Aneuploidy Screening

Dr.m.ali mohammadiNima Mohammadi

2010-11

amirkabirimagincenter.com

ONTD Screening

hCG

Free Beta

DownScreeningAFP Only

1st TrimesterFree Beta

1st TrimesterPAPP-A

NT

1st TrimesterBiochem

+NT

PRENATAL SCREENING 28 YEAR HISTORY

NB

Maximizing First

Trimester Screening

Approaches to Prenatal Testing

Diagnostic Tests Screening Tests

Amniocentesis AFP CVS Quad Screen Cordocentesis Ultrasound Ultrasound

Characteristics of an Ideal Screening Test

• High sensitivity - Identifies a high percentage of affected individuals

• High specificity - Does not alarm a high percentage of unaffected individuals

• Positive early enough in gestation to allow intervention

• Easy and inexpensive to perform

Prenatal Chromosome Screening

Low risk patients ≥ 35 yrs, other chromo risks (FH and U/S) Biochemical screens U/S screen 1st or 2nd Trimester Risk cutoffs: ≥ 35 yrs risk at specific gestation False Positive Rate = 5%

First Trimester Screening FreeBeta / PAPP-A / NT/ NB Screening 11-14 wk Fetal Ultrasound Exam CVS (Early Amniocentesis N/A)

Second Trimester Screening AFP &/or Ultrasound NT 3.0mm in 1st then Fetal Anatomy

and Heart Ultrasound in 2nd

NT General Population Screening● 96,127 patients 10-14 wks DR= 82.2% at 8.3% SPR (1:300 cut off) DR= 77% at 5% SPR

● All patients had 3 NT’s, largest NT used● Only 4.2% lost to follow-up● True DR after 4% bias adjustment for 7%

SAB’s between 12-16wks = 78% at 8.3% (Spencer 2003), not 60%.

USA National FMF/SMFM/ NT Standard

1. Standardized training, practical NT certificate, external audit, and NT certificate renewal required for 1st. Same Standard for > 70 countries

2. Over 500,000 NT’s (USA + UK) in database3. Consistent NT’s in 19 prospective studies of over

120,000 patients by Operator-Specific FMF certificate operators. More now with SMFM.

4. Non-certified operators are inconsistent at 9-15 week.

5. SMFM has entered the Educational Certification

A National Standard Must Be Maintained

Everyone who does ultrasound should NOT do NT unless they have gone through the training for numerous reasons but mostly

for consistency.Most labs for bloods (PAPP A and hCG will

not accept specimen without certification number)

Fetal Abnormalities At Birth

12 weeks NT Percentile 3mm 10%

6mm 90%

Nuchal Translucency:Advancing Gestation

0

2

4

6

8

10

%

10 11 12 13

Gestation (wk)

NT >2.5 mm

Anomalies in 4,116 Chromosomally Normal Fetuses with Increased Nuchal Translucency

> 95th centile at 10 -14 weeks of gestation n(%)Major cardiac defects 43(27%)(increases exponentially with NT)Talipes 15(9%)Body stalk anomaly 10(6%)Diaphrag. hernia 9(6%)Megacystis 8(5%)Exomphalos 7(4%)Akinesia deformity 6(4%)Ventriculomegaly 6(4%)Anencephaly 5(3%)Encephalocele 4(3%)Multicystic kidney 4(3%)Spina bifida 4(3%)Hydronephrosis 3(2%)

1st Trimester Screening - USA National FMF Standard NT Training, Certificate and External Audit. Orlandi, et al., (1997) Krantz, et al., (2000) 10,581 pts NICHD-BUN Trial (2002) 8,514 pts CYRANO Nasal Bone Study NICHD FASTER Research 2004

First Trimester : High Free Beta hCG (2.0 MOM) Low Pregnancy-Associated Plasma Protein-A (PAPP-A) (0.4 MoM) NOT B-hCG (1.2 MOM) – not associated with DS

Second Trimester: AFP for open spine and skull defects Free Beta hCG (2.8 MoM) or B-hCG (1.8 MOM) Estriol

Which Biochemical Marker?

freeBeta/PAPP-A

AFP/freeBeta

2nd Trimester ResearchNO PUBLISHED USA PROSPECTIVE OR NIH STUDIES TO SHOW BENEFIT, NO ACOG STATEMENTS.

