Serum Screening in Pregnancy Case File

Serum Screening in Pregnancy Case File

Eugene C. Toy, MD, Patti Jayne Ross, MD, Benton Baker III, MD, John C. Jennings, MD

CASE 7

A 20-year-old G1P0 woman at 16 weeks’ gestation by last menstrual period (LMP) has received a serum maternal α-fetoprotein (AFP) test that returned as 2.8 multiples of the median (MOM). She is fairly sure of her last menstrual period and has regular menses. She denies a family history of congenital anomalies or chromosomal abnormalities. On examination, she is afebrile, her blood pressure (BP) is 100/70 mm Hg, and her heart rate (HR) is 70 beats per minute (bpm). The heart and lung examinations are normal. The fundal height is midway between the symphysis pubis and the umbilicus. Fetal heart tones are in the range of 140 bpm.

» What is your next diagnostic step?

» What is the purpose of the maternal serum α-fetoprotein (msAFP) test?

ANSWER TO CASE 7:

Serum Screening in Pregnancy                                                  

Summary: A 20-year-old G1P0 woman at 16 weeks’ gestation by a fairly certain last menstrual period has received a serum maternal α-fetoprotein test that returned as 2.8 MOM.

• Next diagnostic step: Basic obstetric ultrasound examination to assess for dates and multiple gestations.
• Purpose of msAFP: The purpose is to assess the risk for a fetal open neural tube defect, and can also be used to assess for the risk of aneuploidy, such as fetal Down syndrome or trisomy 18.

ANALYSIS

Objectives

1. Understand that the most common causes of abnormal serum screening are wrong dates and multiple gestations.
2. Know that an elevated maternal serum α-fetoprotein level may be associated with an open neural tube defect.
3. Know that a low msAFP level may be associated with fetal Down syndrome.
4. Be aware of the large number of noninvasive and invasive tests for fetal anomalies and aneuploidy.
5. Be aware of some of the teratogens and their fetal effects.

Considerations

This patient is at 16 weeks’ gestation by a fairly certain last menstrual period, which is consistent with the clinical examination. The gestational age window of 16 to 20 weeks is the appropriate time to screen with serum testing. The msAFP returned as 2.8 MOM, which exceeds the usual cutoff of 2.0 or 2.5 MOM. The interpretation of the msAFP depends on gestational age and number of fetuses. The components of a certain last menstrual period are (1) patient sure of date of last menstrual period, (2) regular menses, (3) LMP was normal, and (4) patient has had no spotting or bleeding after LMP. The uterine size correlates with the dates. At 16 weeks’ gestation, the fundus is usually midway between the symphysis pubis and the umbilicus. At 20 weeks’ gestation, the fundal height is generally at the level of the umbilicus. Although this patient has a sure LMP and size and date consistency, there is still a significant risk of a dating abnormality or a multifetal gestation. Hence, the next appropriate step is the basic ultrasound examination. If there is a dating error, the msAFP result would be recalculated based on the corrected gestational age. If the msAFP is still abnormally elevated, then at an early gestational age such as 16 weeks, repeating the serum test is an option. For women with abnormally elevated msAFP at a later gestational age, such as 20 weeks, genetic counseling and referral for amniocentesis may be considered.

APPROACH TO:

Abnormal Serum Screening in Pregnancy                                                     

DEFINITIONS

ALPHA-FETOPROTEIN: A glycoprotein made by the fetal liver, analogous to the adult albumin.

FIRST-TRIMESTER SCREENING: Use of biochemical markers (PAPP-A and β-hCG) and/ or transvaginalsonography measuring the aspect in the posterior neck region called the “nuchal translucency” giving a risk of Down syndrome and trisomy 18. This is performed between 10 and 13 weeks’ gestation.

NEURAL TUBE DEFECT: Failure of closure of the embryonic neural folds leading to an absent cranium and cerebral hemispheres (anencephaly) or nonclosure of the vertebral arches (spina bifida).

OPEN NEURAL TUBE DEFECT: A neural tube defect that is not covered by skin.

MATERNAL SERUM α-FETOPROTEIN: α-Fetoprotein level drawn from maternal blood; this may be elevated due to increased amniotic fluid α-fetoprotein.

TRISOMY SCREEN: Three or four serum markers that may indicate an increased risk of chromosomal abnormalities, drawn in the second trimester (typically between 15 and 21 weeks’ gestation). A common combination includes maternal serum α-fetoprotein, human chorionic gonadotropin (hCG), inhibin-A, and unconjugated estriol.

TERATOGEN: An agent or factor that causes a malformation in the embryo.

