An Update in Genetics of Epilepsy Dr Michelle Demos Pediatric Neurologist, Neurogeneticist BC Children’s Hospital

Outline Overview of Genetics Role of Genetics in Epilepsy Implications for Patients/Families Advances in Genetics of Epilepsy and Genetic Testing

Overview of Genetics What is Genetics? Study of heredity or how traits are passed from parent to child

Overview of Genetics Traits: Notable feature or characteristic in a person Each person has a different combination of traits which makes us unique

Physical Traits

Physical Traits

Examples of Traits: Height, body weight, health or disease Genetic and environmental contribution Environmental contribution

Examples of Traits: Height, body weight, health or disease Genetic and environmental contribution Environmental contribution

Gene Basic unit of heredity Made of lengths of DNA (deoxyribonucleic acid) Genes proteins normal body function

Human Genetic Make-up DNA ~ 20, 000 genes The entire collection of DNA is called your Genome

Human Chromosomes OR Female Male

Sex Chromosomes

DNA, genetic code and making of human protein for normal function of human body The most important component of chromosomes is DNA; make up of substances or bases called GCTA The order of these bases make up our genetic code, which provides info needed to make proteins, Important for proper structure and funciton of human body Proteins made up of amino acids; each 3 base unit of DNA determines amino acid; gene is unit containing Blueprint for making specific protein; about 20000 genes packed in each parental chromosome set (23)

Outline Overview of Genetics Role of Genetics in Epilepsy Implications for Patients/Families Advances in Genetics of Epilepsy and Genetic Testing

Twin Studies Vadlamudi et al. Neurology 2004. Lennox, JAMA 1951.

Family Studies Relatives of individuals with epilepsy more likely to develop epilepsy compared to general population

Linkage Analysis Early Epilepsy Gene Discovery

Types of Genetic Disorders in Epilepsy Single Gene Chromosome Complex Mitochondrial

Genetic Changes that can Predispose to Epilepsy: Single Gene

Single Gene Epilepsy Disorders caused by changes (mutations) in single genes recognizable inheritance pattern (dominant, recessive, X-linked) or arise by new mutation or de novo very few cause primary epilepsy syndromes Childhood Absence Epilepsy Juvenile Myoclonic Epilepsy many more will frequently have epilepsy as one of their symptoms

Features of the single gene epilepsy disorders same gene mutation may produce multiple different seizure types or severity not all individuals inheriting the affected epilepsy gene will develop seizures more than one gene can cause the same disorder

Types of Genetic Disorders in Epilepsy Single Gene Chromosome Complex Mitochondrial

Genetic Change that can Predispose to Epilepsy: Loss of part of a chromosome

Chromosome Disorders result in loss (deletion) or extra (duplication) chromosome content often new change in child so recurrence risk is low, but if present in parent then recurrence risk is 50% often associated with other medical problems

Types of Genetic Disorders in Epilepsy Single Gene Chromosome Complex Mitochondrial

Complex Disorders many epilepsies with a genetic basis Childhood Absence Epilepsy Juvenile Myoclonic Epilepsy interactions between multiple genes with or without environment pattern of inheritance less clear

Types of Genetic Disorders in Epilepsy Single Gene Chromosome Complex Mitochondrial

Mitochondria Mitochondria Chromosome Nucleus

Mitochondrial Disorders mitochondrial DNA is passed from mother to child every child of a mother carrying mitochondrial DNA mutation is at risk of being affected affects multiple organs within the body symptoms vary amongst individuals

Mitochondrial Disorders Affected Abnormal Mitochondria Normal Mitochondria Insert pedigree here

Question: If a parent or child has epilepsy, what is the risk to other family members? CAUSE

Cause is Known Acquired brain injury including stroke, brain tumor, brain infection or severe head trauma Risk is not significantly increased

Cause is Known Genetic Disorder (single gene, chromosome, mitochondrial) Risk will depend on genetic cause Remention chromosome and mitochondrial

