Cutting Edge Genetics Made Research funding, Natera Easy - - PowerPoint PPT Presentation

6/7/2018 1

Cutting Edge Genetics Made Easy

Mary E Norton, MD Department of Obstetrics, Gynecology, and Reproductive Sciences University of California, San Francisco

Disclosures

• Research funding, Natera
• Consultant and advisory board member, Invitae

Outline

• Brief history
• What’s here now (and what you need to know about

it)

• Cell free DNA, microarray, expanded carrier screening
• What’s coming (and what you will need to know soon)
• Microdeletion and single gene disorders by cell free DNA
• Whole exome sequencing

Screening Tests

• Cell free DNA screening
• First trimester screening
• Quad screening
• Carrier screening (ethnic, panethnic, or expanded)

Diagnostic Tests

• Karyotype
• Chromosomal microarray

Side Dishes

• FISH (fluorescence in situ hybridization
• NT ultrasound
• First trimester anatomy scan
• Fetal MRI

Coming soon….

• Whole exome or genome sequencing
• In utero therapies

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20 40 60 80 100 120 Age >35 Triple screen Quad screen First trimester screen Integrated screen cfDNA

Detection rate of prenatal screening for Down syndrome has improved over time

Detection Rate (%)

The history of prenatal diagnosis is focused largely

• n Down

syndrome Lejeune, 1959 Tijo and Levan, 1956

Newborn vs prenatal screening

Disorder is important  test is developed and introduced Technology is developed  test is introduced

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1997 DR: 99.2% (98.5 - 99.6)

cfDNA screening for T21: meta-analysis

(Gil et al, Ultrasound Obstet Gynecol, 2015

FPR: 0.09% (0.05 - 0.14) Detection Rate: 99.7% False Positive Rate: 1/2500

cfDNA screening for T21: meta-analysis

(Gil et al, Ultrasound Obstet Gynecol, 2017)

DOWN SYNDROME!!

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Spectrum of Congenital Disease

Structural Malformations

Autosomal recessive Autosomal dominant X-linked Chromosomal/ karyotype

Conditions found by microarray

• 1/300

pregnancies

• 20% of infant

deaths

Half of these are Down syndrome

Copy number variants

Which of the following conditions are NOT detected by a chromosomal microarray?

• A. Down syndrome
• B. 22q microdeletion syndrome
• C. Klinefelter syndrome
• D. Hemophilia
• E. What is a chromosomal microarray?

D

• w

n s y n d r

• m

e 2 2 q m i c r

• d

e l e t i

• n

s y n d r

• m

e K l i n e f e l t e r s y n d r

• m

e H e m

• p

h i l i a W h a t i s a c h r

• m
• s
• m

a l m i c . . .

8% 7% 16% 55% 14%

Microdeletions are genomic imbalances detected by microarray but not karyotype

Miller et al, 2010, AJHG

Microdeletions are detected by chromosomal microarrays (CMA)

• “Lab-on-a-chip”
• Detects thousands of

microdeletions and duplications at the same time

• Have largely replaced

karyotype in pediatrics

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Russell Silver syndrome: Small, asymmetric Mild learning disabilities Prader-Willi: Hypotonia, significant learning disabilities,

• besity

Angelman syndrome: Severe intellectual disability and speech impairment

Some Microdeletion Syndromes Ch 7 Ch 15 Ch 5

Cri du chat syndrome: Poor feeding and growth, microcephaly, severe learning difficulties

Chromosomal Microarray (CMA) for Prenatal Diagnosis

Indication for Testing Clinically Relevant (N=96) U/S Anomaly (N=755) 6.0% AMA (N=1,966) 1.7% Positive Screen (N=729) 1.7% Other (N=372) 1.3%

Rate of abnormalities by maternal age

Increasing maternal age 

(Microarray abnormalities)

• “In patients with fetal structural abnormalities undergoing

prenatal diagnosis, microarray is recommended.”

• Microarray should be available to all pregnant women.
• Patient acceptance limited by concern about uncertain findings,

fear of diagnostic testing, and lack of adequate insurance coverage

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Abnormalities found by microarray 1-6% Chromosomal microarray: bottom line

• Diagnostic testing with chromosomal

microarray is the best test for pregnant women who want as much information as possible about their fetus.

• Women should be counseled about the chance
• f finding a variant of uncertain significance
• Cell free DNA can detect only a tiny number of

the total microdeletions

Availability of Genetic Tests

Most of these are for inherited disorders

Ethnicity Based Carrier Screening

Ashkenazi Jews Tay Sachs disease, Canavan disease, cystic fibrosis, familial dysautonomia Louisiana Cajun, Tay Sachs disease Fr Canadian Caucasians Cystic fibrosis Africans, African Sickle cell anemia, beta Americans thalassemia Southeast Asians Alpha thalassemia Mediterraneans Beta thalassemia

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Traditional Carrier Screening

• Focus on ancestry and family history
• Small number of diseases
• High frequency in a certain subpopulation
• Severe morbidity or mortality
• Fetal, neonatal or early childhood onset
• Well-defined phenotype

Sickle Cell Tay-Sachs disease

Multiplex Panel Screening: Universal (Expanded) Carrier Screening

• Multiplex screening now allows carrier testing for many

(hundreds) disorders at once

• This is relatively inexpensive ($100-350)
• Should it be recommended for everyone?

