Chapter 17Prenatal Diagnosis of Congenital Disorders Ronald J. Wapner, MD, Thomas M. Jenkins, MD, and Nahla Khalek, MDPrenatal diagnosis, a term once considered synonymous with invasive the disease on the positive and negative predictive values is describedfetal testing and karyotype evaluation, now encompasses pedigree in Chapter 16 and is shown in Tables 16-6 and 16-7.analysis, population screening, fetal risk assessment, genetic counsel- Use of screening tests requires that cutoff values for “positive” testsing, and fetal diagnostic testing. Although prenatal evaluation of the be set. Performance of the test depends on this cutoff; for example,fetus for genetic disorders can have a huge impact on individual fami- increased detection rate can be obtained by lowering the cutoff thresh-lies, most screening and testing is done for events that occur in less old, but the concomitant lowered speciﬁcity would result in morethan 1% of pregnancies. In this chapter, we describe different modali- false-positive results. Table 17-1 shows the performance of second-tri-ties available for in utero fetal diagnosis of congenital disorders, the mester maternal serum screening for Down syndrome based on variousapproach to screening ongoing pregnancies for genetic disease, and the cutoffs.1 A receiver operating curve can be used as a statistical methodcounseling requirement for each. to ﬁnd the “best” balance between sensitivity and speciﬁcity. A line diagram is plotted with sensitivity on the vertical axis and the false- positive rate plotted horizontally (Fig. 17-1). The greater the area under the curve (toward the upper left corner), the better the test’sScreening for Fetal performance is, with increasing sensitivity and a reduced false-positiveGenetic Disorders rate. When screening for Down syndrome, cutoff values are importantDetecting or deﬁning risk for disease in an asymptomatic low-risk for laboratories that provide the testing and for clinicians who inter-population is the goal of screening. As opposed to diagnostic testing, pret the results. When viewed from the patient’s perspective, reportingintended to identify or conﬁrm an affected individual, screening is tests as positive or negative can be confusing. Receipt of a “positive”intended to identify populations who have an increased risk for a spe- result of 1 in 250 may lead to a choice of a diagnostic test that carriesciﬁc disorder, and for whom diagnostic testing may be warranted. An a risk of complications, whereas a “negative” result of 1 in 290 mayideal perinatal genetic screening test should fulﬁll the following provide greater reassurance than intended, when in fact the actual riskcriteria: of Down syndrome is similar for both patients. Often, explaining the signiﬁcance of a positive or negative result before the screening test is Identify common or important fetal disorders performed will assist patients in understanding their results. Many Be cost-effective and easy to perform centers report the absolute risk to the patient to further help in inter- Have a high detection rate and a low false-positive rate pretation. Regardless of the counseling approach, understanding the Be reliable and reproducible concept of screening is difﬁcult for many patients. From the perspec- Test for disorders for which a diagnostic test exists tive of the laboratory or clinician, selection of a cutoff that is too high Be positive early enough in gestation to permit safe and legal or too low will lead to overutilization or underutilization of diagnostic options for pregnancy termination if desired tests and the consequent risk of procedure-related pregnancy loss or false reassurance, respectively. Sensitivity and speciﬁcity are two key concepts in screening testperformance (see Chapter 16). Sensitivity is the percentage of affectedpregnancies that are screen positive. Speciﬁcity is the percentage of Likelihood Ratiosindividuals with unaffected pregnancies who screen negative. The The impact of a positive screening test depends on the pretest (a priori)reciprocal of speciﬁcity is the false-positive rate. Sensitivity and speci- risk of an affected pregnancy. Likelihood ratios are statistical meansﬁcity are independent of disease frequency, and they describe the to modify an individual’s risk based on known data for a population.anticipated performance of a screening test in the population. Alter- For binary risk factors that are either present or absent, likelihoodnatively, positive and negative predictive values are dependent on ratios are determined by comparing the frequency of positive tests indisease prevalence and are vital in the interpretation of the test result affected pregnancies to the frequency in normal pregnancies. This isfor an individual patient. These latter two values represent, respectively, calculated as the sensitivity of the test divided by its false-positive ratethe likelihood that a person with a positive or negative test does or (likelihood ratio = sensitivity/false-positive rate). For tests that usedoes not have an affected pregnancy. The impact of the prevalence of continuous variables (such as serum marker measurements), likeli-
222 CHAPTER 17 Prenatal Diagnosis of Congenital Disorders TABLE 17-1 DOWN SYNDROME DETECTION: TABLE 17-2 ADJUSTING THE RISK FOR DOWN FALSE-POSITIVE RATES (FPR) AT SYNDROME USING LIKELIHOOD DIFFERENT CUTOFF VALUES RATIOS FOR BINARY ULTRASOUND MARKERS Triple Quadruple Screen Detection Screen Detection A priori risk of Down syndrome (age or serum screen) 1 : 1000 Cutoff Rate FPR Cutoff Rate FPR Positive ultrasound marker for Down syndrome Rate in Down syndrome population (sensitivity 10% 1 : 200 57 4.3 1 : 200 60 3.5 of marker) 1 : 250 61 5.6 1 : 250 64 4.5 Rate in general population (FPR of marker) 1.0% 1 : 300 64 6.8 1 : 300 67 5.5 Likelihood ratio (sens/FPR) 10.0 1 : 350 67 8.1 1 : 350 69 6.5 Adjusted risk for Down syndrome 1 : 400 69 9.3 1 : 400 72 7.6 (A priori risk × likelihood ratio) = 1/1000 × 10 1 : 100 Data from Huang T, Watt H, Wald N: The effect of differences in the FPR, false-positive rate; sens, sensitivity. distribution of maternal age in England and Wales on the performance of prenatal screening for Down’s syndrome. Prenat Diagn 17:615, 1997. level and subsequent public policy was determined more than 25 years 100 ago and was based on a maternal age risk of 35 years at delivery. The Rate of detection of Down syndrome (%) 90 factors considered in determining this value included the prevalence 80 of disease, a perceived signiﬁcant increase in the trisomy 21 risk after this age, the risk of invasive testing, the availability of resources, and 70 a cost-to-beneﬁt analysis. Since that time, a number of additional 60 screening tests for Down syndrome have become available that chal- lenge the validity of maternal age as a single indication for invasive 50 testing. 40 30 Maternal Age as a Screening Test 20 The association of Down syndrome with advancing maternal age was 10 ﬁrst reported in 1909.2 Fifty years later, karyotype analysis was devel- oped and correlated the Down syndrome phenotype with an extra G 0 0 10 20 30 40 50 60 70 80 90 100 chromosome.3 This led to the development of genetic amniocentesis, False-positive rate (%) through which the prenatal diagnosis of Down syndrome became fea- sible. To standardize the use of this emerging technology, a consensusFIGURE 17-1 Receiver operating curve demonstrating sensitivity report from the National Institutes of Health in 1979 suggested thatfor Down syndrome detection versus false-positive rate. Cutoff amniocentesis be routinely offered to women aged 35 years or older atpoints for sensitivity versus false-positive rate are determined by delivery. At that time, maternal age risks of Down syndrome werethese curves. (From Haddow JE, Palomaki GE, Knight GJ, et al: available only in 5-year groupings. Using these data, the age of 35Reducing the need for amniocentesis in women 35 years of age or seemed a natural cutoff, because women in the 30- to 34-year groupingolder with serum markers for screening. N Engl J Med 330:1114, had a risk of 1 : 880, and the risk for women aged 35 to 40 was almost1994.) fourfold higher. This cutoff was based on a number of factors, includ- ing the availability of experienced operators and cytogenetics labora- tories, the cost-to-beneﬁt ratio, and the balance betweenhood ratios are calculated from the log gaussian distributions of procedure-related losses and the possibility of a positive ﬁnding. Thisnormal and affected pregnancies. Once a likelihood ratio is deter- cutoff continues to be used, and the second-trimester risk of 1 : 270 ormined, it can be used to modify the a priori risk (Table 17-2). If more liveborn risk of 1 : 380 remains the standard value for offering womenthan one likelihood ratio is available and is independent of other invasive testing.parameters, it also can be used to modify risk. In this way, multiple The risk for Down syndrome is now recognized to be continuous,factors (such as maternal age, serum analytes, and ultrasound ﬁndings) which emphasizes the arbitrary nature of an absolute age threshold ofcan be simultaneously used to modify risk. 35. In addition to maternal age, the risk of trisomy 21 depends on the gestational age at which testing is performed, because only 69% of ﬁrst-trimester and 76% of second-trimester Down syndrome pregnan- cies are viable (Table 17-3).4 Currently, more than 80% of diagnostic procedures to determine fetal karyotype performed in the UnitedAntenatal Screening for States are performed solely for “advanced maternal age,” yet 70% ofDown Syndrome affected pregnancies are born to women outside this group. With many women delaying childbirth until later in life, more than 14% of preg-There has been a general consensus in the United States that invasive nant women in the United States are being offered testing based on agetesting for Down syndrome should be offered to women with a second- alone. In Switzerland and the United Kingdom, almost 15% of birthstrimester risk of 1 : 270 or higher (liveborn risk of 1 : 380). The cutoff now occur in women older than 35.
