a downstream effect of PTCHD1 on these genes via a PDZ-domain containing protein. Positive PTCHD1 immunolabeling was visible in the hippocampal neurons. Preliminary results also suggest localization of PTCHD1 in cilia.
Conclusions We report here a potential role for the PDZ binding domain in PTCHD1 in regulating expression of Nlgn1 and Cntnap1. PDZ domains are found in many eukaryotic and bacterial proteins, and in particular many proteins that are important for synaptic functioning.
We speculate that PTCHD1 localization to primary cilia in hippocampal neurons could inhibit the Hh pathway by excluding SMO, allowing cilia to function as Shh sensors, in a manner similar to that of PTCH1, during neurodevelopment.
The PTCHD1 gene has a role in a neurobiological pathway that delivers information to cells during brain development and CNVs and mutations may disrupt crucial developmental processes, contributing to the onset of autism.