The University of Adelaide

X- and Epilepsy.

Jozef Gecz, PhD

1 Pavia, November 17, 2018 Dung Hoang Art Males & Females: Alike OR Different?

XY XX

Marlina Vera, 2017

2 Males & Females: Alike OR Different?

XY XX

Marlina Vera, 2017

3 Males & Females: Alike OR Different?

Science 11/2014

Marlina Vera, 2017

4 Males & Females: Alike OR Different?

Epilepsy

IGE FCD Photo’ty Heterot

Marlina Vera, 2017 The Scientist, October 2015

5 Males & Females: Alike OR Different?

Cardiovascular

Asthma

Marlina Vera, 2017 Ober et al NRG 2008

6 Males & Females: Alike OR Different?

13% penetrance 10% penetrance

Prader-Willi/Angelman

Rubinstein-Taybi

Smith-Magenis

Marlina Vera, 2017 N Engl J Med. 2012 Oct 4;367(14):1321-31

7 Males & Females: Alike OR Different?

…” Our findings show that females systematically carry more neurodevelopmentally deleterious variants than do males. This is true whether individuals (1) are ascertained for NDDs or (2) are parents of a proband referred for those symptoms.” …

A Higher Mutational Burden in Females Supports a ‘‘Female Protective Model’’ in Neurodevelopmental Disorders.

Jacquemont et al. AJHG 94, 415–425, 2014

Marlina Vera, 2017

8 Male Female Differences

 Hormones?

(= )?

 Environment/epigenetics?

 Combination of above?

9 Male Female Differences

 Hormones?

 Chromosomes (= genes)?

 Environment/epigenetics?

 Combination of above?

10 X-chromosome genes can directly affect brain Carruth et al. Nature Neurosci. 5, 933-934, 2002

XY Sry XX

XY Sry XX Sry XY XX

Dopamine Neur’s Dopamine Neur’s E14.5 – mesencephalon cell (dopamine neuron) for XY vs All Neur’s for XY cultures (before testosterone)

11 ESR differences in male and female brain.

Neurite outgrowth promoting effect of 17-β estradiol.

Pooley et al, Brain Res 2015 P2 olfactory neurons, DIV8

12 Male Female Differences

 Hormones?

 Chromosomes (= genes)?

 Environment/epigenetics?

 Combination of above?

13

• 5% of the genome

• 3% of the exome

• 846 coding genes

• 621 non-coding genes

• 892 pseudogenes

• 8-10% of ID May 25, 2002

14 X-chromosome NDD identification

pter SMS 1994 GK DMD OTC MAOA, NDP • ~15 genes

PGK1, ATP7A

PLP • 100s of unresolved families TIMM8A

OCRL HRPT

FMR1 IDS L1CAM qter

15 X-chromosome NDD gene identification

pter 2018

• ~164 genes (as of 13/11/2018) • no obvious function enrichment • 1 gene = multiple disorders • 50+ unresolved families

• females (with de novo mutations) • X does not explain the sex bias in NDDs or in general

qter

16 X-chromosome & epilepsy

pter 2012

~20 % of individuals with severe ID have seizures

~10% of individuals with mild ID have seizures

~ 50% of ~ 150 XLID syndromes (102 genes) with seizures

‘Seizures likely do not cause ID, both are a consequence of disturbed neurodevelopment.’ qter

17 X-chromosome & epilepsy pter 2018

~ 164 XLID genes i.e. 20% of X-chr coding genes are ID genes

~ 99 (60%) XLID genes with seizures (45/’12 – 54/’18)

• seizures are not always described for all patients/gene or not described/looked at all

• multiple animal models available and several with seizures or reduced threshold for seizures

• several XLID genes are ‘jumping’ the boundary, i.e. No seizures or Yes (e.g. IQSEC2) qter

18 Identification of IQSEC2 as an XLID gene

2010 Cheryl Shoubridge

Functional validation 4 unique non-synonymous SNPs in IQSEC2 in 4 separate families • No termination mutations • Missense mutations in functional domain

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t s Sec 7-Wt R758Q Q801P R863W E849A

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Mutations lead to diminished GTP binding to ARF6 compared to wild-type protein

19 Golden Age of Disease Gene Discovery 1986-2016

Next Generation >4 300 genes Sequencing >5 982 diseases

Human Genome

PCR

The American Journal of Human Genetics 100, 695–705, 2017

20 Novel Gene-Phenotype Discoveries 2010-2015

Less Bias!

