Genomics in Mendelian Disorders: from the

Lab to the Clinic

CCCAGATGCCCTGTTCCAGGAGGACAGCTACAAGAAACACCTGAAGCATCACTGTAACAAGTATGTTATTAGAGGGTGGACCTGGAGAGCTTAATTCCCTTTTTATTCTTTAAAAAATACATGCAGCCGGCCCTTCACGTCTGCAGATGCAGAACTCGCAGATTTGGAGGGTCAACTGAGGGACCTGAGCATCTGCGGATCTTGGTGTCTGAGGGGGGTCCTGGAACCATACTCCCGCGGATATGGAGGGACAGCTCTGTTATTAAGACTTTTAAATGGTATAGTTATTGCCTTTGCACAGCCTTATCATTTTTCTTGAAATGTGGTGTCAAGTTGCAGGAGAGCGTACCTTTAGGTGACTGATTATTTTTTAACATGGTAAGATACACAACACAACGTTTACCATTTTTACCATTTATAAGTGAACAATTCATTGGCATTAATTACACTCACAATGCTGTATACTCACTATCTGTACCTGAAATGTTTCCATCTTCCCAAATATAAACACTGTATCAATTAAACA

CCCAGATGCCCTGTTCCAGGAGGACAGCTACAAGAAACACCTGAAGCATCACTGTAACAAGTATGTTATTAGAGGGTGGACCTGGAGAGCTTAATTCCCTTTTTATTCTTTAAAAAATACATGCAGCCGGCCCTTCACGTCTGCAGATGCAGAACTCGCAGATTTGGAGGGTCAACTGAGGGACCTGAGCATCTGCGGATCTTGGTGTCTGAGGGGGGTCCTGGAACCATACTCCCGCGGATATGGAGGGACAGCTCTGTTATTAAGACTTTTAAATGGTATAGTTATTGCCTTTGCACAGCCTTATCATTTTTCTTGAAATGTGGTGTCAAGTTGCAGGAGAGCGTACCTTTAGGTGACTGATTATTTTTTAACATGGTAAGATACACAACACAACGTTTACCATTTTTACCATTTATAAGTGAACAATTCATTGGCATTAATTACACTCACAATGCTGTATACTCACTATCTGTACCTGAAATGTTTCCATCTTCCCAAATATAAACACTGTATCAATTAAACA

Tony Roscioli FRACP PhD

Clinical Geneticist, Sydney Children’s HospitalVisiting Scientist, the Garvan Institute

Summary

• Next Generation Sequencing in Context– Some Australian initiatives with a major health impact– The identification of disease genes

• The Costs to NSW of Specific diseases and cost savings achievable through genomics– Drug reactions, inherited diseases with disability

• Families where genomics has provided the answer with associated cost savings

• Overall pilot results of genomic testing in genetic disorders

Penicillin was one of the most important discoveries during WWIIBy the end of the war almost 2 million lives had been saved

Penicillin

Barry Marshall and Garry Warren

Peptic Ulcers treatment with Ranitidine/Clarithromycin

$650 million annually saved in the US

Peptic Ulceration

Lithium treatment for bipolar disorder

$9 billion saved annually in the US (Kirschner et al 1994)

Bipolar Disorders

John Cade

Genomics is Unique in our Time

- WGS is allowing many diagnoses to be made at a population scale

- It sits in a historical context – more important than the microscopewhich created a revolution in understanding in microbiologyby allowing the invisible to be investigated

- We only see what is visible – WGS makes all genes accessible to study

•Exome = Protein Coding Genome

•split amongst >180,000 exons in humans

• total length ~30Mb

• includes canonical splice sites, miRNAs and 5’ and 3’ untranslated regions

•Estimated that this 1% of the genome contains 85% of human mutations

Technology and Plummeting Costs are Driving

Genomic Approaches

Moore’s Law: the Mandatory Genomic Talk Slide

Detecting disease causing mutations:

A good thing but too many tests prior to NGS

• Confirms / Excludes specific diagnosis

• Determines the patient therapy eligibility

• Enable the systematic design of therapies based on

molecular targets

• Accurate Genetic Counselling

• Recognition of new diseases / disease subgroups

CCCAGATGCCCTGTTCCAGGAGGACAGCTACAAGAAACACCTGAAGCATCACTGTAACAAGTATGTTATTAGAGGGTGGACCTGGAGAGCTTAATTCCCTTTTTATTCTTTAAAAAATACATGCAGCCGGCCCTTCACGTCTGCAGATGCAGAACTCGCAGATTTGGAGGGTCAACTGAGGGACCTGAGCATCTGCGGATCTTGGTGTCTGAGGGGGGTCCTGGAACCATACTCCCGCGGATATGGAGGGACAGCTCTGTTATTAAGACTTTTAAATGGTATAGTTATTGCCTTTGCACAGCCTTATCATTTTTCTTGAAATGTGGTGTCAAGTTGCAGGAGAGCGTACCTTTAGGTGACTGATTATTTTTTAACATGGTAAGATACACAACACAACGTTTACCATTTTTACCATTTATAAGTGAACAATTCATTGGCATTAATTACACTCACAATGCTGTATACTCACTATCTGTACCTGAAATGTTTCCATCTTCCCAAATATAAACACTGTATCAATTAAACA

