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Genetic Testing in Pregnancy Johanna Warren, MDOAFP Women’s Health Winter ConferenceJanuary 19, 2014

+What genetic screening test do you routinely offer your patients?

A. Quad/Penta screenB. Integrated ScreeningC. Nuchal Translucency (NT) onlyD. Stepwise Sequential ScreenE. NIPT (cffDNA)F. none

+ Learning ObjectivesUse the concepts of pre-test probability, positive

and negative predictive values as they apply to testing for fetal aneuploidy.

Outline advantages and disadvantages of various approaches to first-trimester and second-trimester screening as well as invasive diagnostic testing for Down syndrome.

Discuss emerging cell-free fetal DNA (cffDNA) technology and review indications for use in screening.

Summarize the current recommendations for cystic fibrosis screening.

+StatisticsPre-test probability

Prevalence of the disease Post-test probability after one or more preceding tests Rough clinical estimation

Positive predictive value (PPV) the likelihood that an individual with a positive test result

truly is affected/has the genetic condition inherently dependent upon the prevalence

Negative predictive value (NPV) the likelihood that an individual with a negative test result

is truly unaffected/does NOT have the genetic condition measure of test accuracy

+What Makes a Good Screening Test? The condition should be an important health problem. There should be an acceptable test for the condition.

High sensitivity to detect not yet clinically detectable condition High specificity to minimize false positives

The test should be available to the population. The test should be affordable. There should be acceptable treatment for the condition. There should be a “latent stage” of the disease, allowing

for detection/testing before a critical point, during which treatment would be optimal.

+What is the purpose of antenatal genetic testing?Assess risk for chromosome abnormalities

particularly Down syndrome & Trisomy 18Determine which pregnancies should be

offered invasive prenatal diagnosis

+Who Should Be Screened?ACOG Practice Bulletin (2007)

All pregnant women, regardless of their age, should be offered screening and diagnostic testing for aneuploidy.

Women should be counseled regarding the differences between screening and invasive diagnostic testing.

Ideally, patients seen early in pregnancy should be offered screening that combines 1st and 2nd trimester testing Screening test chosen will depend on

availability of NT US as well as CVS

+ Testing for Fetal Aneuploidy – Focus on Down Syndrome (trisomy 21) Explosion of available screening tests Detection of Down Syndrome 90+% with non-invasive

screening tests Confirmation diagnosis still requires invasive testing

Amniocentesis Chorionic villus sampling

+Quick ReviewSeveral methods for combining first-

and second-trimester screening reach higher detection rates for Trisomy 21 than either first- or second-trimester screening alone.

Common options: Ultrasound: Nuchal Translucency Integrated Screenings Quadruple/Penta Marker Screenings Stepwise Sequential Screening

+First Trimester - UltrasoundNuchal Translucency (NT)

Normal: 1-3mm Increased NT is an indication of a chromosomal

abnormality, single gene defect, or birth defects (commonly cardiac defects)

Nasal Bone Absent/hypoplastic in 70% T21

Ductus Venosus – reversed a-wave flow Detection rate 75%, FPR 5%

Tricuspid Regurgitation Detection rate 67.3% in T21, FPR 5.2%

+ Integrated ScreeningsIntegrated Screening

Ultrasound for Nuchal Translucency (NT) + serum PAPP-A/hCG analysis between 10-13.6wks GA; results of these tests are HELD

Serum quad screen test between 15 -19 wks GA At that time, the results of all the studies, combined

with risk assessment due to the patient's age, are used to present a single-risk figure

Serum Integrated Screening first-trimester serum PAPP-A/hCG test result is

combined with a second-trimester quad screen test to provide a single-risk figure (no NT US)

+Quadruple Marker ScreeningMeasure raw values of:

AFP (alpha fetoprotein) ue3 (unconjugated estriol) hCG (human chorionic gonadotropin) DIA (dimeric inhibin A)

Compare to median value for the appropriate gestational age (MoM)

Valid between 14-22.9 weeks GA (optimal 16-18wks); risk of NTD not provided for samples collected prior to 15 weeks.

+Stepwise Sequential ScreeningFirst Trimester

NT US + serum PAPP-A analysis between 10-13.6wks GA

Results combined with the patient's age-associated risk,

Patient is given a risk assessment for aneuploidy may choose at this time to undergo invasive testing

(e.g., amnio or CVS), or add quad screen test at 15-19 wks GA

Second Trimester Quad screen test at 15-19 wks GA a new risk is assessed based on the results of patient’s

age and both the first- and second-trimester screening test results

+ Sequential ScreeningHow do you decide when to proceed with

invasive testing?

Risk of miscarriage (approximate; operator-specific) chorionic villus sampling (CVS) ~ 0.5-1/100 amniocentesis ~1/1000

After 1st trimester results return: If risk is greater than ~1 in 50, offer CVS If risk is less than ~1 in 1,000, advise no further testing is

necessary. If risk is between these two (arbitrary) cutoffs, offer quad

screen test after 15 wks GA, and determine a new risk assessment

+ FASTER Trial DataScreening Test Best

Detection Rate for Down Syndrome

NT alone 70%Quad screen alone (2nd trimester) 81%First Screen (with NT) 87%Serum Integrated (1st tri PAPP-A/hcg + 2nd tri quad)

88%

Sequential Screen (1st tri PAPP-A + NT + 2nd tri quad)

94%

Integrated Screen (1st tri PAPP-A/hcg + NT + 2nd tri quad); patient does not receive results until 2nd trimester testing complete

96%

+What do you do with…

Normal Ultrasounds Other Abnormal Serum Studies

+Normal UltrasoundsNormal ultrasound: 50-60% decrease

in risk for chromosome abnormalitiesRemember that at least 30% of fetuses

with Down syndrome have NO abnormal ultrasound findings!