CLINICAL APPROACH

The triple (or trisomy) screen is used in pregnant women between 15 and 21 weeks’ gestation to identify those pregnancies that may be complicated by neural tube defects, Down syndrome, or trisomy 18. It is a multiple marker test, and the term “triple” is often used to denote that it analyzes three chemicals in the maternal serum to determine the risk for neural tube defects or fetal aneuploidy: α-fetoprotein, human chorionic gonadotropinhCG, and unconjugated estriol. Although the triple screen may be offered to women over the age of 35 years, or advanced maternal age, genetic amniocentesis provides more diagnostic information.

α-Fetoprotein is a glycoprotein synthesized initially by the fetal yolk sac and then later by the fetal gastrointestinal tract and liver. It passes into the maternal circulation by diffusion through the chorioamniotic membranes. When there is an opening in the fetus not covered by skin, levels of AFP increase in the amniotic fluid and maternal serum. Maternal serum AFP is measured in MOM. Different laboratories have different cutoff levels for abnormal AFP; in general, levels >2.0 to 2.5 MOM are suspicious for neural tube defects and warrant further evaluation. However, an abnormally elevated serum AFP level does not necessarily coincide with fetal neural tube defects. Other causes of elevated and decreased maternal serum AFP are listed in Table 7– 1.

Table 7–1 • CAUSES OF ELEVATED AND LOW msAFP

Elevated msAFP

Underestimation of gestational age Multiple gestations Neural tube defects Abdominal wall defects Cystic hygroma Fetal skin defects Sacrococcygealteratoma Decreased maternal weight Oligohydramnios

Decreased msAFP

Overestimation of gestational age Chromosomal trisomies Molar pregnancy Fetal death Increased maternal weight

In contrast to neural tube defects, which have an abnormally elevated maternal serum AFP, those pregnancies complicated by Down syndrome have a low maternal serum AFP.

Unconjugated estriol is also decreased in fetuses with Down syndrome. Human chorionic gonadotropin, however, is elevated in these fetuses. By combining these serum chemicals into a multiple marker screening test, approximately 60% of all Down syndrome pregnancies can be identified. With trisomy 18, all of the serum markers are abnormally low. Different variations of the multiple marker test exist, such as one that adds inhibin A as a fourth analyte to further improve detection rates. (Table 7– 2 lists the noninvasive and invasive tests).

First-trimester Down syndrome screening has also become available to women. This allows prediction of abnormal pregnancies at an earlier gestational age. Firsttrimester screening combines two serum analytes: pregnancy-associated plasma protein (PAPP-A) and free β-hCG with sonographic measurement of nuchal

Table 7–2 • OVERVIEW OF PRENATAL GENETIC TESTS

Noninvasive Techniques

For fetal structural anomalies (ultrasound, magnetic resonance imaging) For neural tube defects (msAFP) For fetal Down syndrome (msAFP, maternal uE3, hCG, inhibin A) For various aneuploidy (cell-free fetal DNA)

Invasive Techniques

Fetal visualization (fetoscopy) Fetal tissue sampling (amniocentesis, chorionic villus sampling, percutaneous umbilical blood sampling) Preimplantation blastocyst biopsy (with IVF)

Table 7–3 • SERUM ANALYTES ASSOCIATED WITH FETAL ANOMALIES
Genetic Disorder	AFP	uE3	hCG	Inhibin A	PAPP-A	Beta hCG
Trisomy 21	↓	↓	↑	↑	↓	↑
Trisomy 18	↓	↓	↓	N/A	↓	↓
Trisomy 13	N/A	N/A	N/A	N/A	↓	↓

The first step in the management of an abnormal triple screen is a basic ultrasound to determine the correct gestational age, to identify the possibility of multiple gestation, and to exclude fetal demise. The most common cause of abnormal serum screening is wrong dating. If the risk of trisomy or neural tube defects is still increased after a basic sonogram, amniocentesis or targeted ultrasound is offered. A targeted examination can correctly identify fetuses with neural tube defects by direct visualization of the fetal head and spine. Furthermore, ultrasound may also detect those fetuses suspicious for having Down syndrome by identification of a thickened nuchal fold, shortened femur length, or echogenic bowel. Other conditions associated with an abnormally high or low maternal serum AFP, such as abdominal wall defects, oligohydramnios, and fetal skin defects, can be identified with ultrasound.

Because high-resolution sonography can detect up to 95% of neural tube defects, some practitioners will not proceed with invasive testing for an elevated msAFP. However, when amniocentesis is chosen for an elevated msAFP, the amniotic fluid is tested for AFP levels. Fetal karyotype is also obtained through amniocentesis, which will identify fetal aneuploidy, such as the trisomies. Fetal loss rate from an amniocentesis is about 0.5%. Other complications include rupture of membranes and chorioamnionitis.