CHAPTER 17 Prenatal Diagnosis of Congenital Disorders 223 TABLE 17-3 RISK FOR DOWN SYNDROME are elevated in Down syndrome pregnancies (1.3 to 2.5 MoM) and do BASED ON MATERNAL AND not vary with gestational age in the second trimester. There is, however, a small correlation with hCG levels, making the added sensitivity for GESTATIONAL AGES Down syndrome detection less robust.19 Detection rates when a quad Gestational Age screen of α-fetoprotein (AFP), hCG, unconjugated estriol, and inhibin Maternal Age A are used are about 75% (screen positive rate, 5%) in the population (yr) 12 Wk 16 Wk 20 Wk Liveborn under age 35 years.20 For women older than 35, the detection rate is 20 1/1068 1/1200 1/1295 1/1527 approximately 92% with a screen-positive rate of 13%. 25 1/946 1/1062 1/1147 1/1352 Other analytes or combinations of analytes have been tested to 30 1/626 1/703 1/759 1/895 further increase sensitivity. Hyperglycosylated hCG excreted in mater- 31 1/543 1/610 1/658 1/776 nal urine has been tested as a marker for Down syndrome. One study 32 1/461 1/518 1/559 1/659 of nearly 1500 women (1448 control subjects and 39 Down syndrome 33 1/383 1/430 1/464 1/547 34 1/312 1/350 1/378 1/446 pregnancies) reported a sensitivity of 96% of affected pregnancies with 35 1/249 1/280 1/302 1/356 a 5% false-positive rate and 71% detection with a 1% false-positive 36 1/196 1/220 1/238 1/280 rate when a combination of hyperglycosylated hCG, urine β-core hCG 37 1/152 1/171 1/185 1/218 fragment, MSAFP, and maternal age was used.21 This detection rate, 38 1/117 1/131 1/142 1/167 however, has not been duplicated by others. 39 1/89 1/100 1/108 1/128 With the addition of extra markers, the potential beneﬁt must be 40 1/68 1/76 1/82 1/97 balanced against the cost. With each additional marker, costs to society 42 1/38 1/43 1/46 1/55 can balloon into the millions because of the number of pregnancies 44 1/21 1/24 1/26 1/30 tested each year with only a minimal improvement in detection. The 45 1/16 1/18 1/19 1/23 relative cost and value of raising the sensitivity or lowering the false- Data from Hook EB: Rates of chromosome abnormalities at different positive rate a few percentage points is an ongoing debate. maternal ages. Obstet Gynecol 58:282, 1981. Abnormal Second Trimester Maternal Serum Markers in Pregnancies with aSecond-Trimester Maternal Normal KaryotypeSerum ScreeningSerum screening may be used to identify the 70% of Down syndrome Unexplained Elevated Maternalpregnancies in women less than 35 years of age. This approach is Serum a-Fetoproteinderived from a 1984 report of lower maternal serum α-fetoprotein When an elevated MSAFP is reported in pregnancies in which the ges-(MSAFP) levels in women carrying a Down syndrome fetus. Women tational age is correctly assigned and the fetus is structurally normal,with Down syndrome pregnancies had a median MSAFP value of 0.75 and the amniotic ﬂuid AFP (AFAFP) is normal, the biologic explanationmultiples of the unaffected median (MoM).5,6 Using this deviation to is almost always a breach in the maternal-fetal interface. This leads tocalculate a likelihood ratio, the age-related risk for Down syndrome higher AFP levels in the maternal circulation. Not surprisingly, womencould be modiﬁed. When the standard 1 : 270 cutoff was used, approxi- with unexplained elevation in AFP levels have been found to havemately 25% of Down syndrome pregnancies among women less than increased risk of obstetric complications, including fetal growth restric-35 years of age were screen-positive.7-9 tion, fetal death, prematurity, oligohydramnios, abruptio placentae, and Elevated human chorionic gonadotropin (hCG) (mean, 2.3 MoM) preeclampsia. Table 17-4 summarizes the numerous reports; the higherand reduced levels of unconjugated estriol (mean, 0.7 MoM) were the MSAFP level is, the greater the risk. Crandall and colleagues22subsequently linked to an increased risk of trisomy 21.6,10,11 hCG or studied 1002 women with MSAFP values greater than 2.5 MoM andreduced levels of unconjugated estriol used alone to modify the mater- stratiﬁed them by the degree of elevation. In those with a normal ultra-nal age risk has a Down syndrome detection rate of only 20% to 30%. sound and amniocentesis, the risk of adverse outcome was 27% overallHowever, because they are independent variables, they can be analyzed but varied with the degree of elevation. Adverse outcome occurred insimultaneously with maternal age and AFP to form a composite risk 16% when the MSAFP was 2.5 to 2.9 MoM, 29% when it was 3.0 tocalculation (frequently called a triple screen). 5.0 MoM, and 70% when it was greater than 5.0 MoM. Waller and The sensitivity of the triple screen for Down syndrome detection coworkers23-25 investigated 51,008 women screened with MSAFP inin women younger than 35 years ranges between 57% and 67% if the California to evaluate the predictive value of high MSAFP comparedfalse-positive rate is held constant at 5%.12-15 Overall, the odds of with low levels. The risk of delivery before 28 weeks was 0.4% with lowhaving an affected pregnancy with a positive screen are approximately values (<0.81 MoM) and 3.2% for those with high values (>2.5 MoM),1 in 33 to 1 in 62, depending on the age range of the population an eightfold difference. The rates for delivery before 37 weeks were 2.6%studied,16,17 an improvement over the 1 : 100 odds when maternal age for the low MSAFP group and 24.3% for the high MSAFP group.is the sole screening parameter. Because of the impact of maternal age Notably, women with MSAFP values greater than 2.5 MoM had a 10.5-on the risk analysis, screening women who will be 35 years of age or fold increase in preeclampsia and a 10-fold increased risk of placentalmore increases the sensitivity using the same cutoffs to approximately complications, suggesting that an elevated value in the absence of an87% but with a false-positive rate of nearly 25%.13,18 anomaly may derive from a fetal-maternal hemorrhage of sufﬁcient Inhibin A, a protein produced initially by the corpus luteum and volume to have clinical signiﬁcance. This observation may explain thelater by the placenta, is now routinely included in second-trimester elevation in MSAFP, but to date no management protocol has beenDown syndrome screening resulting in a “quad screen.” Inhibin A levels demonstrated to improve outcome in these cases.
