should we be offering non-invasive prenatal screening for ... · should we be offering non-invasive...
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Should we be offering Non-Invasive Prenatal Screening for Chromosome Abnormalities in ALL pregnancies?
Marta Martha Dudek, MS LCGC Genetic Counselor
Senior Associate Obstetrics and Gynecology
Division of Maternal-Fetal Medicine
Vanderbilt School of Medicine 1
[email protected] #marthadudek
Notice of Disclosure I have no relevant financial relationships with
commercial interests to disclose. The views and opinions expressed here are my own.
They are not necessarily those of Vanderbilt University Medical Center and they may not be used for advertising or product endorsement purposes.
40th Annual High Risk Obstetrics Seminar 2
Objectives
• Review of traditional prenatal maternal serum screening and update on cell free DNA (cfDNA) for chromosomes abnormalities (CA) in pregnancy.
• Present a protocol for cfDNA screening for the average risk population for CA in pregnancy.
• Discuss how to manage high risk cfDNA/ non-invasive prenatal screening (NIPS) results.
• Discussion of how to choose a laboratory for cfDNA/NIPS for CA in pregnancy.
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0
20
40
60
80
100
20 25 30 35 40 45
Down syn. likelihood atdelivery
4 Maternal Age
% 48 yrs ~6.3% 1 in 16 women
35 yrs ~0.3% 1 in 365 women
40 yrs ~1% 1 in 109 women
45 yrs ~ 3% 1 in 32 women 25 yrs ~ 0.08%
1 in 1200 women
30 yrs ~0.1% 1 in 885 women
Ultrasound Detection rate is operator and patient/fetus dependent Minimal risk
Serum screening 1st &/or 2nd (combined or sequential)
5% False positive No risk of loss 80-90% detection
Invasive testing CVS ~0.5-1% SAB Amnio 1/200-1/500 SAB >99% detection
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Traditional prenatal screening/testing for Trisomy
Evolution of Trisomy Risk Assessment
NIPS using cfDNA
ACOG Practice Bulletin No. 77. Obstet Gynecol 2007;109:217-27.
FPR=5.0% FPR<0.1% Detection rate for Down syndrome
Cell-free DNA
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Cell-free DNA in Maternal Blood • Cell-free DNA (cfDNA) are short DNA fragments • In pregnancy, cfDNA from both the mom and fetus are in maternal blood • Amount of fetal cfDNA present is a small fraction of the maternal cfDNA
Non-Invasive Prenatal Screening (NIPS)
• Blood draw > 10 weeks
gestation • Initially offered for patients
considered at “high risk” for fetal chromosomal aneuploidy
• Turn around time 3-14 days • Cost
• List price $795-$2700 • Patient out of pocket cost
dependent on insurance
NIPS Products by Assay
12
Massively Parallel Shotgun
Sequencing (MPSS)
Targeted Sequencing
Sequenom MaterniT21TM
Verinata Verifi®
Ariosa
HarmonyTM
Targeted Sequencing
Natera
PanoramaTM
COUNTING SNPs
October 2011
February 2011
December 2012
May 2012
Illumina LabCorp
October 2014
Massively Parallel Shotgun Sequencing
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GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10 GACACGGTGGAGCTCGGCCACACCAGGCCCAGCTGG chr14 GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10 ACAGTGGTGGGGCCCATCCCTGGGTGAGGCTCAGTT chr21 GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10 GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10
Counting Method
GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10 TCCGCCCAGGCCATGAGGGACCTGGAAATGGCTGAT chr21 GACACGGTGGAGCTCGGCCACACCAGGCCCAGCTGG chr14 GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10 ACAGTGGTGGGGCCCATCCCTGGGTGAGGCTCAGTT chr21 GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10 GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10 GACACGGTGGAGCTCGGCCACACCAGGCCCAGCTGG chr14 GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10
Sequencing tells you which chromosome the ccf fragment comes from
TCCGCCCAGGCCATGAGGGACCTGGAAATGGCTGAT chr21
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 X Y
14
21 18 13
Mom’s Blood Mom’s DNA
1
2
3
1
2
3
1
2
3
Trisomy 21 (Down syndrome)
Fetal Fraction
Expected ratio for Trisomy
4% 1.02
10% 1.05
20% 1.10
40% 1.20
Counting Method
MPSS vs. Directed
Chr 21, 18, 13, X, Y cfDNA Other Chr cfDNA Unmapped cfDNA
cfDNA in blood
Massively Parallel Shotgun Sequencing
(MPSS) Directed analysis
GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10 GACACGGTGGAGCTCGGCCACACCAGGCCCAGCTGG chr14 GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10 ACAGTGGTGGGGCCCATCCCTGGGTGAGGCTCAGTT chr21 GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10 GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10
Counting Method
GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10 TCCGCCCAGGCCATGAGGGACCTGGAAATGGCTGAT chr21 GACACGGTGGAGCTCGGCCACACCAGGCCCAGCTGG chr14 GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10 ACAGTGGTGGGGCCCATCCCTGGGTGAGGCTCAGTT chr21 GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10 GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10 GACACGGTGGAGCTCGGCCACACCAGGCCCAGCTGG chr14 GGCCCTGGGGACAGTCTCCAATCCACTGAGTCATCT chr10
Directed or Targeting sequencing of Reference chromosomes and chromosomes of interest
TCCGCCCAGGCCATGAGGGACCTGGAAATGGCTGAT chr21
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 X Y
17
SNP = Single Nucleotide Polymorphism
• Polymorphisms occur when a single base pair (nucleotide) is changed: A, T, C, or G
• These are normal genetic changes that occur in every person and mark where people differ from one another
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SNP patterns
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Buffy coat contains mainly maternal DNA
plasma mixture of maternal and fetal DNA
Buffy coat = Maternal DNA
Plasma = Maternal + Fetal
DNA SNP
Sequencing
SNP Sequencing
Maternal Genotype
Maternal + Fetal Genotype
Fetal Genotype
19K+ SNPs are simultaneously targeted
Maternal blood
Algorithm to predict the most probable fetal genotype
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40th Annual High Risk Obstetrics Seminar NIPS for ALL 21
How accurate is NIPS?
Modified from Rebecca Carter’s presentation at PLIDA 2014
Maternal age
QUAD screen
First trimester screen
NIPS/ cfDNA
ACCURACY
Am
nio C
VS
The technology is fabulous!
40th Annual High Risk Obstetrics Seminar 22
NEJ MED 369; 6 8/8/13 p499
NIPS Sensitivity
(true positive of test )
Specificity
(true negative of test)
Positive predictive value
PPV (varies with disease
incidence)
Trisomy 21 99-100% 99.8-100% ?
Trisomy 13 80-100% 99-100% ?
Trisomy 18 97-100% 99.6-100% ?
Sex chromosome abnormalities
96%-100 99.7-100% ?
Source: 2009. Provider handbook for The California Prenatal Screening Program
Should average risk patients be offered
NIPS?