CCCAGATGCCCTGTTCCAGGAGGACAGCTACAAGAAACACCTGAAGCATCACTGTAACAAGTATGTTATTAGAGGGTGGACCTGGAGAGCTTAATTCCCTTTTTATTCTTTAAAAAATACATGCAGCCGGCCCTTCACGTCTGCAGATGCAGAACTCGCAGATTTGGAGGGTCAACTGAGGGACCTGAGCATCTGCGGATCTTGGTGTCTGAGGGGGGTCCTGGAACCATACTCCCGCGGATATGGAGGGACAGCTCTGTTATTAAGACTTTTAAATGGTATAGTTATTGCCTTTGCACAGCCTTATCATTTTTCTTGAAATGTGGTGTCAAGTTGCAGGAGAGCGTACCTTTAGGTGACTGATTATTTTTTAACATGGTAAGATACACAACACAACGTTTACCATTTTTACCATTTATAAGTGAACAATTCATTGGCATTAATTACACTCACAATGCTGTATACTCACTATCTGTACCTGAAATGTTTCCATCTTCCCAAATATAAACACTGTATCAATTAAACA

“Last” Generation Sequencing

-Robust and Reliable but

-Labour intensive – one gene at a time

-Difficulties when wishing to test more than 3-4 genes

-Costs rise rapidly

-No diagnosis if gene not known or sequenced

Next Generation sequencing addressed these issues

But still important for the moment for confirmation and segregation

1st recessiveDisease gene identified by

Exome sequencing

1st recessiveDisease gene identified by

Genome sequencing

1st DominantDisease genes identified byExome/Genome sequencing

De novo mutations in Intellectual Disability

NeanderthalGenome

Rapid Advances in Genomic Studies: 4 years

2014WGS improves diagnosis

in MR by 15%Exon dosage

Mendelian DisordersRare (but not collectively) Monogenic DiseasesMany thousands solved, >2000 left

The Effects of Genomics since 2009: Rapid Gene IDs

Rapid falls in Impact Factors!

0

20

40

60

80

100

3/2

00

9

4/2

00

9

1/2

01

0

2/2

01

0

3/2

01

0

4/2

01

0

1/2

01

1

2/2

01

1

3/2

01

1

4/2

01

1

1/2

01

2

2/2

01

2

3/2

01

2

4/2

01

2

1/2

01

3

2/2

01

3

Mendelian Disease Genes Published per Quarter

We may have 5-8 years until the primaryallele for many recognized Mendelian disorders are identified

(assuming current rates of Gene discovery)

Mendelian gene discovery: Gene discovery is accelerating to 1 new Mendelian disease gene/day

0

10

20

30

40

50

60

70

80

90

3/2

00

9

4/2

00

9

1/2

01

0

2/2

01

0

3/2

01

0

4/2

01

0

1/2

01

1

2/2

01

1

3/2

01

1

4/2

01

1

1/2

01

2

2/2

01

2

3/2

01

2

4/2

01

2

1/2

01

3

2/2

01

3

Rare Diseases: A misnomer

• Rare diseases affect fewer than 5/10,000 people BUT

• Although individually rare, they represent the majority of paediatrichealth budgets and admissions collectively

• Often chronic and complex, requiring many different specialised health services and community support

• Resources required are disproportionate to their individual frequencies

>80% are genetic:

Often related to consanguinity

high recurrence risk

Amenable to improved genetic diagnosis through NGS

100s of families in Sydney South West (Liverpool/Bankstown)

Garvan-SCH cohort:

Mendelian Disorders and Clinical need

• 54 Families• Phenotype

– Intellectual disability (23): Non-syndromic (3)

– Epileptic encephalopathy (11)

– Skeletal (5)– Immune (4)– Syndromic (3)– Eye (3)– Haematological (3)– Neurological (seizures) (1)– Metabolic (1)

Confirmed diagnosis

• Single affected female• Non-Consanguineous family• MR plus Cornelia de Lange gestalt• Possible de novo inheritance• Multiple investigations, microarray normal

• Exome performed in proband

• Both CADD and RVIS/PROVEAN consistent with pathogencity for SMC1A mutation – phenotype crucial