The identification of a fetus affected by a neural tube defect or a chromosomal abnormality can be an ethical and moral dilemma for the parents, whose previous hopes and dreams for having a “normal” child are now extinguished. The parents should not be forced into any decision, but should be provided information in an unbiased fashion.

Emerging Concepts

At the writing of this chapter, the use of noninvasive blood test on maternal blood assays fetal DNA for trisomy 21, 18, and 13, and several other aneuplopiodes, and is for use between 10 and 22 weeks’ gestation. Approximately 13% of cell-free fetal DNA (cfDNA) in the maternal serum is fetal in origin. The technology known as massively parallel shotgun sequencing analyzes and amplifies cfDNA to detect excessive fetal chromosomes. Initial studies indicate a 97% sensitivity and near 99% specificity rate for Down syndrome, but the study populations were small. Other algorithms are being developed to likewise incorporate age-related risk together with laboratory cfDNA results. This technology has not been well studied in large populations yet; and its role at the time of printing was delegated to those women at high risk for Down syndrome. Both the American College of Obstetricians and Gynecologists and the Society for Maternal Fetal Medicine cautioned against the use of cfDNA as a definitive diagnostic test (acting on results), but still as a screening test, since there were false positive results noted.

Table 7–4 • SELECT LISTING OF TERATOGENS
Agent	Effect on Embryo
Androgens	Masculinization of female fetus, labial fusion
Alcohol	Fetal alcohol syndrome, IUGR, microcephaly
Phenytoin (Dilantin)	Fetal hydantoin syndrome, IUGR, microcephaly, facial defects
Lithium carbonate	Heart and great vessel defects (Epstein anomaly)
Methotrexate	Skeletal defects, limb defects
Retinoic acid (vitamin A)	Facial defects, neural tube defects
ACE inhibitors	Skull anomalies, limb defects, miscarriage; renal tubule dysgenesis, renal failure in neonate, oligohydramnios
Warfarin	CNS and skeletal defects
Valproic acid, carbamazepine	Neural tube defects

Teratogens

Every baby has a 3% to 5% baseline risk of a birth defect. Specific exposures toagents during organogenesis—days 15 to 60—may lead to malformations. During the first 2 weeks of gestation, a teratogen usually has an “all or nothing” effect. In other words, the embryo is killed or recovers from the exposure. For some prescribed medications, the benefits may outweigh the risks. There is a consensus, however, that some agents should never be used during pregnancy (retinoic acid derivatives). See Table 7– 4 for a partial listing of common teratogens.

COMPREHENSION QUESTIONS

7.1 A 23-year-old G1P0 woman at 20 weeks’ gestation undergoes an ultrasound examination for size greater than her dates. The ultrasound reveals hydramnios with an amniotic fluid index of 30 cm. The fetal abdomen reveals a cystic mass in the right abdominal region and a cystic mass in the left abdominal area. Which of the following is the most likely associated condition?

A. Gestational diabetes

B. Congenital ovarian tumors

C. Down syndrome

D. Rh isoimmunization

7.2 A 28-year-old woman delivers a baby with a cleft palate and cleft lip. The baby is otherwise healthy. The patient asks about whether there is a genetic reason for this anomaly. Which of the following is the best explanation of the genetics of this condition?

A. Autosomal dominant

B. Autosomal recessive

C. X-linked dominant

D. X-linked recessive

E. Multifactorial

7.3 A 22-year-old G2P1 woman at 25 weeks’ gestation with a sure last menstrual period asks for serum screening. The patient’s sister has one child with Down syndrome and, otherwise, there is no family history of anomalies or genetic disorders. Which of the following is the most appropriate response?

A. Amniocentesis is the appropriate test.

B. Serum screening should be performed.

C. Explain to the patient that it is too late for serum screening, but that her risk for Down syndrome is not much higher than her age-related risk.

D. The patient being only 22 years of age does not need serum screening.

E. The patient has a 25% chance of her baby having Down syndrome.

7.4 A 28-year-old G1P0 woman at 16 weeks’ gestation is noted to have an elevated msAFP at 2.9 MOM. She underwent a targeted ultrasound examination which did not reveal a neural tube defect. Her physician also undertakes a diligent search for an etiology for the elevated msAFP without identifying an etiology. Which of the following conditions is this patient at increased risk?

A. Increased incidence of stillbirth

B. Gestational diabetes

C. Placenta previa

D. Molar pregnancy

E. Down syndrome

7.5 A 22-year-old woman is seen for her first prenatal visit at 16 weeks’ gestation with a family history of congenital deafness and neonatal renal disease. The patient’s hearing is normal. Which of the following is the best next step?