What criteria are required by laboratories before including gene variants on panels?

What is on expanded panels and how are disorders chosen?

Disorders should be:

• Severe
• Common
• Have a well-described natural history and phenotype
• Have a high detection rate

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Expanded Carrier Screening: The Wild West

• Evaluated commercially available panels
• 27% of included disorders meet criteria per ACMG and

ACOG

ACOG 2017 Updated Screening Recommendations

• Screening should be offered to all women before or during

pregnancy for:

• Cystic fibrosis
• Spinal muscular atrophy
• MCV should be offered to all women who are currently

pregnant

• To those at risk for hemoglobinopathies, Hb electrophoresis

should be offered (African, Mediterranean, Middle Eastern, SE Asian, West Indian) or if MCV is low

ACOG 2017 Updated Screening Recommendations

• Fragile X screening should be offered to all women with:
• a family history of FraX related disorders
• unexplained ovarian insufficiency or failure
• Tay Sachs screening should be offered to those who are:
• French Canadian
• Cajun
• Ashkenazi Jewish

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Is expanded carrier screening accurate?

Molecular DNA testing is not the recommended screening approach for certain conditions

• Tay-Sachs disease
• Hexosaminidase A enzyme analysis is best test for non-

Jewish individuals

• Hemoglobinopathies
• Gold standard is MCV and hemoglobin electrophoresis
• Carrier screening platforms do not include all mutations

for alpha and beta thalassemia

The Question: The Answer: Not always!! How Often Do Tests Find Something?

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What Then?

24-45% will have something

• 1. Explain to the patient
• 2. Test the partner (he might not have insurance)
• 3. He will often be a carrier for something else
• 4. If low detection rate on original panel, do

gene sequencing

• 5. Explain all this to the patient

A real patient story

• Patient reports that she carries SMA
• Partner has expanded carrier screening through panel covered

by his insurance

• Carries Fanconi Anemia, group A
• Patient undergoes expanded carrier screening with panel

covered by her insurance

• She carries Pompe disease but was not tested for Fanconi

group A, just group C (updated panel)

• They are frustrated and seek a second opinion
• He undergoes gene sequencing for this condition
• All he really needed was testing for SMA

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What should you do?

It is acceptable to offer either:

• 1. Ethnicity based screening
• 2. Pan ethnic screening (offer everyone CF, SMA, and

assess MCV, regardless of ancestry)

• 3. Expanded carrier screening
• Offer everyone the same thing
• Consider creating a custom panel

Non-Invasive Single Gene Tests

• Maternal and fetal cell free DNA cannot be easily

distinguished

• However, can identify de novo or paternal gene mutation
• This includes blood type if mother is Rh negative and fetus is

Rh positive

37 yo G1P0 at 27 wks

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• N=47 cases
• Correct in 46 (96.2%)
• Useful tool in 3rd trimester to distinguish IUGR from

achondroplasia

Chitty LS, et al. Prenat Diagn 35:656, 2015

• De novo mutations
• Many associated with

advanced paternal age

• No prospective data on

performance

• Not ready for routine use

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What is the future?

Whole Genome and Whole Exome Sequencing

• Whole Genome Sequencing
• Obtaining the complete sequence of all 3 billion base pairs of DNA

in any individual

• Whole Exome Sequencing
• Obtaining the complete sequence of the ~2% of the genome

containing the exons that encode proteins

What is whole exome sequencing?

whole exome sequencing (WES), is a genomic technique for sequencing all of the protein- coding genes in a genome (known as the exome).

A genome is like watching the game from beginning to end An exome is like reading the highlights

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The future: Noninvasive whole genome sequencing?

What is “CRISPR”? What is “CRISPR”?

• Clustered
• Regularly
• Interspaced
• Short
• Palindromic
• Repeats

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CRISPR 101

Gene editing: fixing the errors

Clinical applications

• Edit pig DNA so organs can be transplanted into humans
• Inject CRISPR-edited WBCs into humans to treat cancer
• Treat illnesses such as cystic fibrosis, sickle-cell anemia and

Huntington's disease

• Modify and repair embryos
• Technical details aside,

Crispr-Cas9 makes it easy, cheap, and fast to move genes around—any genes, in any living thing, from bacteria to people. –David Baltimore

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What should I do?

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What should I do?

• Explain difference between diagnostic testing and

screening

• Ask patients if they “need to know as much as possible” or are

OK figuring out if they are high risk or low risk

• Offer aneuploidy screening to everyone
• Explain difference between traditional (broad, general screen for

lots of things) and cfDNA (very targeted)

• Offer aneuploidy cfDNA, not microdeletions
• Offer carrier screening to everyone
• Choose one offering and stick with it for all patients
• Know your lab!
• Be sure you have a plan for positive results

Thank you!