HGMD database: http://www.hgmd.cf.ac.uk/ac/index.php

ClinVar: https://www.ncbi.nlm.nih.gov/clinvar/

Trait 1 = Epilepsy Trait 2 = CP Trait 3 = ID Trait 4 = Autism

The American Journal of Human Genetics 100, 695–705, 2017

21 IQSEC2: XLID & Epileptic Encephalopathy

2010 2013 ------2018 Cheryl Shoubridge Affected females # of families Phenotype Males Females cases (often limited information) ID ID + Seizures Speech ASD / Seizures Speech ASD / Seizures deficits features deficits features Familial 14 7 7 49 15 10 14 22 6 7 2 30% 20% 28% 27% 22% 9% De Novo 65 12 53 32 29 28 11 33 24 22 12 90% 87% 34% 72% 66% 36% 79 19 60 81 47 38 25 55 30 29 14 58% 46% 30% 54% 52% 25%

~136 patients directly affected by pathogenic variants in IQSEC2

+ Microdeletions / microduplications / whole gene duplications

22 Current Ge-Phe landscape of IQSEC2

Mutation Type Inheritance Cheryl Shoubridge

Phenotype in heterozygous females is Nonsense De Novo Familial similar to hemizygous males Missense Male Male Splice Female Female

Phenotypes ID + Seizures

ID -/+ Seizures

XLID ` Tolerated variation CC IQ-like Sec7 PH PDZ Allele Frequency Hemizygous males

0.01 to 0.05 N>2 <1/10,000 to 0.001 N=1 N=0 Singleton

Shoubridge, Harvey, Dudding, Hum Mut, 2018

23 PCDH19 – Cell adhesion, axon bundling pter

CDKL5 Dibbens et al. Nat Genet, 2008 Families (EFMR) ARX USP9X & DDX3X singletons (DEE) IQSEC2

PCDH19 Variable penetrance

Depienne et al Plos Genet, 2009

mut + + mut /+ PHF6 + +/+ + +/+ HCFC1 MECP2 + mut qter +/del + +/mut +/mut +/

24 Male Females Differences

 Hormones?

 Chromosomes (= genes)?

 Environment/epigenetics?

 Combination of above?

25 Identical DNA = different outcome! PCDH19 Girls Clustering Epilepsy

80-95%

Cognitive outcomes 70%

20% XCI 65%

5-20%

26 Identical DNA = different outcome! PCDH19 Girls Clustering Epilepsy

PCDH19: p.Asn340Ser (c.1019A>G) Kristy Kolc Age Seizure onset Seizure offset Language Psychiatric ID Reference Seizure type (semiology) Inheritance ID (years) (months) (years) Delay Comorbidity 32 Depienne 2009 3 9 Continuing FS, G (TC) De novo Yes Mild None 33 Depienne 2009 6 8 Continuing P De novo Yes Mild ADHD 71* Dibbens 2011 6 13 ? (TC) Maternal - Moderate ASD 72* Dibbens 2011 3 17 ? (TC) Maternal Yes Severe Unknown 73* Dibbens 2011 - 12 14 ? De novo - Normal None 85 Higurashi 2012 5 13 Continuing FS, F De novo - Normal None 86 Higurashi 2012 5 25 2 AS, G (TC) De novo - Normal None 93 Higurashi 2013 8 5 Continuing FS, F (T) De novo - Severe None 100 Higurashi 2013 5 8 Continuing FS, AS, F (TC) Maternal Yes Severe Multimorbidity 116 Liu 2017 9 7 Continuing FS, F (GTC) De novo - Yes Multimorbidity 117 Liu 2017 3 8 Continuing FS, F (GTC, My) Maternal - Yes Multimorbidity 131 Liu 2017 7 18 Continuing FS (GTC) De novo - Normal None 138 Marini 2010 9 10 Continuing FS, F (TC, H) De novo - Mild None 146* Marini 2010 15 6 Continuing FS, F (TC, SE) Maternal - Mild None 147* Marini 2010 44 8 17 (TC) ? - Normal None 153 Marini 2012 5 10 Continuing? FS (motor or hypomotor) De novo - Normal Other 164 Marini 2012 16 6 Continuing? FS (hypomotor) Maternal - Mild None 170 Marini 2012 11 10 Continuing? FS (motor) De novo - Moderate None 182 Marini 2012 9 5 Continuing? FS (hypomotor) De novo - Moderate ASD 184 Marini 2012 8 12 Continuing? FS (motor) Maternal - Normal None 218 Specchio 2011 7 12 Continuing FS, F, SG De novo - Normal None 221* Terraciano 2012* 8 11 5 FS (CJ) Maternal No Normal None 222* Terraciano 2012* ? N/A N/A NP Unknown No Normal None Yes? None 226** Terraciano 2016 3 ? ? FS De novo -