224 CHAPTER 17 Prenatal Diagnosis of Congenital Disorders TABLE 17-4 STUDIES EVALUATING THE RELATION OF UNEXPLAINED ELEVATIONS OF MSAFP AND POOR PREGNANCY OUTCOME Pregnancies MoM IUGR Premature Abruption IUFD Perinatal Source Location (Year) Screened Cutoff LBW Risk Risk Delivery Risk Risk Risk Death Brock et al (505, 506) Scotland (1977, 15,481 2.3 2.5× + + 1979) Wald et al (507, 508) England (1977, 3,194 3.0 4.7× 5.8× 3.5× 1980) 4,198 Macri et al (509) New York (1978) 6,031 2.0 2.0× Gordon et al (510) England (1978) 1,055 2.0 3.5× 4.5× Smith (511) England (1980) 1,500 2.0 + + + + Evans and Stokes Wales (1984) 2,913 2.0 3.0× + 8.0× (512) Burton et al (513, 514) North Carolina 42,037 2.5 2.0× 8.0× 10.0× (1983, 1988) Persson et al (515) Sweden (1983) 10,147 2.3 2.8× 2.0× 10.0× 3.0× Haddow et al (516) Maine (1983) 3,636 2.0 3.6× 2.0× Purdie et al (517) Scotland (1983) 7,223 2.5 2.5× 20.0× Fuhrmann and Weitzel West Germany 50,000 2.5 3.5× 8.6× (518) (1985) Williamson et al (519) Iowa (1986) 1,161 Poor outcomes Robinson et al (520) California (1989) 35,787 2.0 3.5× Ghosh et al (521) Hong Kong (1986) 9,838 2.0 + Schnittger and Kjessler Sweden (1984) 18,037 2.0 + + (522) Hamilton et al (523) Scotland (1985) 10,885 2.5 10.0× 2× >10.0× 3.0× 8.0× Doran et al (524) Ontario (1987) 8,140 2.0 6.0× + Milunsky et al (525) Massachusetts 13,486 2.0 4.0× 3.0× 8.0× + (1989) IUFD, intrauterine fetal demise; IUGR, intrauterine growth retardation; LBW, low birth weight; MoM, multiple of the median; MSAFP; maternal serum a-fetoprotein; +, increased risk but unquantiﬁed. Data from Milunsky A (ed): Genetic Disorders and the Fetus: Diagnosis, Prevention, and Treatment, 3rd ed. Baltimore, Johns Hopkins University Press, 1992, p 656. 0.0 to 0.15 MoM suggest biochemical abnormalities of the fetusUnexplained Elevated Human Chorionic or placenta, including placental steroid sulfatase deﬁciency, Smith-Gonadotropin Levels Lemli-Opitz syndrome, congenital adrenal hypoplasia, adrenocor-The risk for adverse pregnancy outcome with elevated hCG levels ticotropin deﬁciency, hypothalamic corticotropin deﬁciency, andappears to be independent of those associated with elevated AFP. anencephaly.Studies have shown that unexplained elevated hCG (greater than Smith-Lemli-Opitz syndrome is an autosomal recessive disorder2.0 MoM) is associated with an increased risk of preeclampsia, preterm resulting from a defect in 3αβ-hydroxysteroid-α7-reductase, alteringbirth, low birth weight, fetal demise, and possibly hypertension.26 It cholesterol synthesis and resulting in low cholesterol levels and theappears that the higher the hCG is, the greater the risk. accumulation of the cholesterol precursor 7-dehydrocholesterol in blood and amniotic ﬂuid. Because cholesterol is a precursor of estriol,Elevated Human Chorionic Gonadotropin and the defect results in reduced or undetectable levels of estriol in mater-Maternal Serum a-Fetoprotein nal serum and amniotic ﬂuid. Smith-Lemli-Opitz syndrome is charac-The combination of elevated MSAFP and hCG levels occurs rarely but terized by low birth weight, failure to thrive, and moderate to severemay have an overall pregnancy complication rate exceeding 50%. A mental retardation. It is associated with multiple structural anomalies,study of 66 singleton and 33 multiple pregnancies with an MSAFP of including syndactyly of the second and third toes, microcephaly, ptosis,more than 2 MoM and an hCG of more than 3.0 MoM found that 60% and a typical-appearing facies.32-34 Undermasculinization of the gene-of singletons and 81% of twins had at least one of several obstetric talia including complete sex reversal can be seen in male fetuses.complications, including preeclampsia, preterm birth, growth restric- Bradley and colleagues35 summarized ﬁndings in 33 women whotion, placental abnormalities, and fetal death.27 Conﬁned placental delivered infants with Smith-Lemli-Opitz syndrome. Twenty-four ofmosaicism for chromosome 16 has been reported to be associated with 26 women who had second-trimester estriol values obtained had levelsextremely high levels of both analytes, as well as with similarly poor less than the 5th percentile (<0.5 MoM). The median level in thisoutcomes.28,29 group was 0.23 MoM (below the ﬁrst percentile). A risk assessment based on low maternal serum unconjugated estriol levels has beenLow Second-Trimester Maternal Serum Estriol suggested but is not presently available.36 Reliable and inexpensiveLow maternal serum unconjugated estriol levels have been linked to prenatal testing for Smith-Lemli-Opitz syndrome based on amnioticadverse pregnancy outcomes.30,31 Very low or absent estriol levels of ﬂuid cholesterol or 7-dehydrocholesterol levels is available.37
CHAPTER 17 Prenatal Diagnosis of Congenital Disorders 225 Placental steroid sulfatase deﬁciency is an X-linked recessive disor- TABLE 17-5 SECOND-TRIMESTERder resulting from deletion of Xp22.3. This enzyme deﬁciency prevents ULTRASOUND MARKERSremoval of the sulfate molecule from fetal estrogen precursors, pre- ASSOCIATED WITH DOWNventing conversion to estriol. The fetal phenotype depends on the SYNDROMEextent of the deletion, with greater than 90% of cases presenting as X-linked ichthyosis that can be treated with topical keratolytic agents. BrachycephalyHowever, in about 5% of cases, there can be a deletion of contiguous Increased nuchal thicknessgenes causing mental retardation. The deletion can, on occasion, Congenital heart defectsextend to cause Kallman syndrome or chondrodysplasia punctata. The Hyperechoic bowellack of estrogen biosynthesis may result in delayed onset of labor, Shortened femurprolonged labor, or stillbirth. Shortened humerus Renal pyelectasis Prenatal diagnosis for the deletion leading to placental sulfatase Duodenal atresiadeﬁciency and congenital ichthyosis can be performed by identifying Hypoplasia of the midphalanx of the ﬁfth digitthe gene deletion by karyotype or ﬂuorescence in situ hybridization.38-40 Echogenic intracardiac focusAlthough very low estriol levels, usually below the level of detection, “Sandal gap” of the footcan identify males at risk for this disorder, testing in these cases is not Widened ischial spine angleroutinely offered because the phenotype is usually mild. However, the Foot lengthrarer, more serious cases of extensive deletions will be missed.41 Short or absent nasal boneSecond-Trimester Ultrasound Markersof Down Syndrome pleted and the results of serum screening have been reported. MarkersThe clinical suspicion of Down syndrome is suspected when an infant commonly sought to assess the risk of Down syndrome include theis found to have speciﬁc physical ﬁndings that occur frequently in following:Down syndrome infants but can also occur in normal individuals.These include a simian crease in the fetal hand, a short femur or 1. An increased nuchal fold (>6 mm) in the second trimester is thehumerus, clinodactyly, and excessive nuchal skin. Similarly, the in utero most distinctive marker. The fetal head is imaged in a transversediagnosis of Down syndrome can be suspected when anomalies or plane similar to that for measuring the biparietal diameter. Thephysical features that occur more frequently in Down syndrome than thalami and the upper portion of the cerebellum should be in thein the general population are noted on an ultrasound examination. plane of the image. The distance between the external surface ofCertain of these congenital anomalies, such as atrioventricular canal the occipital bone and the external surface of the skin is then mea-or duodenal atresia, strongly suggest the possibility of Down syndrome sured. About 35% of Down syndrome fetuses but only 0.7% ofand are independent indications to offer invasive testing. Although normal fetuses have a nuchal skin fold measurement greater thanwhen these anomalies are present there is a high risk of trisomy 21, 5 mm. This ratio yields a likelihood ratio of 50 but includes fetusesthese anomalies have low sensitivity and thus are not useful in screen- with more than one marker. When an increased nuchal fold is aning. For example, when duodenal atresia is identiﬁed, there is an isolated ﬁnding, the likelihood ratio is still strong at 20-fold. Thisapproximate 40% risk of Down syndrome, yet it is seen in only 8% of high likelihood ratio is obtained because of the rarity of an increasedaffected fetuses. Physical characteristics that are not themselves anom- nuchal fold in an unaffected population (i.e., high speciﬁcity). Foralies but that occur more commonly in