Single Gene Disorders New dominant mutation recurrence risk is low – severe epilepsy disorders If not new, depends on inheritance pattern Dominant, recessive, X-linked Remention chromosome and mitochondrial

Dominant Inheritance: 50% risk

Recessive Inheritance: 25% risk

X-linked Inheritance: 50% risk to have an affected son

Cause Unknown Many of epilepsy syndromes with genetic basis (Childhood Absence Epilepsy) Complex inheritance Risk is an estimate based on past studies

Estimated Risks Risk in close relatives is about 2 to 4 times higher than people in general population Risk is less than 1 in 10 that a child of a person with epilepsy will also develop epilepsy Risk is higher in relatives of a person with generalized epilepsy, young onset epilepsy and mothers vs fathers with epilepsy

Outline Overview of Genetics Role of Genetics in Epilepsy Implications for Patients/Families Advances in Genetics of Epilepsy and Genetic Testing

Epilepsy Genetics and Genomics – Now Chromosome Microarray or Array Comparative Genomic Hybridization (CGH) Next-Generation Sequencing Complex etiology Genetic generalized epilepsies and Nonacquired focal epilepsies with multiple genetic and environmental Factors contributing to risk requires larger sample sizes to have sufficient power to Detect responsible genes. One approach is to perform large-scale-genome-wide linkage or Genome-wide association studies with large collections of families or cases and controls 1-Kasperavicuiute and colleagues 2010 >3000 individuals with focal epilepsies and almost 7000 controls; focal epilepsy any etiology; identified associations modest effect—none reached Genome-wide significance 2012: Han chinease Guo Y: 1087 focal epilepsy with acquired or unknown; Single SNP reached singificance on 1q32.1 CAMSAP1L1-calmodulin regulated Spectrin associated cytoskeletal protein

Methods: Array CGH Normal Control DNA Patient DNA DNA fragments Advances in genetics of epilepsy; technology allows identification of genomic imbalances at higher resolution compared to routine cytogenetic studies. DNA fragments representing entire genome are place on array. Genomic comparison is made by hybridization of differentially labeled test and reference DNA. relative intensity of test and reference sample fluorescence for each DNA fragment used to detect gains or losses. Location of that is then mapped back to the genome. Eg Agilent aberration detection method uses algorith to detect consistently high or low log2 ratios indicating del/dup. Patient DNA Normal Control DNA From http://www.utoronto.ca/cancyto

Examining chromosomes or genomes for genomic imbalances or Copy Number Variants (CNVs) causing Epilepsy ArrayCGH 1000X resolution Karyotype

Copy Number Variants Deletion or Copy Loss Duplication or Copy Gain >1000bp

Chromosome Microarray in Epilepsy: What have we found? A cause or abnormal CNV is identified in up to 15% of patients Treatment resistant epilepsy or other severe forms of epilepsy Epilepsy plus other neurological problem or birth defect A cause is identified less often in patients with benign epilepsy syndromes (Juvenile Myoclonic Epilepsy) Including BC 15q13, 15q11, 16p13

Chromosome Microarray in Epilepsy: What have we found? Limitations to testing: negative test does not rule out genetic cause Results of unclear significance Incidental findings: finding an abnormality in a gene predisposing to unrelated health problem (example breast cancer gene) Including BC 15q13, 15q11, 16p13

Chromosome Microarray in Epilepsy: Who should be Tested? Cause of Epilepsy is unknown and: a more severe form of epilepsy or epilepsy plus global delay, intellectual disability, autism, other neurological problem and/or birth defect Patient and/or Family who has received appropriate genetic counselling Including BC 15q13, 15q11, 16p13

