Prenatal Testing And Screening

Lecture Outline

1. Definitions

2. Age related risks

3. Etiology and phenotype of chromosome anomalies

4. Risks, phenotype and testing options of ONTD

5. 1st and 2nd Trimester Prenatal Testing Options

6. 1st and 2nd Trimester MS Screening Options

7. New approaches to combining these tools

8. Ultrasound as a screening tool

Testing Defined

A procedure that (in most cases) provides adefinitive answer to the question that is beingasked. Tests that have low false positives andnegatives are considered diagnostic.

In the case of prenatal testing there is a risk ofmiscarriage associated with all the currentlyavailable diagnostic invasive tests.

Screening Defined• Identify those at increased risk who are

not be perceived to be at risk

• Does not dx definitively

• Follow-up options available for definitive information

• Sensitivity=True positives/all affected

• Specificity=True negatives/all unaffected

Baseline Risk for Having a Child With a

Serious Birth Defect

3-5%

WE DON’T GET BETTER WITH AGE

Maternal

Age

DS RiskBirth

All Chrom AbnBirth

DS risk Mid trimester

20 1/1231

30 1/185 1/384 1/685

33 1/592 1/285 1/452

34 1/465 1/243 1/352

35 1/385 1/178 1/274

36 1/287 1/149 1/213

37 1/225 1/123 1/166

38 1/177 1/105 1/129

39 1/139 1/80 1/100

40 1/109 1/63 1/80

41 1/85 1/48 1/61

NORMAL MALE KARYOTYPE 46,XY

NORMAL FEMALE KARYOTYPE 46, XX

WHEN MEIOSIS FAILS

Normal Sex cell production

Monosomy

Non-disjunction

Sex Cells

Sex Cells

T21 Tid Bits• ~70% T21 due to an error in maternal meiosis I

• ~ 20% maternal meiosis II

• ~ 5% occur during spermatogenesis (meiosis II)

• 5% of trisomic 21 error in mitosis

– no advanced maternal age and there is no preference for which chromosome 21 is duplicated in the mitotic error

• most common chromosomal abnormalities in liveborn children

~1/700

Phenotype• Moderate mental retardation

• Characteristic facies – upslanting palpebral fissures – epicanthic folds, – midface hypoplasia, – macroglossia

• Congenital malformations

– heart (30-40%), atrioventricular canal – gastrointestinal tract, such as duodenal stenosis or atresia,

imperforate anus, and Hirschsprung disease – Leukemia (both ALL and AML) 10-20x– acute megakaryocytic leukemia occurs 200 to 400 times

more frequently in the Down syndrome – 90% have significant hearing loss, usually of the

conductive type

Trisomy 18, Edward syndrome

1/8000

Facial•microcephaly with prominent occiput •narrow bifrontal diameter •short palpabral fissures •low-set malformed ears •cleft lip +/- palate •narrow palatal arch •micrognathia Skeletal•neck •webbed •chest •short sternum •widely spaced nipples •hips: •small pelvis, congenital dislocation of the hips, limited hip abduction •extremities: •phocomelia •rockerbottom feet or equinovarus short dorsiflexed big toes fixed flexion deformity of the fingers (overlapping of the 2nd and 5th fingers over the 3rd and 4th fingers) simple arch pattern of the fingers and toes hypoplasia of fingernails single crease of 5th finger or all fingers (absence of interphalangeal flexion creases) simian crease

Central Nervous Systemsevere mental retardation hypotonia -> hypertonia neural tube defects poor suck and weak cry failure to thrive ocular anomalies Respiratory•apnea Cardiovascular( >95%)•major: VSD, ASD, PDA •minor: transposition, ToF, coarctation, anomalous coronary artery, dextrocardia, •aberrant subclavian artery, arteriosclerosis, PS, bicuspid aortic and/or pulmonic valves Gastrointestinal•inguinal, umbilical, and/or diaphragmatic hernia •congenital defects: •diastasis recti, heterotopic pancreas, malrotation, Meckel's, tracheoesophageal fistula Genitourinary•cryptorchidism •congenital defects: double ureter, ectopic kidney, horseshoe kidney, hydronephrosis, polycystic kidney

Trisomy 18

47,XXY

• T13 Patau syndrome• 1/5,000

•severe mental retardation •coloboma, •(a cleft palate) and/or a cleft lip •hypotonia •skeletal abnormalities (polydactyly)•Renal heart defects•holoprocencephaly

Kleinfelter syndrome

1/1000

45, X

45, X

Miscarriage Turner syndrome1/3000 liveborns

Neural Tube Defects

• Second most common major congenital defect (1-2/1000)

• Not a chromosome anomaly

• Routinely tested and screened for in pregnancy

• Failure neural tube to close at 28 days gestation

• 20% are closed lesions and difficult to detect prenatally

Open Neural Tube Defects

Closed lesions

Open lesions

In 1976 the American College ofObstetrics and Gynecology

recommendedprenatal diagnosis be offered toall women 35 years of age and older atdelivery.