fetuses with Down syndrome women with an a priori risk of less than 1 : 1600 (age-related riskare called markers. By comparing the prevalence of markers in Down for a 20-year-old), a 20-fold increase results in a risk estimate of atsyndrome fetuses to their prevalence in the normal population, a likeli- least 1 : 270. Thus, the presence of an increased nuchal fold alone ishood ratio can be calculated that can be used to modify the a priori an indication to offer invasive testing.42-47age or serum screening risk. This is the basis for ultrasound screening 2. The fetal nasal bone has been demonstrated to be hypoplastic orfor Down syndrome. absent in up to 60% of Down syndrome pregnancies imaged in the For a marker to be useful for Down syndrome screening, it should second trimester and only about 1% to 2% of unaffected pregnan-be sensitive (i.e., present in a high proportion of Down syndrome cies. Complete absence will occur in about 37% of affected casespregnancies), speciﬁc (i.e., rarely seen in normal fetuses), easily imaged with hypoplasia occuring in about half. In normal pregnancies,in standard sonographic examination, and present early enough in the absence is seen in 0.9% of cases and hypoplasia in 2.4%. Nasal bonesecond trimester that subsequent diagnostic testing by amniocentesis length can be converted to a likelihood ratio and used for Downcan be performed so that results are available when pregnancy termi- syndrome risk assessment. When performed by experienced opera-nation remains an option. A list of available markers and their likeli- tors, nasal bone evaluation may be the best single ultrasound markerhood ratios are seen in Tables 17-5 and 17-6, respectively. for second trimester risk assessment.48 Before considering each marker individually, it is important to 3. Down syndrome fetuses in the second trimester may have shortremember that the predictive value of any test (e.g., a marker) depends proximal extremities (humerus and femur) relative to the expectedon the prevalence in the population of the condition being tested for. length for their biparietal diameter. This can be used to identifyIn the case of sonographic markers for trisomy 21, the clinical impor- at-risk pregnancies by calculating a ratio of observed to expectedtance of a marker, therefore, varies according to the a priori risk as femur length based on the fetus’s biparietal diameter. An observed-determined by maternal age, the results of multiple serum markers, to-expected ratio of less than 0.91 or a biparietal diameter-to-femurand the presence of any other sonographic markers detected at the ratio of more than 1.5 has a reported likelihood ratio of 1.5 to 2.7same examination. It is wise, therefore, to defer discussion of the when present as an isolated ﬁnding. A short humerus is moreimpact of markers until the ultrasound examination has been com- strongly related to Down syndrome, with reported likelihood ratios
226 CHAPTER 17 Prenatal Diagnosis of Congenital Disorders TABLE 17-6 LIKELIHOOD RATIOS (LR) FOR ISOLATED MARKERS IN THREE STUDIES AAURA Nyberg et al. Smith-Bindman et al. LR* LR (95% CI)† LR (95% CI)‡ Sonographic Marker (N = 1042) (N = 8830) (N = meta-analysis of >131,000) Nuchal thickening 18.6 11.0 (5.2-22) 17.0 (8.0-38) Hyperechoic bowel 5.5 6.7 (2.7-16.8) 6.1 (3.0-12.6) Short humerus 2.5 5.1 (1.6-16.5) 7.5 (4.7-12) Short femur 2.2 1.5 (0.8-2.8) 2.7 (1.2-6) Echogenic intracardiac focus 2.0 1.8 (1.0-3) 2.8 (1.5-5.5) Pyelectasis 1.5 1.5 (0.6-3.6) 1.9 (0.7-5.1) *LR assumed by the original age-adjusted ultrasound risk adjustment (AAURA) model by Nyberg DA, Luthy DA, Resta RG, et al: Age-adjusted ultrasound risk assessment for fetal Down’s syndrome during the second trimester: Description of the method and analysis of 142 cases. Ultrasound Obstet Gynecol 12:8, 1998. †Nyberg DA, Souter VL, El-Bastawissi A, et al: Isolated sonographic markers for detection of fetal Down syndrome in the second trimester of pregnancy. J Ultrasound Med 20:1053, 2001. ‡LR of meta-analysis by Smith-Bindman R, Hosmer W, Feldstein VA, et al: Second-trimester ultrasound to detect fetuses with Down syndrome: A meta-analysis. JAMA 285:1044, 2001. CI, conﬁdence interval. ranging from 2.5 to 7.5. Bahado-Singh and coworkers49 combined calculate an age-adjusted ultrasound risk assessment for Down humerus length with nuchal skin fold to estimate Down syndrome syndrome in 8914 pregnancies (186 fetuses with Down syndrome, risk and calculated the likelihood ratios for various measurements 8728 control subjects). Some type of sonographic ﬁnding (major to adjust estimated Down syndrome risk for each patient. abnormality, minor marker, or both) was observed in 68.8% of fetuses4. Echogenic intracardiac foci occur in up to 5% of normal pregnan- with trisomy 21 compared with 13.6% of control fetuses (P < .001). cies and in approximately 13% to 18% of Down syndrome gesta- The observation that about one third of fetuses with Down syndrome tions.50 The likelihood ratio for Down syndrome when an echogenic have neither a marker nor an anomaly has been used to adjust focus is present as an isolated marker has ranged from 1.8 to 2.8. the estimated risk of Down syndrome downward by approximately The risk does not seem to vary if the focus is in the right or left 60% to 65% (likelihood ratio, 0.4) when the “genetic ultrasound” ventricle or if it is unilateral or bilateral. is normal. This sensitivity was observed in a single experienced center.5. Increased echogenicity of the fetal bowel, when brighter than the It is doubtful that the same sensitivity can be achieved in every surrounding bone, has a Down syndrome likelihood ratio of 5.5 to center.61 6.7.51-53 This ﬁnding can also be seen with fetal cystic ﬁbrosis (CF), A positive likelihood ratio can be used to estimate an increase in congenital cytomegalovirus infection, swallowed bloody amniotic risk. The magnitude of the increase depends on the marker(s) or ﬂuid, and severe intrauterine growth restriction. Therefore, if anomalies seen. Nyberg and colleagues reviewed their own data60,62 and amniocentesis is performed for this ﬁnding, testing for the other the data of others63 to estimate a likelihood ratio for each marker as an potential etiologies should be considered. isolated ﬁnding (see Table 17-6). An isolated minor or “soft” marker6. Mild fetal pyelectasis (a renal pelvis anterior-posterior diameter was the only sonographic ﬁnding in 42 (22.6%) of 186 fetuses with greater than 4 mm) has been suggested as a potential marker for trisomy 21, compared with 987 (11.3%) of 8728 control fetuses (P < Down syndrome. As an isolated marker, the likelihood ratio ranges .001). Nuchal thickening, nasal bone hypoplasia, and hyperechoic from 1.5 to 1.9 (see Table 17-6). This has been found by Snijders bowel showed the strongest association with trisomy 21 as isolated and coworkers54 not to be signiﬁcantly more frequent in Down markers, followed by shortened humerus, echogenic intracardiac focus, syndrome pregnancies than in normal pregnancies (i.e., low shortened femur, and pyelectasis. Echogenic intracardiac focus was the speciﬁcity). single most common isolated marker in both affected fetuses (7.1%)7. Other markers described include a hypoplastic ﬁfth middle phalanx and control fetuses (3.9%) but carried a low risk. of the hand,55 short ears, a sandal gap between the ﬁrst and second toes,56,57 an abnormal iliac wing angle,58 and an altered foot-to- Combined Ultrasound and Second- femur ratio.59 These markers are inconsistently used because of the time and expertise required to obtain them. Trimester Maternal Serum Marker Risk Assessment Ultrasound markers can also be combined with serum markers if theyUse of Second-Trimester Ultrasound to are independent. Souter and coworkers64 demonstrated a relativelyEstimate the Risk of Down Syndrome small correlation that needs to be taken into consideration if a quan-As with other screening modalities, second-trimester ultrasound can titative approach is used. Bahado-Singh and colleagues65 combinedbe used to alter the a priori risk in either direction. A benign second- ultrasound markers with maternal analytes, including urinary hyper-trimester scan having none of the known markers and no anomalies glycosylated hCG and urinary α-core fragment of hCG. In a sample ofhas been suggested to have a likelihood ratio of 0.4, assuming the image 585 pregnancies, the sensitivity was 93.7%, with a false-positive rate ofquality is satisfactory. Nyberg and coworkers60 used this approach to 5%.