Epilepsy Genetics and Genomics – Now Chromosome Microarray or Array Comparative Genomic Hybridization (CGH) and Copy Number Variants Next-Generation Sequencing Complex etiology Genetic generalized epilepsies and Nonacquired focal epilepsies with multiple genetic and environmental Factors contributing to risk requires larger sample sizes to have sufficient power to Detect responsible genes. One approach is to perform large-scale-genome-wide linkage or Genome-wide association studies with large collections of families or cases and controls 1-Kasperavicuiute and colleagues 2010 >3000 individuals with focal epilepsies and almost 7000 controls; focal epilepsy any etiology; identified associations modest effect—none reached Genome-wide significance 2012: Han chinease Guo Y: 1087 focal epilepsy with acquired or unknown; Single SNP reached singificance on 1q32.1 CAMSAP1L1-calmodulin regulated Spectrin associated cytoskeletal protein

Examining genes, exomes and genomes for sequence variants causing Epilepsy Sanger sequencing Next-generation sequencing

Next-generation sequencing Massively parallel sequencing Sequence millions of fragments simultaneously Whole genome, exome (protein coding), targeted or gene panels Massive parallel sequencing as many genes as desired Sequence quality depends on minimum coverage at a base-pair position and optimization of enrichment procedures of the genes of interest (target sequence)

Whole genome, exome or targeted sequencing in Epilepsy Targeted= 10 to 500 epilepsy genes Exome = 1 %

Whole Exome sequencing Single gene disorders 20 000 variants per individual Exome contains 85% of disease-causing mutations

Next Generation Sequencing: What is the impact? Rapid increase in Gene Discovery

Next Generation Sequencing: Epilepsy Impact ~ 500 genes associated with epilepsy Significant increase in diagnoses: Diagnosis 10 – 50% of those tested Severe epilepsy conditions or Epilepsy plus New dominant mutations in multiple different genes

Next Generation Sequencing: Challenges Negative test does not rule out genetic cause Variants of unclear significance Proving new or novel gene identified is the cause of epilepsy Availability in Canada Ethical Issues Cost

Whole Genome Sequencing

Next Generation Sequencing: Positive Impact on Patients/Families Earlier diagnosis including disorders with specific treatment implications More accurate counselling regarding outcome and recurrence risk Prevent additional investigations A clear genetic diagnosis improves access to therapies/community support

Next Generation Sequencing: Negative Impact on Patients/Families Identifying a disease without specific treatment Discrimination regarding life, disability or long-term care insurance Incidental findings (finding an abnormality in a gene that increase risk for developing cancer) Genetic Counselling Before Testing

Genetic Testing in Patients/families with Epilepsy Chromosome Microarray Analysis: Available in BC Indicated in patients with epilepsy of unknown cause and: Severe forms of epilepsy Epilepsy plus delay, intellectual disability, autism, other neurological/behavioral problems and/or other congenital abnormalities Pre-test genetic counselling required

Genetic Testing in Patients/families with Epilepsy Next Generation Sequencing: Limited Availability in BC Research, MSP funding Consider in patients with negative chromosome microarray and: Severe forms of epilepsy Epilepsy Plus Early onset epilepsy Pre-test genetic counselling required Gene panels or whole exome sequencing

Future Directions Standardized approach to genetic testing Genetic discoveries leading to development of better treatment options

Future Directions Pharmacogenomics of Epilepsy: The study of the interaction of an individual's genetic makeup and response to a drug. Adverse drug reactions (HLA-B*1502) Seizure control Aim to individualize and optimize treatment based on genetic make-up Hla b1502 tegretol induced Han Chinese

Summary Genetic factors play a role in many epilepsy disorders New technology (Chromosome Microarray and Next-Generation Sequencing) has revolutionized gene discovery and genetic testing in epilepsy Recent discoveries and further advances in diagnosis and pharmacogenomics will likely lead to improved individualized treatment of epilepsy

Questions?

Genetic Contribution to Epilepsy Complex Number with Epilepsy Doesn’t address many genetic factors involved in symptomatic epilepsies Genetic factors likely involved most epilepsies; impact and heredity is variable This is important consideration when considering genetic testing Genetic factors likely play a role in most epilepsies but genetic impact is variable and have no role in recurrence risk ie low heritability but impact on seizure predisposition after brain injury And response to medication. Genetic (high) Acquired (low) Genetic Contribution to Epilepsy Helbig I, et al, 2008. 67