QUAD (AFP/ß-hCG/Estriol/Inhibin A) INTEGRATED (1st & Quad) – Can SPR be lowered by

requiring 1st risks be withheld? No, not in clinical practice.

● Medical, Legal (HIPPA), ethical problems of withholding known ↑ risks (NT, NB) from physician/patient until

17-20 wks to add 4 more tests.

● Six markers double cost for lower DR and higher SPR than 1st.

2nd Trimester Research● FASTER Integrated Research - FIRST WORKS better than Integrated when all DS fetuses are counted.● 1st prospective FMF clinical studies have

↑ detection rate (DR) and ↓ screen positive rate (SPR), so 1st obviates any benefit of Integrated.

● Integrated withholds advantages of 1st for no benefit. Is it an acceptable medical procedure?● Impact of immediate results after NT!!!!!!

Eliminates broken tubes, biohazard, hemolysis, specimen degradation, and centrifugation

Stabilizes protein markers Reduces Marker Variation (S.D.)

Increases Separation between Distributions

Patient and staff acceptance!!!!

Dried Blood Screening

CUTS SCREEN POSITIVE RATES BY HALF (~1/2) RAISES DETECTION RATES

First Trimester Screening:When?

freeBeta / PAPP-A

freeBeta / PAPP-A / NT

Weeks

24-84mm CRL

9w 0d - 13w 6d LMP

0 5 9 11 12 13 14

(Adjusted for MA, GA, FH, Wt., Ethnicity)

45-84mm CRL

11w 0d – 13w 6d LMPNot 10-14 wks

DATING BY CRL

Immediate results*

DS freeBeta/PAPP-A DR @ 5% SPR

Study-FMF NT DS Weeks Detection (%)

Krantz, 1996 22 10-13 63Wald, 1996 77 8-13 62Berry, 1997 47 9-13 55Orlandi, 1997 11 9-13 61Haddow, 1998 48 9-15 60Wheeler & Sinosich, 1998 17 9-12 67De Graaf, 1999 37 10-13 55Spencer, 1999 210 10-13 67Tsuckerman, 1999 31 8-13 69Krantz, 2000 50 9-13 63

USA Detection: 63% at 4.5% vs. triple 60% at 7.6%

Why Combine freeBeta / PAPP-A / NT?

FreeBeta / PAPP-A

NT FreeBeta / PAPP-A / NT

% DS Detection

63 82.2 91 (93)*

%SPR 4 8.3 4.4 (2.4)

● NT ALONE DOUBLES SPR AND ↓ DR 10%

● ALWAYS COMBINE NT, freeBeta, PAPP-A TO ↑ DR, ↓ SPR, AND ↑ YIELD

*(Current USA data)

DS freeBeta / PAPP-A /NT DR at 5% SPR

Brizot, 1994, 1995 80 10-13 5.0 89Orlandi, 1997 11 9-13 5.0 87De Graaf, 1999 37 10-13 5.0 85De Biasio, 1999 13 10-13 3.3 85Spencer, 1999 210 10-13 5.0 89Spencer, 2000 7 10-13 5.5 86Krantz, 2000 33 10-13 5.0 91Schucter, 2002 14 11-13 5.0 86Wapner, 2002 61 10-13 5.0 78.7Bindra, 2002 82 11-13 6.8 92Crossley, 2002 37 11-13 6.0 92Spencer, 2003 25 10-13 5.2 92

USA Detection 91% @ 2.4% SPR

Study DS Weeks SPR% Detection (%)

freeBeta/PAPP-A/ NT Krantz et al. Obstet Gynecol 96: 207 (2000)

10,253 patients < age 35 between 9-13 weeks 5,811 patients with NT at 10w4d - 13w3d

SPR Detection Rate YieldDown syndrome 4.8% 91% 1/17Trisomy 18 1.2% 97% 1/17

Yield = 1/17 vs. 1/80 for triple screen

Maternal Age – NT/FreeBeta / PAPP-A

● ≥ 35 YRS – ROUNTINELY OFFER AMNIO

● Some day we will eliminate this FEAR of >35 years!!!!!