CHAPTER 17 Prenatal Diagnosis of Congenital Disorders 227 TABLE 17-7 ASSOCIATION OF ULTRASOUND MARKERS WITH ANEUPLOIDY Isolated Multiple Ultrasound Finding (%) (%) Trisomy 13 Trisomy 18 Trisomy 21 Other 45X Holoprosencephaly 4 39 30 7 — 7 — n = 132 Choroid plexus cysts 1 46 11 121 18 11 — n = 1806 Facial cleft 0 51 25 16 — 6 — n = 118 Cystic hygroma 52 71 — 13 26 11 163 n = 276 Nuchal skin fold 19 45 — 9 85 19 10 Diaphragmatic hernia 2 34 — 18 — 14 — n = 173 Ventriculomegaly 2 17 10 23 13 14 — n = 690 Posterior fossa cyst 0 52 10 22 — 8 — n = 101 Major heart defects 16 66 30 82 68 31 30 n = 829 Duodenal atresia 38 64 — — 21 2 — n = 44 Hyperechoic bowel 7 42 — — 22 17 — n = 196 Omphalocele 13 46 28 108 — 31 — n = 475 Renal anomalies 3 24 40 52 48 62 — n = 1825 Mild hydronephrosis 2 33 8 6 27 9 — n = 631 Intrauterine growth restriction (early) 4 38 11 47 — 18 36 (triploidy) n = 621 Talipes 0 33 — — — — — n = 127 Isolated, isolated ﬁnding; multiple, multiple ﬁndings on ultrasound. Adapted from Snijders RJM, Nicolaides KH: Ultrasound Markers for Fetal Chromosomal Defects. New York, Parthenon, 1996.Second-Trimester Ultrasound First-Trimester Ultrasound ScreeningScreening for Other for AneuploidyChromosomal Abnormalities In his initial description of the syndrome that bears his name, LangdonFetal aneuploidy other than Down syndrome can be suspected based Down described skin so deﬁcient in elasticity that it appeared to be tooon ultrasound ﬁndings (Table 17-7). Choroid plexus cysts occur in 1% large for the body. This was particularly noticeable in the neck area.of fetuses between 16 and 24 weeks’ gestation and have been associated The skin in the fetal neck can now be seen with ultrasound at as earlywith trisomy 18. Thirty percent to 35% of fetuses with trisomy 18 have as 10 to 12 weeks of gestation and is known as a nuchal translucencychoroid plexus cysts. Among fetuses with a choroid plexus cyst, about (NT). The quantiﬁcation of this additional “skin behind the neck” can3% have trisomy 18, most (65% to 90%) of whom have other ultra- be used for ﬁrst-trimester Down syndrome screening.72sound ﬁndings (Table 17-8). Although an isolated choroid plexus cyst The NT is a ﬂuid-ﬁlled space in the posterior fetal nuchal area. NTwas estimated to yield a probability of trisomy 18 of 1 of 150 in one is deﬁned as a collection of ﬂuid under the skin behind the neck inreview, many of the series reviewed contained a high proportion of fetuses between 11 and 14 weeks’ gestation. This can be successfullyolder women, which would overstate the risk. Snijders and coworkers66 measured by transabdominal ultrasound examination in approxi-calculated that an isolated choroid plexus cyst has a likelihood ratio mately 95% of cases.for trisomy 18 of 1.5 and can be used to calculate an individual’s risk Studies conducted in women with increased risk of aneuploidyfor trisomy 18. The size, location, or persistence of the cyst does not demonstrated an association between increased NT and chromosomalalter this risk.67-71 defects.73-90 Subsequent studies demonstrated that an NT thickness Table 17-7 displays the magnitude of the associations between above the 95th percentile was present in approximately 80% of trisomyvarious ultrasound ﬁndings and aneuploid conditions as estimated 21 fetuses.89 As with other serum and ultrasound markers, the signiﬁ-from a referral population. The rates noted may overestimate the cance of the NT thickness depends on the a priori risk for a chromo-strength of the association when such ﬁndings are noted on a screening somal abnormality. NT thickness increases with gestational age orexamination. crown-rump length. Figure 17-2 illustrates the NT between 11 and 14
228 CHAPTER 17 Prenatal Diagnosis of Congenital Disorders TABLE 17-8 ULTRASOUND FINDINGS weeks’ gestation. These observations suggested that NT could be used ASSOCIATED WITH TRISOMY 18 as a screening test for Down syndrome by converting the deviation from the expected mean to a likelihood ratio. Frequency NT combined with the maternal and gestational age to assess the Finding (%) risk for Down syndrome was studied in more than 100,000 pregnan- cies.91 NT was greater than the 95th percentile in more than 70% of Growth restriction 46 Hand or foot abnormalities* 39 fetuses with trisomy 21. The risk of Down syndrome was calculated Cardiac abnormality 31 by the maternal age and gestational age prevalence multiplied by the CNS abnormality 29 likelihood ratio. A cutoff of 1 : 300 was used. The studied sample Diaphragmatic hernia 13 included 326 fetuses with trisomy 21. Eighty-two percent of trisomy Ventral wall defect 10 21 fetuses were identiﬁed, with a false-positive rate of 8.3%.91 When a Facial abnormality 7 screen-positive rate of 5% was selected, the sensitivity was 77% (95% At least one abnormality 90 conﬁdence interval [CI], 72% to 82%). Subsequent studies have dem- onstrated similar Down syndrome detection rates, between 70% and *Including rocker bottom feet, overlapping ﬁngers. CNS, central nervous system. 75% (Table 17-9). From Gupta JK, Cave M, Lilford RJ, et al: Clinical signiﬁcance of fetal The screening paradigm using an ultrasound measurement to choroid plexus cysts. Lancet 346:724, 1995. determine a likelihood ratio is reliable only if NT is measured in a standard fashion. Standards for NT measurements include the following: 1. The minimal crown length should be 45 mm and the maximal, 3.5 84 mm. The success rate for accomplishing a measurement for these gestational ages is between 98% and 100%. The success rate falls to 3.0 90% at 14 weeks and onward.92 95th 2. Either transabdominal or transvaginal scanning can be used, with Nuchal translucency (mm) 2.5 about 95% of cases able to be imaged by the transabdominal 75th route.93 2.0 50th 3. A true sagittal section of the fetus as for measuring the fetal crown- rump length must be obtained. 