MACF1

POU4F3

EEF2K

SMC1A

OGT-8

-6

-4

-2

0

2

0.00 0.20 0.40 0.60 0.80 1.00 1.20

RVIS vs. PROVEAN SCOREZSCAN20

SFMBT2EVPLSEPT2

ZMYD19

WDR85

SMC1A

0

5

10

15

20

25

30

35

40

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27

CADD score

SMC1AI: the diagnosis

• Missense mutation in X-linked gene

with predicted high pathogenicity

• SMC1A gene mutations in 5% of

individuals with a clinical diagnosis of

Cornelia de Lange

• Escapes X inactivation and so girls and

boys may be affected

• In silico pathogenicity predictions: probably damaging / deleterious

• Sequencing reads in IGV: unlikely to be a sequencing error

• Confimed as de novo by Sanger

sequencing

Epileptic encephalopathy: high diagnostic

rate in de novo families

Singletons n=2/11

• 2 pathogenic results confirmed from 11 patients after singleton sequencing. Trio sequencing in progress

• 1 de novo KCNQ2 mutation

• Compound heterozygous ASNS mutations – newly described MR, seizures

and microcephaly

Family trios n=5/6 families diagnosed

• 5 likely pathogenic findings from 6 trios.

• Previously reported de novo ALG13 (X linked CDG)

• Compound heterozygote ADSLmutations in girl with infantile spasms and movement disorder

• De novo SCN2A mutation

• De novo SCN1B mutation in child with infantile spasms (extend previously reported phenotype)

Other confirmed/In Progress diagnoses

• Single affected male MR and cerebral abnormalities

• Confirmed de novo GRIN2B mutation

• “We are so happy to know the cause and to know it

wasn’t our fault”

• Consanguineous family, MR and epilepsy

novel gene, Drosophila studies

in progress

• Non-consanguineous family, PID

de novo STAT3 mutation, confirmed

-Provide Diagnosis, accurate management

-Accurate recurrence risk and testing

-New Genes

Patient 1: an undiagnosed

immunodeficiency 11 year old girl

• Recurrent skin abscesses refractory to antibiotics

from birth

– Non-related parents of Indian background

• 2 well siblings

– Age 3 Left intra-orbital abscess – Multiple surgical drainage procedures

– Mycobacterium cultures

-Genetic susceptibility to mycobacteria?

- IFNg/ IL-12/ NFKB pathways

- Individual Genes sequenced at NIH

- no mutation identified

• Referred to SCH immunogenetics clinic

• Impressions: • ? AD HIGE – however no reports of mycobacterial infection

• ? Other Mendelian susceptibility to mycobacterial disease

– Gene not identified at NIH

– Strategy

• Whole exome sequencing organised

• Anticipated inheritance model

– Autosomal recessive

– De novo Dominant

Patient 1: NGS corrects a

diagnosis

Definitive Diagnosis

• Diagnosis – AD Hyper IgE syndrome

– De novo Heterozygous STAT3 mutation –

previously reported

• Previous IFNg treatment

– Estimated cost for 3-4 years total therapy

• Estimate c. 1400 vials x $226.82

• Total estimate (current price) c. $317,000 (IPU)

• Ceased as now known to be ineffective

because of the confirmed NGS diagnosis

Summary to Date from half of the cohort

Total: 30 families; 53 exomes

10/30 probably diagnosed (30%)

Of these, 2 genes potentially novel

Consistent with international practice:

WES leading to rapid diagnoses and gene discovery

Health Costs which could be

addressed through NGS at a great saving to the

health budget

Drug reactions

inherited diseases with disability

Pre-conception testing

Australian and NSW population

NSW = 7.5 million March 2014NSW = ~100,000 births/year 2011

Australia = 23,660,200 Nov 11 2014Australia = 308,065 births per year

Australia = 3.154X NSW population

Drug Reactions: an expensive affair

-Medication-related hospital admissions have been

estimated to be 2-3% of all Australian hospital admissions

-20%-30% of all admissions >65 years are medication-

related

-230,000 Medication-related hospital admission annually

-The average cost per each admission in 2011-12 is $5,204

-The annual cost of medication-related admissions is

$1.2 billion

Birth 60 years

Drug ReactionAverage Cost $5200

Birth

WGS

60 years

Flagged Genomic RecordAlternative Drug

Cost Saving $5200

Pharmacogenomics

Population WGS

A proportion of $380 MILLION: 10-20%??$30-50 million saved?