238 Van Harssel 2013 6 15 Continuing FS (H) Unknown - Moderate Multimorbidity

27 PCDH19 GCE - Review

271 cases reviewed (duplicates removed) Kristy Kolc

Kolc et al., Mol Psych, 2018

28 Pcdh19 KO mice - II (Taconic, P Thomas).

Daniel Pederick

Wild type Pcdh19

Mutant Pcdh19

Pcdh19HA-FLAG/-

E14.5 brain D Pederick et al, Neuron 2018

29 Pcdh19 KO mice - II (Taconic, P Thomas).

Daniel Pederick Affected

Unaffected

Variable Expressivity

due to Xi?

E14.5 brain – different females D Pederick et al, Neuron 2018

30 Pcdh19 KO mice - II (Taconic, P Thomas).

Daniel Pederick

PCDH19-GCE Discordant MZ twins

E14.5 brain – different females D Pederick et al, Neuron 2018

31 Discordant MZ twin research, lesson from PCDH19 GCE

wt/wt wt/wt wt/wt wt/wt

wt/wt wt/mut wt/mut wt/mut

e.g. SCN1A e.g. PCDH19 NEJM 363;14, 2010

Discordant twins = Discordant variant Discordant twins = Identical variant

32 Neurodevelopmental disabilities ~1 in 20 individuals (life-time, not all genetic) ‘Splitting’ Group of disorders in which the development OTHR ID ASD of the central nervous system is disturbed.

EPI CP BEH • Intellectual disabilities • Autisms • Epilepsies LNG HEAR VIS • Movement disorders (CP) • Mood & Behavior • Speech & Language

33 Neurodevelopmental disabilities ~1 in 20 individuals (life-time, not all genetic) ‘Lumping’ Group of disorders in which the development of the central nervous system is disturbed.

• Intellectual disabilities NDDs • Autisms • Epilepsies • Movement disorders (CP) • Mood & Behavior • Speech & Language

34 Neurodevelopmental disabilities ~1 in 20 individuals (life-time, not all genetic) ‘Lumping’

Trait 1 = Epilepsy

Trait 2 = Mov NDDs Trait 3 = ID Trait 4 = Autism

35 Neurodevelopmental disabilities ~1 in 20 individuals (life-time, not all genetic) ‘Lumping’

NDDs

36 Neurodevelopmental disabilities ~1 in 20 individuals (life-time, not all genetic) Common Features • Excess of Males

• Female Protective Model

• Highly Heterogeneous • 1000s of genes NDDs • Variable Clinical Expressivity • Highly Co-morbid

• Increased Cancer Risk

Must be appreciated when considering intervention.

37 X-chromosome & epilepsy

pter

• ~60% (99/164) XLID genes with seizures

• X-chromosome mutations frequently found in singleton females with seizures

• X-chr. inactivation (epigenetic silencing) is a (major) disease modifier

• ID & epilepsy are likely independent consequences of disturbed development

qter

38 Acknowledgements

NEUROgenetics at UofA Paul Thomas

Cheryl Shoubridge

Major collaborators: Funders:

Marini

39