We are getting close but it may take a generation!!!!

Age(y) SPR Detection Rate (%)

24 3.2 8825-29 3.7 8830-34 5.3 9035-39 10.3 9140 24.7 97

(Krantz, et al, 2000)

All Ages 4.8 91

Second Trimester Screening AFP / B-hCG / UE3 (16w - 20w) (Triple) SPR DR Yield ONTD 5.0% 80% 1 / 42 Down Syndrome 7.6% 60% 1 / 80 Trisomy 18 0.5% 60% 1 / 16

12%

AFP / freeBeta hCG (13w 4d to 22w 3d) SPR DR

Yield ONTD 2.7% 98% 1 / 25 Down Syndrome 2.8% 80% 1 / 25 Trisomy 18 0.1% 70% 1 / 6

5.6%● HIGHEST DR AND YIELD, LOWEST SPR ●

DS SCREENINGDR % SPR %

Maternal AgeTripleAFP/freeBeta

306080

57.62.9

freeBeta/PAPP-A 63 4.5NT Only 80 5NT/freeBeta/PAPP-A 91 2.4

Down Syndrome Detection

0

20

40

60

80

100

A B C D

A: freeBeta/PAPP-A/NT (11w 0d - 13w 6d)

B: freeBeta/PAPP-A (9w 0d - 13w 6d)

C: AFP/freeBeta (13w 4d - 22w 3d)

D: Triple / Quad (16w - 20w)

93%

63%80%

60%

% D

etec

tion

First Trimester SecondTrimester

2.4% 4.4% 2.9% 7.6%

Patient’s Prefer 1st

All 6 published surveys show patients strongly prefer 1st.

(Monni G, et al. Lancet 1998)(Mulvey S and Wallace, EM. Br J Obstet Gynecol 2000; 107; 1302-5)

Should 2nd Be Moved To 1st?Cuckle & Lith Prenat Diag 19:505 (1999)

“There is overwhelming evidence that screening for Down syndrome could be moved from the second to the first trimester of

pregnancy.” (The Royal College of Obstetricians and Gynaecologists, 1997)

The Time for First Trimester NT Has ComeChasen et al. J. Ultrasound Med 20:1147, 2001

“Given the scientific rigor and the results of the FMF database, in our view nuchal translucency should not be considered

investigational but instead a highly reliable diagnostic screen when performed in expert hands.”

1st & 2nd Trimester Screening Genetic Counseling

Nondirective counseling and obtain Consent

Discuss 1st and 2nd screening in the 1st trimester Copy patients’ Consent to her physician to avoid

automatic 2nd chromosomal screen

Impact of IMMEDIATE RESULTS!!!!!

SMFM supports 1ST (Feb,2004). ACOG to review in 2004.

Jan 2007 ACOG Bulletin

• If patients present in the first trimester, she should be offered a first trimester or combined screening

• If patients present in the second trimester, sh ould be offered second trimester screening

Does a 2nd Trimester Follow-up Chromosome Screen Help or Hurt?

Because 1st ↓ PREVELENCE 91-97%:

Yield will ↓5 fold from 1/17 to ~ 1/250

2nd Triple will ↑ SPR to 13% (2.4% 1st + 10.3% 2nd-BUN)

2nd AFP/freeBeta will ↑ SPR to 4.4% (2.4% 1st + 2% 2nd)

A F/UP FOR SNP AND SPP PATIENTS CANNOT BE ACCURATE (DATABASE, PRIOR RISK) AND WILL CONFUSE.

A 2nd F/UP IS NOT STANDARD OF CARE. PLAN: FOR SNP AFTER 1st DO MSAFP OR U/S?????