25th 4. The magniﬁcation must be such that the fetus occupies at least three 1.5 fourths of the image. The magniﬁcation should be increased so that 5th each increment in the distance between calipers should only be 1.0 0.1 mm. Studies have demonstrated that ultrasound measurements 0.5 can be accurate to the nearest 0.1 to 0.2 mm.94 5. Care must be taken to clearly distinguish between the fetal skin and 0 the amnion. At this gestational age, both structures appear as thin 45 50 55 60 65 70 75 80 85 membranes. This can be accomplished by either waiting for spon- Crown-rump length (mm) taneous fetal movement away from the amniotic membrane or by bouncing the fetus off the amnion by asking the mother to coughFIGURE 17-2 Normative curves for nuchal translucency or tap on her abdomen (Fig. 17-3).measurement between 11 and 14 weeks’ gestation. (From 6. The maximal thickness of this subcutaneous translucency betweenNicolaides KH, Sebire NJ, Snijders RJM: The 11-14 Week Scan. New the skin and the soft tissue overlying the cervical spine should beYork, Parthenon, 1999.) measured by placing the calipers on the lines as illustrated in Figure 17-4. TABLE 17-9 STUDIES OF IMPLEMENTATION OF FETAL NUCHAL TRANSLUCENCY (NT) SCREENING Gestation Successful NT Cutoff False-Positive Rate Detection Rate Source (ref) (wk) N Measurement (mm) (%) of Trisomy 21 Pandya (89), 1995 10-14 1,763 100% >2.5 3.6 3 of 4 (75%) Szabo (90), 1995 9-12 3,380 100% >3.0 1.6 28 of 31 (90%) Bewley (97), 1995 8-13 1,704 66% >3.0 6.0 1 of 3 (33%) Bower et al (526), 1995 8-14 1,481 97% >3.0 6.3 4 of 8 (50%) Kornman et al (527), 1996 8-13 923 58% >3.0 6.3 2 of 4 (50%) Zimmerman et al (528), 1996 10-13 1,131 100% >3.0 1.9 2 of 3 (67%) Taipale et al (529), 1997 10-16 10,010 99% >3.0 0.8 7 of 13 (54%) Hafner (179), 1998 10-14 4,371 100% >2.5 1.7 4 of 7 (57%) Pajkrt (181), 1998 10-14 1,547 96% >3.0 2.2 6 of 9 (67%) Adapted from Nicolaides KH, Sebire NJ, Snijders RJM: The 11-14 Week Scan. New York, Parthenon, 1999.
CHAPTER 17 Prenatal Diagnosis of Congenital Disorders 229 + + + + + + + + + -5 + Correct FIGURE 17-4 Proper placement of the calipers for measuring the 0.09 cm nuchal translucency. (From Nicolaides KH, Sebire NJ, Snijders RJM: A The 11-14 Week Scan. New York, Parthenon, 1999.) NT measured but were not acted on, reported that, in the interventional groups, successful measurement was achieved in 100% of cases, whereas the noninterventional centers were successful in only 85%.97 In a recent prospective study,98 the NT was measured by two to four operators in 200 pregnant women, demonstrating that after an initial measurement, a second one made by the same operator or another operator varied from the initial measurement by less than 0.5 and 0.6 mm, respectively, in 95% of cases. It is suggested that a large part of the variation between operators can be accounted for by placement of the calipers rather than generation of the appropriate image. Subsequent studies99-101 have con- tinued to report small interoperator differences. -5 Because NT values are incorporated into a standardized algorithm along with biochemical analytes, it is critical that these ultrasound 0.09cm measurements be performed and monitored appropriately. To accom- B plish this, certiﬁcation and quality review programs have been devel- NT oped to ensure that accurate and precise NT measurements areFIGURE 17-3 First-trimester nuchal translucency (NT) obtained. The Fetal Medicine Foundation of London was the ﬁrst tomeasurement. Clear distinction of the amnion as opposed to the offer formalized NT training and quality review. In the United States,skin edge is made by waiting for fetal movement. Measurement the Nuchal Translucency Quality Review (NTQR) program was initi-before the fetus moves (A) is less accurate than after fetalmovement (B). ated in 2005. Both programs teach the mechanics of obtaining an NT measurement, have an image review process to ensure that the stan- dard technique is used correctly, and perform ongoing epidemiologic monitoring of sonographer and sonologist performance. Two studies7. During the scan, these measurements should be taken and the have evaluated the techniques used to ensure consistent NT results. maximum one recorded and used for Down syndrome risk Both conﬁrmed that ongoing expert review of images is an inefﬁcient calculation. and impractical approach. Epidemiologic monitoring in which indi-8. The NT should be measured with the fetal head in the neutral posi- vidual operator’s performance is compared with expected standards is tion. When the fetal neck is hyperextended, the measurement can preferable.101,102 be increased by 0.6 mm, and when the neck is ﬂexed, the measure- ment can be decreased by 0.4 mm.959. The umbilical cord may be found around the fetal neck in approxi- First-Trimester Biochemical Screening mately 5% to 10% of cases, which may produce a falsely increased Two serum analytes are useful for ﬁrst-trimester screening. Pregnancy- NT, adding about 0.8 mm to the measurement.96 In such cases, the associated plasma protein A has been demonstrated to have a mean measurements of NT above and below the cord differ, and the value of 0.4 MoM in trisomy 21 pregnancies. The free β subunit of smaller measurement is the most appropriate. hCG is elevated in Down syndrome pregnancies, with a mean value of 1.8 MoM. Screening using pregnancy-associated plasma protein A Even with these criteria, standardization of NT measurements (PAPP-A) alone identiﬁes about 40% to 45% of trisomy 21 pregnan-remains difﬁcult. Certiﬁcation courses are available with continuous cies, and free β-hCG identiﬁes about 23%, both with a false-positivequality assessment to maintain proper technique. The ability to achieve rate of 5%.103-105 Combining both free β-hCG and PAPP-A can identifya reliable measurement has been linked to the motivation of the sonog- 60% to 65% of trisomy 21 pregnancies, for a similar 5% false-positiverapher. A study comparing the results obtained from hospitals where rate.106 This is a serum analyte detection rate similar to that seen withNT was clinically used compared with those where they were merely triple screening in the second trimester.