INDICATIONS FOR PRENATAL DIAGNOSIS

Maternal age > 35 years at EDC

Abnormal maternal serum screening for: DS cut off > 1/270 Increased risk Trisomy 18 or 13

Smith-Lemli-Opitz syndrome ONTD

Previous child with chromosomal abnormality or diagnosable genetic disorder

Balanced translocation carrier

Ultrasound anomaly (soft sign vs frank anomaly)

PRENATAL DIAGNOSTIC PROCEDURES

AMNIOCENTESIS

CHORIONIC VILLUS SAMPLING

PERCUTANEOUS UMBILICAL CORD SAMPLING

PERFORMED ROUTINELY 16-20 WEEKS

AMNIOCENTESIS

ULTRASOUND GUIDED AMNIOCENTESIS

Amniocentesis Testing

• Chromosome analysis

• AF-AFP levels• Acetylcholinesterase

• Risk of miscarriage associated with procedure 1/100-1/400

ONTD

Advantages

• Highly reliable results 99+%• Familiar• Long standing reputation• NTD detection

Disadvantages• Late in gestation

– Decision making– Privacy– Mom feels movement

• Fear of needles

• Needle invades the sac

Fetus: 12 weeks gestation

Transcervical

Chorionic Villus

Sampling

Transabdominal

Performed >10 wks-13 weeks

Chromosome analysis

Risk 1/100-1/200

• trophoblastic shell cells • Syncitiotrophoblasts – poly-proliferate

tissue type=directs• cytotrophoblasts• Mesodermal core=tissue culture• frorm finger-like extensions

Disadvantages• Placental mosaicism 1% of CVS is confirmed in the fetus ~ 10-

40%

• Second trimester amniocentesis mosaicism ~ 0.1-0.3% & confirmed in a fetus up to 70% of the time.

• ?LRD risk prior to 70 days gestation (10 weeks)

• Higher loss rate

• Less access to procedure

• Higher chance of insufficient sample

• Early test=risk of sampling a fetus potentially destined to miscarry

• No ONTD testing

• More risk of vaginal bleeding

• Speculum

Benefits(1) Earlier in gestation

(2) rapidly growing cell cultures practically free of maternal

cell contamination

(3) an efficient direct method to obtain high quality metaphases from the of the syncitiotrophoblasts tissue which the fetal karyotype is defined within a few hours of chorionic villi sampling (specialty cyto techinque)

(4) is suitable for a rapid, direct diagnosis of the related metabolic diseases.

(5) placental mosaicism (trisomic rescue in fetus) can increase the risks of genetic abnormalities such as uniparental disomy

G. Simoni1, Human Genetics 1983  

Fetal Blood Sampling“PUBS”

Percutaneous Umbilical Cord Sampling(PUBS)

or Cordocentesis• ~2% risk of loss• Technically difficult prior to 20 weeks • Blood disorders such as hemophilia and

anemia– Useful for detection of Rh isoimmunization of the

fetus (blood cell count and oxygen level)→erythroblastosis fetalis (HDN)

• Chromosomal abnormalities Fetal karyotype in 48 hours

• Infections such as toxoplasmosis and rubella. • The procedure is also used to perform blood

transfusions to the fetus and to administer medication directly into the fetal blood supply.

Reproductive Decision Making

RISK Fetal Aneuploidy

Procedure Related

RISK

TO TEST OR NOT TO TEST

I want to know The benefits outweigh the risks Options are desirable Because my doctor says so…..

Not sure I want to know Risks are a big worry Options stink Because my doctor says so….

So what to do what to do…………..

SECOND TRIMESTERMATERNAL ANALYTES FOR ANEUPLOID

SCREENING

FETAL

Alpha-fetoprotein- AFPEstriol- uE3

PLACENTAL

Estriol- uE3Human chorionic gonadotrophin- hCGInhibin-A

2nd Trimester MSS Overview

Used for detection of:

1. ONTD 2. Down syndrome3. trisomy 18 4. Smith-Lemli-Opitz syndrome

Second trimester MSS

AFP

HCG

uE3

Inhibin-A

ONTD DS T18 SLO

Seru

m M

ark

er

XMoM

0.7

2.1*

0.7

2.1

0.21*

0.65

0.36

0.4*

Smith Lemli Opitz Syndrome

• Defect enzyme in the conversion of 7-dehydrocholesterol to cholesterol.

