IN THE NAME OF GOD First trimester screening for aneuploidy Dr.parichehr pooransari perinatalogist SHOHADA hospital of shahid beheshti university

Why to diagnose prenatally? Is better to know! preparing family for dealing with anomaly planning time and place of delivery Multideciplinary mx In-utero treatment Termination of pregnancy Prematurity and congenital anomalies are the most common causes of neonatal death

How to Diagnose? Diagnostic Procedures Therapeutic Procedures Screening Procedures Diagnostic Procedures Therapeutic Procedures

Screening procedure Screening is intended to identify populations who have an increased risk for a specific disorder, and for whom diagnostic testing may be warranted.

Screening by Sonography Confirmation of gestatioanl age Confirmation of viability Detection of number of fetus and chorionicity Detection of fetal anomalies Detection of high risk groups for chromosomal abnormalities Detection of placental site Detection of amniotic fluid volume Detection of fetal growth Detection of fetal Well-being Evaluation of fetus and placental blood flow Detection of fetal anemia ( Isoimmunization),…

Screening by biochemical markers Screening and diagnosis of maternal anemia Screening and diagnosis of GDM Screening of maternal diseases ( Hypothyroidism,…) Screening of chromosomal abnormalities Screening and diagnosis of fetal infections Screening of fetal anemia Screening of preterm birth Screening of Intrauternine growth restriction and SGA Screening for macrosomia Screening for miscarriage and stillbirth Screening for preeclampsis Screenongfor neural tubal defects

Screening for Chromosoaml abnormalities First trimester Versus Second trimester Sonography Biochemistry Combined sonography and biochemistry

Assessment of Risk Maternal age 0.0001 0.001 0.01 0.1 1 10 20 25 30 35 40 44 Years Risk % Trisomy 21 Trisomy 18 Trisomy 13 xxx/xxy/xyy 45x Triploidy For every case of trisomy 21 there is one with another defect The risk for trisomies increases with maternal age The risk for sex chromosome defects and triploidy does not change with maternal age

Measurement of Nuchal Translucency Gestation 11-13+6 wks CRL 45-84 mm Mid-sagittal view Image size: calipers 0.1mm Neutral position Away from amnion Maximum lucency Callipers on-to-on More than one measurement Umblical cord around the neck

Technique: maximum lucency Screening for Trisomy 21 Technique: maximum lucency 200 400 600 800 1000 1200 1400 1 2 3 4 5 Nuchal translucency (mm) Risk (1 in) 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 45 55 65 75 85 Crown-rump length (mm) NT (mm) 2.7mm 2.0mm 1.5mm 1.6mm 1.2mm 0.9mm 3.5mm 1 in 1000 1 in 100

Nasal bone

Tricuspid valve Doppler Big Magnification Apical 4-chamber Angle < 30º Gate: 2.5 - 3 mm Tricuspid regurgitation Half systole Velocity > 60 cm/s

Advantages of first trimester scan To confirm pregnancy & viability To confirm GA & Dating if needed To confirm the number of gestational sacs, fetuses and chorionicity To check fetal anatomy for structural anomalies To measure nuchal translucency / screen for chromosomal abnormality To check uterus & adnexa

First trimester biochemical markers Free beta hCG Pregnancy associated plasma protein A

Biochemical markers 100 Screening in the first trimester by a combination of maternal age, fetal NT, FHR and serum free ß-hCG and PAPP-A identifies about 90% of trisomy 21 pregnancies for a false positive rate of 3% NT FHR ß-hCG PAPP-A 90% 90 80 ß-hCG AFP E3 IA 71% hCG AFP E3 IA 67% 70 ß-hCG AFP E3 65% 61% ß-hCG AFP hCG AFP E3 60% hCG AFP 56% 60 Detection rate (%) 50 40 30 20 10