Avoided Costs to NSW Health System:

Pharmacogenomics

Results flaggedIn Childhood

Birth 40-60 years

Birth 30 – 60 years

A child with a Severe disability

The Costs to Families and Society of

Severe Genetic Conditions

- Costs of Medical/Surgical care- Assisted home living- Medications- Loss of employment opportunities for parents- Relationship breakdown- Loss of reproductive confidence with fewer future children

The Costs to Families and Society of

Severe Genetic Conditions

- Overall costs to families and carers for all aspects of daily life:$500,000/year

- Many of the causes will be new (“de novo”) mutations with a low chance of recurrence

- Some may be inherited with a high chance of recurrence

- Even is a small number of families who carry a gene mutationidentified by WGS make the choice to have a second healthy child with IVF, this will result in a significant economic impact

The Ongoing Psychological Burden:

Guilt and Fear can be removed by Genomics

- Professor Di Donnai: “Never underestimate the power of a diagnosis even if nothingcan be done”

- The majority of families will have had a child with a severeintellectual disability as a “one off” event – WGS will provide them with an answer and restore their reproductive confidence

- The knowledge that the disability was due to a new mutation with a very low chance of occurring again and that the family have not caused the birth of a child with intellectual disability is often enough to lift the burden of guilt

Traditional Diagnosis cost also significant• SMN1 molecular testing $690

• Myotonic dystrophy DNA test on mother $506

• Neurological appt for assessment mother $110

• Myasthenia Gravis DNA testing $2800

• 2 Micoarrays –both babies $1200

• 3 Pathologists opinion both babies incl UK $720

• Laminin A molecular testing $1000

• Postmortem $3000

• Muscle biopsy $144.05

Total $10,170 vs 2 Exomes $2-3000

• MRI brain $441.14

• Muscle biopsy $144.05

• Skin biopsy $50.25

• Surgical Session 4 h $2651.81

• Anaesthetist session 4h $1884.43

• Day Stay $1946

• ICU 24h non ventilated $326

• EM concord $380

• Dry ice to Melbourne $80

• Mito analysis $1400

• SNP arrays $1200

• Nerve conduction $200

Total $10,703 vs 2 Exomes $2-3000

Assuming 50% Exome Diagnostic Rate• Average per family all costs:~$10,000 ( a recurrent estimate over many studies)

• Average per family for 2 Exomes:~$2000

• Assuming would only get to a diagnosis half then time:– Still a saving of $6000 per family

• Assume that half of the costs for medical care are still required– Still an average saving of $3000 per family

– Much more money will be saved in some families and to the health system

Running 100 exomes per year could savehalf a million dollars per year in standard diagnostic costs

And these saving will become more pronounced as genomic costs fall

Population WGS

Parents who are both carriers couldChoose IVF to start a pregnancy knowing

the baby would not a recessive disease if they are at risk

Modulated Costs in the NSW Health System:

Pre-conception genomic testing

Results flaggedIn early adulthoodFor family planning

Birth 20-30 years

Potential Savings in NSW

• Pharmacogenomic testing and management could save $30-50 Million / year

• Confirming de novo mutations in intellectual disability would increase the likelihood that families would have more children – with all the positives that brings

• Genomic testing is going to be the short path to a specific diagnosis with savings of at least $3000 per family where there is a child with a significant genetic disorder

• The lifetime cost for all the care that a person with a severe intellectual disability requires could be ~$20-30 million

• Some families where there is a high risk of recurrence are already requesting IVF to ensure they can have a healthy child – with the secondary effect of shifting future costs to other parts of the health budget

Cost Savings and People Investment

• We are beginning to show that money can be saved for the hospital system

• It will however require Investment for infrastructure and also for staff – geneticists, genetic counselorsand community education

Key messages

• Genomics is the key medical diagnostic technology of our time

• The rollout of NGS technologies for community testing is occurring in NSW but will require adequate funding for genetic and genomic staff as well as ongoing resources to support testing

• The application of NGS to multiple health problems will save resources across the entire health budget which could represent a funding mechanism for NGS

• Collaboration between the KCCG/Garvan and genomicists within hospitals will result in good outcomes and high quality reports to clinicians and families

Thanks to the people at the Garvan and SCHN/SEALS

•Pathologists (Buckley/Mead/Burnett), Genome scientists

•Garvan/KCCG Genomics and Bioinformatics –(Mattick/Dinger/Cowley/Kaplan/Miller/Schofield)•Students: Lisa Ewans, Paul Gray

Hooi Ling TeeohEmma Palmer

•Immunology: John Ziegler, Paul Gray•Neurology: Annie Bye, Michelle FarrarMichael Cardamone, Hugo SampaioIan Andrews, John Lawson•Genetics: Anne Turner, David Mowat, Edwin KirkMary-Louise Freckmann, Michelle Lipke, Rani Sachdev•Genetic Counsellors: Carolyn Shalhoub, Bec MacintoshJacqui Robinson, Lisa Bristowe