• Affects 1 in 20,000 to 40,000 births • Autosomal Recessive• Mental Retardation• Slow growth• Heart defects• Facial cleft

• Screen positive women have uE3 < 0.4 MoM • ~2% baby affected• Testing AF for 7-8- dehydrocholesterol (7/8-DHC) levels

Other……..

AFP

HCG

uE3

Inhibin-A

Seru

m M

ark

er

Turner T13 Triploidy Pregnancy complications

uE3 lower than 0.1 MoM

• Increase risk for x-linked ichthyosis

ONTD screening

• Abnormal Screen MS-AFP > 2.5 MoM

• Elevated MSAFP recommendations:– Ultrasound– 90% r/o spinal lesions

100% r/o anencephaly– VWD reduced with normal scan– If MSAFP >4.0 MoM and NL U/S

offer invasive testing

Detection Rates

• Anencephaly 100% 100%• Spina Bifida 85-90% 90%

• VWD– Omphalocele70%95%– Gastroschisis90%95%

MSAFP Add Ultrasound

Screening for DS 2nd Trimester 1/270 Cut-off

• Age 20 5

• Age AFP 30 5

• Age AFP + HCG 55 5(Double)

• Age AFP+uE3+HCG 61 7 (Triple)

• Age AFP+uE3+HCG+Inhibin-A 755 (Quad)

DR% FPR%

The detection rate for Down syndromeusing the second trimester triplescreen is:

50% for women <35 y/o 88% for women >35 y/o and

~60% for all women

Quad ~75%

First Trimester Integrated Screening For

Trisomies =FIRST

•Entire analysis performed 11-13 6/7 weeks gestation

•MS free Beta hCG and PAPP-A (pregnancy associated plasma protein)

-obtained bw 9-13 weeks

•Nuchal measurement 11-13 6/7

•Maternal age

•Detection Rates:~80% Down syndrome 1/270~90% trisomy 18 1/100

Nuchal Translucency (NT)Nuchal Translucency (NT)

• Gestation 11 to 13 6/7 wksGestation 11 to 13 6/7 wks

•Mid-sagittal viewMid-sagittal view

• Fetus away from amnionFetus away from amnion

• Enlarge head & upper Enlarge head & upper thoraxthorax

• Widest part of NTWidest part of NT

First Trimester Screening

• Free beta HCG

• PAPP-A

• NT

DS T18

>3.0 mm

Down syndrome DR ~1:270 Cut-off

MA + NTMA + NT11 10%10% 71% 71%

MA + BiochemistryMA + Biochemistry33 67% 67%

MA + NT + MA + NT + BiochemistryBiochemistry2 2 5%5% >80% >80%

1. Schuchter et al. Prenatal Diagnosis 22: 211, 2002

FPR DR

2. Wapner et al. NEJM 349: 1405, 20033. 3. Spencer et al UOG 1999Spencer et al UOG 1999

Nasal bone

Recommendations 1st Trimester Nuchal >3.5

• CVS

• Targeted ultrasound evaluation 18-20

• Echocardiogram

• Residual 5-6% risk neonate may have a yet undefined genetic syndrome…

Fetal Nasal Bone• 65% DS have absent nasal bone

• General population 1% – African Americans 8%-10%

• Secondary screen

• Difficult to obtain

• Higher false positive in 1st

Integrated Screening

PAPP-A and Nuchal

Quad Screen

Screen Positive1/270 DS1/100 T182.5> ONTD

Screen Negative

AmnioUS Nothing

10-14

16-19

Results 18-20

Timeline weeks

Decision Making

20+ wks

+

=DR: 94-96%FPR: 5%

Advantages• Increases detection rate• Decreases false positive rate (fewer tests

and fewer procedure related losses)

Disadvantages• Long wait time for result• Unable to utilize CVS and early detection • Late GA by the time amnio results final

Contingent

First Trimester Screening

High risk Low Risk Intermediate

Triple Screening IntegrationOffer CVS

High riskLow Risk

US and AmnioStop

Advantages• Increase detection rate 90%• Decrease FPR 2%• Reduce the number of amnios

performed in low risk pregnancies

Disadvantages

•New (limited data)•Hard to determine uptake•Offered at few institutions

RISK OF ANEUPLOIDY BASED ON GA AND ANOMALY

3D UltrasoundFetal Face24 weeks Gestation

Fetal MRI

FETOSCOPY

Amnion (stuck twin)

Umbilical cord

QUESTIONS ??