+ Low 1st Trimester PAPP-A Pregnancies with PAPP-A of ≤ 5%tile (0.4MoM) are at

increased risk for: Spontaneous fetal loss < 24 wks GA Low birth weight Preeclampsia Gestational HTN Preterm birth and stillbirth Preterm premature rupture of membranes Placental abruption

+ Abnormal 2nd Trimester Serum Values

Elevated hcg Elevated inhibin-A

Low uE3

preeclampsia preeclampsia steroid sulfatase deficiency

preterm delivery IUGR Smith-Lemi-Opitz syndrome

low birth weight infant

preterm delivery congenital adrenal hyperplasia

IUFD adrenocorticotropin deficiency

placental abruption hypothalamic corticotropin deficiency

anencephaly

+What’s next?

+ Cell-free Fetal DNA (cffDNA) Screening or Diagnosis? Known as “Noninvasive Prenatal Testing” or “NIPT” Technology uses circulating cell free fetal DNA found in the

maternal plasma Thought to be derived primarily from placenta

New recognition of limitations of screening with pregnancies with placental mosacisms

Unclear data for egg donor pregnancies Available as early as 10th week of pregnancy Cleared from maternal blood almost immediately after

childbirth

+ Cell-free Fetal DNA (cffDNA) 2012 publications (Sparks et al., Ashoor et al., Bianchi

et al.) targeted (chromosome-selective) sequencing of

chromosomes 18 and 21 highly accurate potentially more cost-effective

Technology can be expected to identify 98% of cases of T21 with a false-positive rate of < 0.5%.

Multiple different labs MaterniT21plus by Sequenom Verifi by Verinata Harmony by Ariosa

+ Cell-free Fetal DNA (cffDNA) Labs MaterniT21plus by Sequenom

>99% detection for T21, T18 ~90% detection for T13 <1% false positive rate Cost: $235-2700

Verifi by Verinata >99% detection for T21, T18 ~80% detection for T13 >90% detection for 45,X <1% false positive rate Cost: $200-1200

Harmony by Ariosa >99% detection for T21, T18 ~80% detection for T13 <1% false positive rate Cost: up to $795

+ Cell-free Fetal DNA (cffDNA) ACOG Committee Opinion, Dec. 2012 – “Noninvasive

Prenatal Testing for Fetal Aneuploidy” cffDNA testing should be an informed patient choice after

counseling and should not be part of routine prenatal laboratory assessment.

cffDNA testing should not be offered to low-risk women or women with multiple gestations (has not been studied).

A negative cffDNA test result does not ensure an unaffected pregnancy.

A patient with a positive test result should be referred for genetic counseling and should be offered invasive prenatal diagnosis for confirmation of test results.

+ Indications for Considering Use of cffDNA for Screening Maternal Age ≥ 35 years at delivery Fetal ultrasound findings indicating increased risk of

aneuploidy CPCs? IEF? Clinodactyly? Absent/hypoplastic nasal bone?

History of a prior pregnancy with a trisomy Positive test result for aneuploidy (any serum test) Parental balanced translocation with increased risk of fetal

trisomy 13 or 21.

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+ConsiderationsPrimary insurers are generally reimbursing

for first-trimester screenings, including NT ultrasounds as well as NIPT.

Are there referral sites available to your patients for appropriate genetic counseling and NT US measurements? What about for CVS and/or amniocentesis?

What testing strategy makes the most sense for your patients?

How do you see it evolving in the next 1-2 years?

+Testing for Cystic Fibrosis (CF) Site of genetic defect = CF Transmembrane

Regulator (CFTR) gene, a chloride channel protein ~ 1700 mutations of CFTR gene have been

described Disease incidence: 1 in 2500 in the non-Hispanic

white population Carrier frequencies:

1/24-25 Caucasians of European descent or Ashkenazi Jews

1/58 Hispanic American 1/61 African American 1/94 Asian American

+ CF Genetic Testing

Difficult to assign a single ethnicity to individuals

ACOG 2011 Recommendation – offer CF carrier screening to all women of reproductive age

Need to be screened only once

+ CF Parental Genetic Testing Sequential testing

Test mother for carrier state If positive, test father

Concurrent testing Test mother and father simultaneously

Advantage: can be done prior to conception Limitation: depends on accurate ID of father If both parents are unaffected but family hx of CF exists:

Genetic counseling Identify if CFTR mutation analysis in affected family member is

available

+ Positive CF Testing

If both parents are carriers… and prior to conception

offer ART options for diagnosis of embryo and currently pregnant

offer CVS or amnio to confirm status of fetus

No in-utero treatments exist Variable clinical scenarios, with median survival for

patients 37yrs

+ “Residual Risk” Potential risk of having an affected child with CF

after testing is completed and is negative Varies by race and current testing panel for gene

abnormalities Will vary over time and by laboratory Newborn screening panels that include CF

screening do not replace maternal carrier testing

+Summary Recommendations Genetic screening in pregnancy (and pre-conception!)

is rapidly getting more complex. OHSU Online Course (FREE!) 0.5 CME credits available www.ohsu.edu/prenatal-screening

Understand your patient population and your local capabilities, specifically as they relate to genetic counseling, ultrasound expertise, and diagnostic testing.

Develop practice workflows that allow women to access early genetic screening should they desire.

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