A. Amniocentesis for karyotype

B. Amniocentesis for rubella PCR

C. Genetic counseling

D. Glucose challenge testing

7.6 A 34-year-old woman is noted to be at 34 weeks’ gestation. The size is less than her dates. An ultrasound is performed which reveals oligohydramnios with an AFI of 2.1 cm. Which of the following agents may be responsible for this condition?

A. Lithium exposure

B. Angiotensin converting enzyme (ACE) inhibitor

C. Oral hypoglycemic agent

D. Phenytoin (Dilantin)

ANSWERS

7.1 C. This baby has the “double bubble” of duodenalatresia. The hydramnios results from the inability of the baby to swallow. Duodenal atresia is strongly associated with fetal Down syndrome. Gestational diabetes is associated with hydramnios occasionally; however, duodenal atresia is not related. Rh isoimmunization can also lead to hydramnios and hydrops but not duodenal atresia.

7.2 E. The genetics for cleft palate and cleft lip in the absence of other anomalies is multifactorial, and not a clear genetic transmission. The risk of recurrence is generally about 5%. The risk is higher if one of the parents also has a cleft lip/ palate. Other disorders that are multifactorial include cardiac malformations and neural tube defects.

7.3 C. The window for serum screening is usually between 15 and 21 weeks, so that her gestational age of 25 weeks is too late. The history of her sister having a baby with Down syndrome confers a very small, if any, increased risk for her own pregnancy. If the patient herself had a prior baby with Down syndrome, the risk would be substantially increased, and genetic counseling with possible amniocentesis for karyotype would be appropriate.

7.4 A. Pregnancies with elevated msAFP, which after evaluation are unexplained, are at increased risk for stillbirth, growth restriction, preeclampsia, and placental abruption. Thus, many practitioners will perform serial ultrasound examinations, monitor for these complications, and perform fetal antenatal testing such as biophysical profile testing.

7.5 C. Genetic counseling is appropriate with a family history of possible heritable syndromes. A glucose challenge test would not be helpful in evaluating heritable syndromes because it is used as a screen for gestational diabetes. Genetic counseling is recommended before a risky procedure, such as an amniocentesis, is performed because based on the family history, it may not be indicated in this situation.

7.6 B. Oligohydramnios is found with fetal exposure to ACE inhibitors. Neonatal renal failure may also be noted. Lithium is associated with Epstein anomaly (a fetal heart malformation); Dilantin is associated with a fetal hydantoin syndrome of intrauterine growth retardation, microcephaly, and facial defects.

    CLINICAL PEARLS    

» The most common cause of abnormal triple screening is wrong dates. » The next step in the evaluation of abnormal triple screening is the basic ultrasound. » Up to 95% of neural tube defects are detectable by targeted sonography. » About 60% of Down syndrome cases are detected with the triple screen with an elevated human chorionic gonadotropin level, low msAFP, and low unconjugated estriol. » An elevated msAFP suggests a neural tube defect, but there are many other etiologies. » Cell-free DNA testing may be very useful especially in high-risk individuals, but should not be used in general in low-risk individuals. » Cell-free DNA testing should not be viewed as a diagnostic test and does have false positives and negatives, especially in low-risk patients. » Teratogenic exposure prior to 2 weeks’ gestation leads to an “all or nothing effect.” Organogenesis occurs between days 15 and 60 of embryonic life.

REFERENCES

American College of Obstetricians and Gynecologists. Screening for fetal chromosomal abnormalities. ACOG Practice Bulletin 77. Washington, DC; 2007. (Reaffirmed 2011.) 

American College of Obstetricians and Gynecologists. Cell-free DNA screening for fetal aneuploidy. ACOG Committee Opinion 640. Washington, DC; 2015. 

Cunningham FG, Leveno KJ, Spong CY. Prenatal diagnosis and fetal therapy. In: Williams Obstetrics. 24th ed. New York, NY: McGraw-Hill; 2014:313-339. 

Lu MC, Williams J III, Hobel CJ. Antepartum care: preconception and prenatal care, genetic evaluation and teratology, and antenatal fetal assessment. In: H acker NF, Gambone JC, Hobel CJ, eds. Essentials of Obstetrics and Gynecology. 5th ed. Philadelphia, PA: Saunders; 2009:71-90. 

Sifakis S, Papantoniou N, Kappou D, Antsaklis A. Noninvasive prenatal diagnosis of Down syndrome: current knowledge and novel insights. J Perinat Med. 2012;40:319-327.

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