23
Majority of babies born with Down
syndrome are born to women LESS than
35 ys of age
Update on NIPS
• Microdeletion panels (Panorama & MaterniT21PLUS) • Change in practices regarding fetal fraction reporting • Sex chromosome abnormality reporting • Ability to report in multiples • Addition of other labs using Illumina platform and
Verifi algorithm • Validation in the average risk population
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Our Current NIPS Protocol
• Offered only to high risk patients • All patients undergo genetic counseling* • NIPS is presented as a screen within the
context of all other screening and diagnostic options
• Written informed consent obtained • NT/ nasal bone ultrasound still in place * NSGC Position Statement; Benn et al, 2011; Bianci et al., 2014
NIPS for ALL
• Provider initiated discussion of patient desire for information – Written information provided – Decision aides used
• NT ultrasound or anatomy US for those desiring information
• Patients who desire NIPS meet with a genetic counselor for written informed consent
Wolfberg, AJ et al. (2014)
Suggested Follow up of NIPS
• After LOW RISK NIPS: – msAFP 15-21 6/7 wks (screen for ONTD) – Anatomy ultrasound ~>20 wks gestation
• HIGH RISK NIPS should be confirmed by: – CVS 11-13 6/7 wks OR – Amniocentesis (>16 weeks) OR – Blood chromosomes at delivery
Suggested Follow up of NIPS
• Genetic Counselor consult for high risk NIPS – Review of results including positive predicted
value – Review condition – Review testing options – Discussion of prenatal care and delivery plan – Offer written resources and support – Follow up with referring provider
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How to choose a NIPS lab/product
1. Accuracy, Accuracy, Accuracy 2. Transparency 3. Reporting 4. Cost 5. Customer support and easy of ordering
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Reporting differences
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Maternit21 Illumina Harmony Panorama
Result reporting
Positive/ Negative
Positive/negative/ aneuplody suspected
Risk assessment range 0.1% to 99%
Risk assessment range 0.1% to 99%
Y only No no yes no
45, X only No no no Yes
Sex chromosome aneuploidy
If noted will report
Opt-in 45,X always reported others as noted
Triplody/ vanishing twin detection
no no no Yes
Microdeletion panel
Yes (opt-OUT)
n/a n/a Yes (opt-IN except for 22q)
What is Fetal Fraction? • Percentage of cell free
DNA originating from pregnancy
• 4% fetal fraction • Some women require
a re-draw • NIPT may not be
possible for all patients – Women with larger
body mass
Illustration modified from: Greenwood Genetics, Genetic Counseling Aids, 6th Edition, 2013
Why is Fetal Fraction important?
• Maternal serum from two NON-pregnant patients were sent to labs
• Two lab reported normal female
• Two labs were unable to provide report due to low Fetal fraction
Measures ff
Reports ff
Sequnom MarterniT21
Yes No
Illumina Verifi
No No
Harmony Ariosa
Yes Yes
Panorama Natera
Yes Yes
40th Annual High Risk Obstetrics Seminar 32 Takoudes, T. and Hamar Benjamin. (2014)
NIPS Products
33
Massively Parallel Shotgun
Sequencing (MPSS)
Targeted Sequencing
Sequenom MaterniT21TM
Illumina Verifi®
Ariosa
HarmonyTM
Targeted Sequencing
Natera
PanoramaTM
COUNTING SNPs
List price $2762 $1200 $795 $2600
TAT 5-7 days 3-6 days 7 days 7-10 days
40th Annual High Risk Obstetrics Seminar
Incidence of conditions
34 Incidence out of 100,000 Live Births
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Additional Panels
Natera (Panorama) • 22q11.2 DiGeorge (in/out) • 1p36 Deletion • 15q Angelman/Prader-Willi • 5p Cri-du-chat
• Triploidy/vanishing twin
Sequnom (MaterniT21 PLUS) • 22q11.2 DiGeorge 1p36 deletion syndrome • 4p deletion Wolf- Hirschhorn • 5p Cri-du-chat 8q Langer-Giedion 11q Jacobsen • 15q Prader-Willi/Angelman
Trisomy 16 Trisomy 22
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More Common Than Down Syndrome in Younger Women
36
Maternal Age
Microdeletions Panel
Down Syndrome
Down Syndrome prevalence from Snijders, et al. Ultrasound Obstet Gynecol 1999;13:167–170. Total prevalence shown for 6 microdeletions using higher end of published ranges from Gross et.
al., Prenatal Diagnosis 2011; 39, 259-266; and www.genetests.org. Total prevalence may range from 1/1049 - 1/2113.
1/2000
1/1000
1/500
1/250
20 22 24 26 28 30 32 34
Pitfalls and Challenges
• When you have a hammer, is everything a nail?