Maternal serum free ß-hCG& PAPP-A at 11-13+6 wks 20 Euploid n 2.0 In trisomy 21 pregnancies maternal serum free ß-hCG is about twice as high and PAPP-A is reduced to about half compared to chromosomally normal pregnancies. Trisomy 21 10 0.4 1.2 2.8 3.6 4.4 4.8 Maternal serum free ß-hCG (MoM) The performance of screening for trisomy 21 by maternal age and serum free ß-hCG and PAPP-A is: Detection rate 65% False positive rate 5% 8 n Euploid 4 0.2 0.4 0.6 0.8 1.0 1.2 1.4 1.6 1.8 Maternal serum PAPP-A (M0M)

Maternal serum free ß-hCG& PAPP-A at 11-13+6 wks Concentration of free ß-hCG and PAPP-A is influenced by: The machine and reagents Gestational age Maternal weight Ethnicity Smoking status Method of conception HyperemesisGravidarum Twin ( Chorionicity ,vanishing) Temperature: In first-trimester screening for aneuploidies, analysis of blood samples should be undertaken within a few minutes of collection, otherwise the samples should be refrigerated at 4 degrees C throughout the interval between collection and analysis

Maternal serum free ß-hCG& PAPP-A at 11-13+6 wks In trisomy 21 pregnancies: Free ß-hCG is higher than in euploid pregnancies and the difference between the two is higher at 13 than at 11 weeks Serum PAPP-A is lower than in euploid pregnancies and the difference between the two is higher at 11 than at 13 weeks The difference from euploid pregnancies in PAPP-A at 11 weeks is greater than the difference in ß-hCG at 13 weeks and therefore the overall performance of biochemical screening is better at 11 than at 13 weeks 5.0 ß-hCG Euploid 1.0 PAPP-A Median MoM 0.5 0.1 75 80 85 90 95 100 Gestation (days)

Screening for trisomy21 by fetal NT & FHR and serum free ß-hCG& PAPP-A at 11-13+6 wks Nuchal translucency (mm) 1 2 3 4 5 Euploid Gestation (days) 75 80 85 90 95 100 Trisomy 21 In trisomy 21 compared to euploid pregnancies: The difference in biochemical markers is greater at 11 than at 13 weeks The difference in fetal NT is greater at 11 than at 13 weeks Therefore the overall performance of screening is better at 11 than at 13 weeks 5.0 ß-hCG Euploid 1.0 PAPP-A Median MoM 0.5 0.1 Gestation (days) 75 80 85 90 95 100

Screening for trisomy21 by fetal NT & FHR and serum free ß-hCG& PAPP-A at 11-13+6 wks The overall performance of combined screening is better at 11 than at 13 weeks and may be best at 10 weeks Ultrasound scanning for fetal abnormalities is better at 12 than at 11 weeks and much better than at 10 weeks A good way of achieving a high performance of screening for trisomy 21 and diagnosing major fetal defects by ultrasound is to carry out the blood test at 10 or 11 weeks and the ultrasound scan at 12 weeks Ultrasound scan at 12 wks 96 10 92 88 11 Detection rate for 3% FPR (%) 84 12 80 76 Blood test Gestation (wks)

Chromosomal defects other than trisomy 21 Free ß-hCG PAPP-A Trisomy 18  0.3 MoM Trisomy 13 Turner’s  1.0 MoM Triploidy  /  0.1/10.0 MoM 0.1/1.0 MoM Trisomy 21 2.0 MoM 0.5 MoM

Screening for trisomies 18 and 13 by fetal NT & FHR and serum free ß-hCG& PAPP-A at 11-13+6 wks Trisomies 18 and 13 are the second and third most common chromosomal abnormalities after trisomy 21 .At 11 -13+6 weeks the relative prevalence of trisomies 18 and 13 to trisomy 21 are about 1 to 2.5 and 1 to 7, respectively 100 90 Trisomy 21 80 Trisomy 13 Trisomy 18 70 60 All three trisomies are associated with increased maternal age, increased fetal NT and decreased maternal serum PAPP-A A beneficial consequence of first-trimester combined screening for trisomy 21 is the early diagnosis of trisomies 18 and 13. At a false positive rate of 3% the detection rate of trisomy 21 is 90% and of trisomies 18 and 13 is about 75% 50 Detection rate at 3% FPR (%) 40 30 20 10