230 CHAPTER 17 Prenatal Diagnosis of Congenital Disorders The total hCG molecule can also be used for ﬁrst-trimester screen-ing but has slightly less discrimination power than does the free β Additional First-Trimester Markers ofsubunit,107 especially at less than 11 weeks’ gestation. Free β-hCG Down Syndromebegins to increase in performance as a Down syndrome marker at asearly as 9 weeks’ gestation, reaching values almost twice those in unaf- Biochemical Markersfected pregnancies by 13 weeks. Levels of total hCG begin to increase ADAM 12 is the secreted form of a disintegrin and metalloprotease 12,above those in unaffected gestations at 11 weeks.108,109 The impact of a glycoprotein of the Meltrin family synthesized by the placenta andsubstituting total hCG for the free β subunit on overall Down syn- secreted throughout pregnancy. ADAM 12 has proteolytic functiondrome screening remains uncertain. A recent meta-analysis showed against insulin-like growth factor (IGF) binding proteins IGFBP-3 andthat in younger patients (<35 years), detection of Down syndrome IGFBP-5 and regulates the bioavailability and action of IGF-1 and -2.116increased by 4, 5, 6, and 7 percentage points at 9, 10, 11, and 12 weeks, Studies have shown that ﬁrst-trimester ADAM 12 levels are reduced inrespectively, when free β was added to pregnancy-associated plasma women carrying a Down syndrome pregnancy, and that the reductionprotein A and nuchal translucency compared with 0, 0, 2, and 4 per- is more pronounced in earlier gestation.117-119 Discrimination appearscentage points when intact human chorionic gonadotropin was best at around 8 to 10 weeks, with an overall median MoM of 0.79added.110 In patients with advanced maternal age (>35), inclusion of in Down syndrome pregnancies.119 Population modeling shows that afree β-hCG reduced the false-positive rate by 2.5, 3.1, 3.8, and 4.4 per- combination of ADAM 12 and PAPP-A measured at 8 to 9 weeks,centage points compared with 0.1, 0.3, 1.0, and 2.2 percentage points combined with NT and free β-hCG measured at 12 weeks, couldfor intact hCG at 9, 10, 11, and 12 weeks, respectively. Other authors achieve a detection rate of 97% with a 5% false-positive rate, or 89%have found less impact. Using samples from the FASTER study, Canick with a 1% false-positive rate.119and coworkers111 showed that at 12 weeks’ gestation, the addition offree β-hCG to NT and PAPP-A added only 0.9% (−3.3 to 6.3) detec- Ultrasound Markerstion. However, at earlier gestational ages the impact of free β-hCG Nasal Bone. Similar to ﬁndings in the second trimester, investiga-would be greater. tors have suggested that assessment of the fetal nasal bone (NB) can be used in the ﬁrst trimester to predict trisomy 21. This is based on the ﬂat nasal bridge area, which is a well-described component of theCombined First-Trimester Down syndrome phenotype, as well as on several histopathologic and radiographic studies demonstrating differences in the nasal bones ofNuchal Translucency and Down syndrome fetuses. Stempﬂe and colleagues120 found that NBBiochemistry Screening ossiﬁcation was absent in one quarter of Down syndrome fetusesCombining NT with serum analytes improves ﬁrst-trimester Down investigated between 15 and 40 weeks’ gestation, compared with nonesyndrome detection rates. Table 17-10 summarizes the large interna- of the controls. Similarly, Tuxen and colleagues121 evaluated Downtional experience with ﬁrst-trimester Down syndrome screening using syndrome fetuses between 14 and 25 weeks’ gestational age by radio-free β-hCG, PAPP-A, and NT measurements. Overall, for a 5% false- graph and pathologic study and found that the NB was absent in onepositive rate, combined ﬁrst-trimester risk assessment provides a third.Down syndrome detection rate of approximately 88% (95% CI, 84.0% Sonek and colleagues48 published the ﬁrst large prospective trial ofto 89.4%). In women older than 35, 90% to 92% of trisomy 21 preg- aneuploid risk evaluation using ﬁrst-trimester ultrasound assessmentnancies can be identiﬁed with a 16% to 22% false-positive rate.20,112 of the fetal nasal bone. They determined that the fetal nasal bone couldFirst-trimester screening can also identify trisomy 18 pregnancies. routinely be imaged and that its absence was associated with trisomyOver 90% of such pregnancies are screen positive when combined 21 (Fig. 17-5). The NB was absent in 73% of trisomy 21 fetuses com-biochemical and NT screening is used.112 pared with only 0.5% of euploid fetuses. They estimated that if NB When combining analytes, differences in gestational age–speciﬁc assessment were combined with maternal age and NT measurement,performance should be considered.113-115 At all gestational ages between 93% of Down syndrome cases would be detected at a false-positive rate9 and 12 weeks, NT and PAPP-A are the most efﬁcient markers. In of 5%, and 85% with a false-positive rate of 1%.combination, they are most efﬁcient at 11 weeks, when free and total A recent review of the literature by Rosen and D’Alton122 evaluatedhCG are least efﬁcient. In practice, screening is performed between 11 35,312 women having ﬁrst-trimester ultrasound assessment for NB. Inand 13 weeks of gestation. 33,314 cases (94.3%), the NB was successfully imaged. The sensitivity TABLE 17-10 STUDIES OF DOWN SYNDROME DETECTION RATES IN FIRST-TRIMESTER SCREENING Pregnancies Down Syndrome Cases Study (ref) Screened (Screen-Positive/Total) Detection Rate BUN (Wapner , 2003) 8,216 48/61 79% FASTER (Malone , 2005) 38,033 100/117 86% SURUSS (Wald , 2003) 47,053 84/101 83% Nicolaides (, 2005) 75,821 321/325 93% TOTAL 167,210 533/604 88.2% Screening tests were for free b-subunit of human chorionic gonadotropin, pregnancy-associated plasma protein A, and nuchal translucency (with a 5% false-positive rate).
CHAPTER 17 Prenatal Diagnosis of Congenital Disorders 231FIGURE 17-5 Ultrasound images of the fetal nasal bone (NB) in the ﬁrst trimester. First-trimester ultrasound images of euploid (left) andtrisomy 21 (right) fetuses demonstrate the presence of the nasal bone in the normal gestation and its absence in trisomy 21. Scanningtechniques are those suggested by the Fetal-Medicine Foundation for assessing NB and include the following: (1) The image is magniﬁed sothat each movement of the calipers causes a 0.1-mm incremental change. (2) A midsagittal view of the fetal proﬁle is obtained. (3) The anglebetween the ultrasound transducer and a line passing from the fetal forehead to the chin is 45 degrees. (4) When the NB is present, threeechogenic lines should be visible. The NB and overlying skin look like an equal sign. In the same view, the skin over the nasal tip should bevisible. If both the nasal tip and skin are present, and the NB echo cannot be visualized or is less echogenic than the skin, the NB is consideredabsent. (Fetal-Medicine Foundation, available at http://www.fetalmedicinefoundation.com/nasal.htm.)of NB alone for detecting trisomy 21 was 65% with a false-positive rate would be detected at a false-positive rate of 5%.127 For a false-positiveof 0.8%. The positive predictive value of the screen was 54%, meaning rate of 0.5%, the detection rate would be 90.5%. Although these datathat approximately 1 in 2 fetuses with an absent NB had trisomy 21. are promising, detection rates using this combined screen would beIf the NB was absent, the likelihood that a fetus had trisomy 21 was expected to be signiﬁcantly lower in an unselected population using aincreased 87-fold. The negative likelihood ratio with a normal NB was similar 5% false-positive rate. In addition, appropriate imaging of0.35 (95% CI, 0.32 to 0.39). the NB appears to be technologically more difﬁcult than measurement As experience with NB has increased, relationships between absent of the NT, making its use in a primary screening program lessNB, fetal crown-rump length (i.e., gestational age), NT, and ethnicity attractive.128have been established. The current data demonstrate that in euploid Tricuspid Regurgitation. Another potential ultrasound markerpregnancies, NB absence occurs more frequently with increasing NT. is tricuspid regurgitation determined by pulsed wave Doppler ultraso-In a series of 5851 high-risk patients containing 333 trisomy 21 fetuses, nography.129,130 This ﬁnding is present in around 8% of normal fetusesabsence of the NB had a likelihood ratio of 37.1 when the NT was less and 65% of those with trisomy 21. Combining tricuspid regurgitationthan the 95th percentile, and this was reduced to 13.4 when the NT with NT and PAPP-A would be expected to achieve a detection rate ofwas 4 or greater.123 The same study showed that the NB was more likely 95% with a 5% false-positive rate, or 90% with a 2% false-positiveto be absent at earlier gestational ages. For example, in euploid fetuses rate.131with a crown-rump length between 45 and 54 mm, the NB was absent Ductus Venosus Wave Form. A third potential marker is abnor-in 4.7% of cases. At a crown-rump length between 75 and 84 mm, the mal blood ﬂow through the ductus venosus. Studies have shown thatNB was absent in only 1.0% of cases. Prefumo and colleagues124 found