• What will be missed by NIPS? • Ambiguous results • Discordant results for gender • Some samples fail. No results and
patient further in gestational age.
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Are we better off?
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Know and know
know that I don’t know
Don’t know that I
know
don’t know don’t know
NIPS +21 Family and
provider notified
NIPS – Low risk /
patient understand limitations
Suspected DX by screening
BUT All testing declined
NIPS – But falsely reassured
Resources
• National Society of Genetic Counselors nsgc.org
• National coalition for Health Professional Education in Genetics nchpeg.org
• Society of Maternal Fetal Medicine smfm.org
• American College of Medical Genetics acmg.net
• Genetics Home Reference ghr.nlm.nih.gov
40th Annual High Risk Obstetrics Seminar
References 1. Ashoor G, Syngelaki A, Wagner M, et. al. (2012) “Chromosome-selective sequencing of maternal plasma cell–free DNA for
first-trimester detection of trisomy 21 and trisomy 18” Am J Obstet Gynecol, 206:322.e1-5. 2. Benn PA, Borrell A, Cuckle H, Dugoff L, Gross S, Johnson J, Maymon R, Odibo A, Schielen P, Spencer K, Wright D, Yaron Y.
(2011) “Prenatal detection of Down syndrome using massively parallel sequencing (MPS): a rapid response position statement from a committee on behalf of the Board of the International Society for Prenatal Diagnosis.”
3. Bianchi DW and Wilkins-Haug L, (2014) "Integration of Noninvasive DNA Testing for Aneuploidy into Prenatal Care: What Has Happened Since the Rubber Met the Road?," Clinical Chemistry, 60;1:78-87.
4. Canick, JA, et al. (2012). “DNA sequencing of maternal plasma to identify Down syndrome and other trisomies in multiple gestations”. Prenatal Diagnosis. 32: 730-734.
5. Devers P, Cronister A, Ormond K, Facio F, Brasington K and Flodman P, (2013) "Noninvasive Prenatal Testing/Noninvasive Prenatal Diagnosis: the Position of the National Society of Genetic Counselors," J Genet Counsel (2013) 22:291–295
6. Fairbrother, G, et al. (2013). “Clinical experience of noninvasive prenatal testing with cell-free DNA for fetal trisomies 21, 18, and 13, in a general screening population”. Prenatal Diagnosis. 33: 580-583.
7. Gregg AR, Gross SJ, Best RG, Monaghan KG,Bajaj K, Skotko BG, Thompson BH, Watson MS. 2013. “ACMG statement on noninvasive prenatal screening for fetal aneuploidy.” Genet Med 15(5):395-398.
8. Livingston, Deborah, (2013)"ACMG Suggests Broader Application for Noninvasasive Prenatal Screening Tests NIPS tests shouldn’t be limited to high-risk pregnancies, " The AJMG Sequence: Decoding news and trends for the medical genetics community, DOI: 10.1002/ajmg.a.36087
9. Nicolaides, K, et al. (2012). “Noninvasive prenatal testing for fetal trisomies in a routinely screened first-trimester population”. American Journal of Obstetrics and Gynecology. 207: 374.e1-6.
10. Takoudes, T. and Hamar Benjamin. (2014), “Non-invasive prenatal testing performance when fetal cell-free DNA is absent,” Ultrasound Obstet Gynecol doie:10.1002/ugo.14715.
11. Wolfberg, AJ et al. (2014), “Nuchal translucency measurement plus non-invasive prenatal testing to screen for aneuploidy in a community-based average-risk population,” Ultrasound Obstet Gynecol 44: 491–494.
40th Annual High Risk Obstetrics Seminar
Genetic Counselors Board certified and licensed genetic counselors:
Martha Dudek Jill Nichols Caitlin Grabarits
Genetic counseling appointments available: One Hundred Oaks Clarksville Springfield Franklin
615-343-5700