Screening for trisomies 18 and 13 by fetal NT & FHR and serum free ß-hCG& PAPP-A at 11-13+6 wks There are differences between the three trisomies: Fetal NT is higher in trisomies 18 and 13 than in trisomy 21 Serum PAPP-A is lower in trisomies 18 and 13 than in trisomy 21 Serum free ß-hCG in trisomy 21 is high whereas in trisomies 18 and 13 this is low Fetal heart rate in trisomy 13, unlike trisomies 21 and 18, is high The use of specific algorithms for trisomy 18 and trisomy 13, in addition to the use of the algorithm for trisomy 21, improves the detection of trisomies 18 and 13 from about 75% to 95% with a minor increase in the total false positive rate from 3% to 3.1% 10 20 30 40 50 60 70 80 90 100 Detection rate at 3% FPR (%) Trisomy 21 Trisomy 18 Trisomy 13

Selected strategies for screening Depends on: Availability of screening test Availability of well trained and certified sonographer Availability of good equipped Laboratory with well trained technician and Lab. Team Cost Availability of diagnostic test Low false positive

Screening strategies NT 1st Trimester 64-70 79-84 90-95 60-69 67-81 Strategy Analytes Detection rate (%) 1st Trimester NT 64-70 NT+ PAPP-A, hCG or free beta-hCG 79-84 NT+ PAPP-A, hCG or free beta-hCG + New markers 90-95 2nd Trimester, Triple test MSAFp, hCG or free l3-hCG, uE3 60-69 2nd trimester , Quadruple (Quad) test MSAFp, hCG or free l3-hCG, uE3, inhibin 67-81

A screen-positive test result indicates that the woman's risk of having a child with Down syndrome is equal to, or exceeds, a specific cut-off level This cut-off level is program/laboratory-specific and based on the combination of markers used, among other factors. A typical cut-off for the combined test is the term risk of Down syndrome of ≥1 in 300. This is associated with a false positive rate (FPR) of about 5 percent (eg, about 5 per 100 women will have risks higher than 1 in 300 – such as 1 in 250 or 1 in 75). The odds of Down syndrome in this group of women is about 1 in 20 (eg, for every 20 women screen-positive, 1 will have a Down syndrome fetus and 19 will have a normal fetus)

First-trimester combined and stepwise sequential testing should not be offered if CVS is unavailable . Alternatively, they may choose to undergo secondary screening using a maternal plasma-based test for cell free DNA. Although not diagnostic, the high sensitivity and specificity of the DNA test will reassure many of these women that their risk of having a Down syndrome pregnancy is very low, which allows them to avoid the risk associated with an invasive diagnostic test. Among those women with a positive DNA test, the risk of Down syndrome is increased, but still requires confirmation by an invasive diagnostic test (eg, amniocentesis to obtain amniocytes for karyotyping).

Screen-negative first-trimester or integrated test results — A negative test result means the patient's risk of having a baby with Down syndrome is less than a specified cut-off level; it does not exclude the possibility of Down syndrome. The patient's risk is usually provided in the report (eg, Down syndrome risk 1 in 9000) and this number can be given to the patient with a discussion of its meaning. With regard to Down syndrome screening, no further testing is recommended.

Conclusion Screening for chromosomal defects should be offered to all pregnant ladies who attend at < 20 weeks Proper counselling must be done with parents Screening strategy must be chosen accordning to: Gestational age Available certified sonographer Availabe certified Laboratory Affordable cost Risk and counselling after result must be done accurately by physician NOT Laboratory or even sonographer unless sonographer is perinatalogist The final decision must be from parents not the physician which is possible if counselling is effective.

…Conclusion NT is the best marker As a single marker IF… Combination of NT with biochemistry will increase the detection rate and may decrease false positive rate Training programs for sonographers ( www. Fetalmedicine.com) Increase the number of certified Laboratory with proper softwares , machines and trained technician) Establish multicenter studies to find out our normal ranges ( MoMs)