Don’t do QUAD screen after patient has had FTS results and risks!!!

NOT the same as a planned sequential or integrated

• What happens when the MSAFP that we normally order after FTS is inadvertently ordered as a QUAD screen? Whoops!!!!

Here is what we do????

Genetic Counseling

Insurers Cover 1st1. freeBeta / PAPP-A – YES – over 85%

2. Second Trimester – YES

3. NT – YES, for ↑ risk patients, for consult referral. For low risk patients, use general ICD codes.

4. CPT Options –● 99242,3, or 4, consult● 76815 (limited) or 76801-22 U/S with documentation

(maternal/fetal, <14wk)● Laboratory bills for biochemistry

Hold on…..

Absence of Nasal Bone in Trisomy 21Fetuses at 11-14 Weeks of Gestation

Lancet (2001) 358:1665-67

Simona Cicero, Patrizia Curcio, Aris Papageorghiou, *Jiri Sonek, Kypros Nicolaides

Harris Birthright Research Centre for Fetal Medicine, King’s College Hospital Medical

School, London, UK*Dept. of Obstetrics & Gynecology Ohio State University, Ohio, USA

Nasal Bone Assessment Absent and/or Short Nasal Bone is associated with

T21, T18, and 58 Other Syndromes Proper NB measurement requires Standardization

and Formal Training (similar to NT Certificate)● Cicero, 2003: After training 100 NB’s

needed for proficiency● Further Study of 1st trimester Short/

Absent NB is need for QC● FASTER, 2004: NB without special training/QC, 9 of 9 DS missed vs.

consistent high DR (70%) for over 20,000 patients, with NB training/QC.

First-Trimester Nasal BoneStudy SPR DR

Cicero (2001) 0.5% 73%

Orlandi (ISUOG 2002) 1.0% 67%

Massoc (ISUOG 2002) 0.8% 75%

Monni (ISUOG 2002) 0.7% 64%

Cicero (2003) 2.8% 66.9%

Orlandi (2003) 1.0% 66.7%

NT + FreeBeta + PAPP-A + Nasal Bone Fetal Medicine Foundation Data

FPR DS Detection Rates

5% 97%

2% 95%

1% 94%

0.5% 91%

vs. Integrated: 1% FPR @85% DR (Wald, 1999)

Prospective DS Screening DR% SPR%

AFP / freeBeta (2nd) 80 2.9Triple (2nd) 60 7.6Integrated(2nd) 76/90/90 3.4/5/5.4

1st 93 2.41st + NB 95 21st + NB 94 1

(CAN,WALD,FASTER)

First PLUS NB has the highest DR and yield with the lowest SPR

First Trimester Advantages Earliest, most effective and safest risk assessment Highest Detection and Yield for chromosomal, heart (40%),

and many other anomalies at the lowest SPR of all screens Early anomaly and perinatal risk management Privacy for patient! Preferred by Patients! Less bonding at early GA Earlier, safer, and lower cost options Early resolution of patient anxiety Early Reassurance for 97.6% of Low Risk patients Meets ACOG guideline for national implementation by

Operator - Specific certificate providers. Cost effective (↓36%), insurance coverage as acceptable

medical procedure.

2nd Trimester ResearchNO PUBLISHED USA PROSPECTIVE OR NIH STUDIES TO SHOW BENEFIT, NO ACOG STATEMENTS.

QUAD (AFP/ß-hCG/Estriol/Inhibin A) INTEGRATED (1st & Quad) – Can SPR be lowered by

requiring 1st risks be withheld? No, not in clinical practice.

● Medical, Legal (HIPAA), ethical problems of withholding known ↑ risks (NT, NB) from physician/patient until

17-20 wks to add 4 more tests.

● Six markers double cost for lower DR and higher SPR than 1st.

2nd Trimester Research● FASTER Integrated Research - FIRST WORKS better than Integrated when all DS fetuses are counted.● 1st USA prospective FMF clinical studies have ↑ DR, ↓ SPR, so 1st obviates any benefit of Integrated.● Integrated withholds advantages of 1st for no benefit. Is it an acceptable medical procedure? YES, so says the 2007 ACOG bulletin!!!!