pulsation of the ductus venosus gives detection rates of 65% to 75%that NB hypoplasia was more common in the euploid fetuses of women with a 4% to 5% false-positive rate,132 and the rate increased to 75%of African descent when compared with either Asian or white popula- to 80% when NT was added. When serum biochemical markers mea-tions (odds ratio, 2.3). Cicero and colleagues125 also found an increased sured at 10 weeks were also added, the modeled detection rate increasedincidence of absent fetal NB in the ﬁrst trimester in women of Afro- to 92% at a 5% false-positive rate, or 84% at a 1% false-positiveCaribbean and southern Asian descent. The NB was absent in 2.5%, rate.1339.0%, and 5.0% of white, Afro-Caribbean, and southern Asian popula-tions, respectively. Likelihood ratios for trisomy 21 with absent NBwere 31.3, 8.8, and 14.2, respectively, in these three populations. Impact of Spontaneous Miscarriages NB status is independent of serum biochemistry, allowing NB on First-Trimester Screeningassessment to be combined with measurements of NT and maternal A potential disadvantage of earlier screening is that chromosomallyserum markers to increase ﬁrst-trimester screening performance.126 In abnormal pregnancies that are destined to miscarry will be identiﬁed.a retrospective case-control study of a high-risk population with a The impact of this can be evaluated because 69% of trisomy 21 fetusesmedian maternal age of more than 38 years assessed by NT, NB, and living in the ﬁrst trimester and 76% of those alive in the second tri-biochemistry, it was estimated that 97% of Down syndrome cases mester will be born alive.4 Using this information, Dunstan and Nix134
232 CHAPTER 17 Prenatal Diagnosis of Congenital Disorderscalculated that a detection rate of 80% in the ﬁrst trimester is approxi- a 1% false-positive rate. If all screen-negative patients proceed tomately equivalent to a second-trimester sensitivity of 75%, suggesting second-trimester screening, an overall detection rate of 95% can bethat when early spontaneous losses of trisomy 21 pregnancies are obtained with a 5% false-positive rate. Although this approach hasconsidered, ﬁrst-trimester screening is superior to that presently avail- excellent performance, with a high proportion of affected pregnanciesable in the second trimester. identiﬁed in the ﬁrst trimester, it is logistically demanding. First-trimester screening would be less desirable if screen-positive Contingent sequential screening is similar to stepwise sequentialpregnancies or those with enlarged NTs were preferentially lost. In a screening, but patients with a very low ﬁrst-trimester combined riskstudy of 108 fetuses with trisomy 21 diagnosed in the ﬁrst trimester do not have second-trimester analysis performed. Using an approachbecause of increased NT, Hyett and colleagues found that six patients in which patients with a ﬁrst-trimester risk of 1 : 1300 or less completeelected to continue the pregnancy.135 In ﬁve of the six fetuses the trans- screening in the ﬁrst trimester, only 15% to 20% of patients have tolucency resolved, and at the second-trimester scan the nuchal fold return for second-trimester analysis.141,142 Contingent sequentialthickness was normal. All six of these trisomy 21 fetuses were born screening has a detection rate of 92% to 94% for a 5% screen positivealive. Wapner and colleagues112 calculated that greater than 80% of rate.140screen-positive trisomy 21 pregnancies would be born alive. Nasal Bone Contingency Screening. Nasal bone assessment is technically more difﬁcult to perform than NT, which may limit avail-Other Approaches to Down Syndrome ability. To address this Nicolaides and colleagues143 proposed a two- stage screen, reserving NB assessment for patients at intermediate riskScreening: Combining First- and after the combined ﬁrst-trimester screen is complete. In this model,Second-Trimester Screening Tests patients evaluated by NT and serum markers with a risk of 1 in 100 orScreening performance may be improved by combining analytes per- greater would be offered CVS, and those with a risk of less than 1 : 1000formed at different gestational ages.136,137 These approaches include the would be deemed to have such a low risk that no further testing isfollowing. offered. Those with a risk between 1 : 101 and 1 : 1000 would have NB Integrated Aneuploidy Screening (Noninformative Sequen- evaluation. In initial studies, performance of this two-stage approachtial). Wald and colleagues137 described a protocol for screening based was similar to using NB assessment as part of the initial screen. Theon tests performed during both the ﬁrst (NT and PAPP-A) and second two-stage approach would have a signiﬁcant advantage because onlytrimesters (quad screen). A single risk estimate is calculated in the about 15% of pregnancies would require NB evaluation, which couldsecond trimester using all six of the measured analytes. Integrated be performed in centers that have developed special expertise in thisscreening has a detection rate of approximately 95% with a 5% false- technique.positive rate.20,137 Approximately 85% of affected pregnancies would bedetected with a false-positive rate of only 0.9%.137,138 Although this Can Maternal Age Be Eliminatedscreening approach is quite sensitive and speciﬁc, withholding the riskestimate until the second trimester precludes earlier prenatal diagnosis as an Indication for Invasiveby chorionic villus sampling (CVS) and is not an acceptable approach Prenatal Diagnosis?for many women.139 Maternal age of 35 or older has been a standard indication for invasive If NT scanning is not available, an integrated serum screen may be testing for more than 35 years. When it was initially suggested, approxi-performed (PAPP-A in the ﬁrst trimester and a quad screen in the mately 5% of births were to women older than 35 years, as were 30%second trimester). This approach has a detection rate of 86% to 90% of trisomy 21 gestations. Presently, almost three times as many womenat a 5% false-positive rate.20,137 giving birth are older than 35, and this group contains about 50% of Sequential Testing. In an attempt to maximize screening perfor- trisomy 21 conceptions. For every invasive procedure done with mater-mance by combining ﬁrst- and second-trimester analytes yet retain the nal age as the only indication, the odds of being affected are approxi-beneﬁt of ﬁrst-trimester diagnosis, various methods of sequential mately 1 : 100.20 As screening has improved, the importance of maternalscreening have been proposed. In these approaches, ﬁrst-trimester risk age as a single indication for testing has been reevaluated.results are calculated and used for clinical management, with second- In women aged 35 years and older, 87% of Down syndrometrimester testing performed in selected cases. pregnancies and 25% of unaffected pregnancies will be triple-screen Three approaches to sequential risk assessment are presently avail- positive at a cutoff of 1 : 250.18 The incidence of Down syndrome inable. In independent sequential testing, a ﬁrst-trimester combined risk this age group is approximately 1 : 100. Table 17-11 demonstratesis calculated with a 1 : 270 screen-positive cutoff. Decisions on invasive that performing an amniocentesis on screen-negative women (risktesting are made on the basis of these results. In the second trimester, <1 : 270) aged 35 years or older would lead to the loss of three normala quad screen is performed and calculated independent of the ﬁrst- pregnancies from procedure-induced complications for every Downtrimester results. This approach provides detection rates greater than syndrome pregnancy identiﬁed. First-trimester screening has greater95%,20,136 but it has an unacceptably high false-positive rate of greater than a 90% sensitivity with a 15% false-positive rate in women agedthan 10% because independent calculation of the quad screen risk 35 years or older.112 Using the approach illustrated in Table 17-1, it candoes not take into consideration the reduced second-trimester preva- be calculated that almost four normal pregnancies will be lost for eachlence of Down syndrome pregnancies after ﬁrst-trimester prenatal Down syndrome pregnancy identiﬁed.diagnosis. Screening the entire population of pregnant women regardless of Stepwise sequential testing reduces the high false-positive rate of age provides the most effective use of resources. Presently, 14.2% ofindependent sequential testing and offers the highest risk patients the women older than 35 are offered invasive testing, as are about 5%option of ﬁrst trimester invasive testing by using a high ﬁrst-trimester of women under age 35 who are screen positive, making greater thanrisk cutoff and calculating the second-trimester risk by integrating 18% of pregnant women eligible for testing. If second-trimester screen-information from both trimesters.140 For example, using a 1 : 65 cutoff ing were used for all patients regardless of age and only screen-positivein the ﬁrst trimester identiﬁes 70% of affected pregnancies with only patients were offered invasive testing, the number of procedures would