Prenatal Screening Tests Comparison• First Trimester Combined (1st): NT/Nasal Bone

(NB)/freeBeta/PAPP-A @ 11-14 wks has 94 % detection rate (DR) @ 1% screen positive rate (SPR) or 97% @ <5% SPR. 1st (NT/freeBeta/PAPP-A) has the highest DR (91%) and yield (1/17) with the lowest SPR (2.4%). 1st is the most effective, earliest and safest screening test. 1st identifies other chromosomal anomalies, 40% of heart defects and ↑ perinatal risks. 1st provides optimal pregnancy management, more options and ↓ cost.

Prenatal Screening Tests Comparison

Sequential: 1st, then a 2nd chromosomal screen (2nd) w/o risk adjustment: Lowest yield, Highest SPR Not Recommended. A

follow-up “2nd” test is not standard of care because the YIELD is ↓ 5 Fold, the SPR is over

13%, and 2nd can’t be accurate due to ↓ DS incidence, revised prior risk, and unadjusted 2nd. New Sequential procedure offers results

but lowers first trimester Sensitivity……

Prenatal Screening Tests ComparisonIntegrated Withholding (2nd): NT/PAPP-A @ 10-14wks

and Quad @ 16-20 wks has 75-85% DR @ 2.8% SPR = Lower DR and yield, Higher SPR than 1st, therefore Integrated has No Benefit over 1st. Can we justify withholding known ↑ risks until 17-21 wks for no documented benefit?

● FASTER published, but does not show benefit over 1st when all Down syndrome fetuses are counted however some will elect this. (my opinion)

● 1st studies with higher DS DR @ lower SPR have been published (Cicero 2003). (my opinion)

● Therefore, 1st has negated any integrated benefit while offering many new advantages. (my opinion)

Integrated Disadvantages: (opinion)● Loss of all 1st benefits by requiring known 1st risks be

withheld from patients and physicians for 6-10wks to keep SPR low.

● Physicians can’t withhold known 1st risks for 6-10wks, so SPR rises = no benefit.

● 1st results are not improved with follow up screen and increases SPR.

● 2-3 visits required, incomplete till 17-21 wks.● A 3rd visit required for screen positive patients.● Drop-Out = 8-35% due to U/S reassurance, cost, time, and

insurance lab preferences.● Drop-Out patients are billed for NT/PAPP-A but don’t

receive an integrated test result.

● Fetal and perinatal risks are withheld 6-10wks for no benefit.

● Patients and physicians should not be denied immediate 1st risks or reassurance (>97%). Integrated give a less specific risk.

● Increased patient anxiety. ● Loss of privacy and optimal pregnancy management

options when you go into the second trimester.● All 6 published surveys show patients strongly prefer

First Trimester Screening. Triple and Quad (2nd): Lowest DR and yield, Highest SPR of

available prenatal screening tests.

What do we do???....

• This screening issue is so confusing to most patients, we offer all the testing but when asked, we give them our suggestion of doing FTS with NB as what we think is the best test.

• We offer Genetic counseling liberally – many people are withholding this option until after they get results of screen. This adds to patient confusion.

Hold on again!

Chromosomal Microarray Analysis (CMA)

• Single test includes all disorders detected by standard cytogenetics and multiple FISH tests

• Probes for virtually all known microdeletion/duplication syndromes and all telomeres are included

• Major advance in the diagnosis of patients with suspected genetic etiology

• Can expand prenatal diagnosis from karyotype to numerous other conditions caused by small deletions and duplications, not visible on routine chromosome analysis

NON –INVASIVE FISH TESTING

• Harvest fetal trophoblastic cells from cervix with cytobrush!!!!!

• Non-invasive • Fish results quickly• NOT A KARYOTYPE AS YET• PRETTY COOL SCREENING TEST BUT….• Controversial issues!!!!!

WOW!!!Are we in exciting times or what…..

Thanks for your attention!!!