Protocols for High-Risk Pregnancies. Группа авторов
such as DiGeorge syndrome, which results from a deletion on chromosome 22 (22q11.2), and Williams syndrome, which results from a deletion on chromosome 7 (7q11.23). Chromosomal microarray analysis can identify submicroscopic abnormalities that cannot be seen with conventional karyotyping, and these copy number variants can be associated with significant genetic diseases.
Pathophysiology
The phenotype of trisomy 21 occurs when there is a triplication of dosage‐sensitive genes on chromosome 21; these genes comprise the Down syndrome critical region. Nondisjunction of the chromosome 21 pair during meiosis of the germ cells accounts for 95% of cases of trisomy 21. In the vast majority of cases, the extra chromosome is maternal in origin, and there is a strong correlation between maternal age and the chances of fetal trisomy 21. In less than 5% of cases, the additional chromosome 21 material is a result of an unbalanced translocation, usually affecting chromosomes 14 and 21, but occasionally also involving chromosomes 15 or 22. About 50% of such cases occur as de novo translocations and 50% are inherited from a parent who carries a balanced translocation. Rarer cases of trisomy 21 are mosaic, in which some cell lines carry three copies of chromosome 21, while others are diploid and normal. Trisomies 13 and 18 also occur due to meiotic nondisjunction in approximately 85% of cases, while 10% of cases are mosaic and 5% are due to a translocation.
Copy number variants (CNV) occur when the number of copies of a particular gene or genomic region varies from one individual to the next; these variants can be duplications or deletions. Large CNVs may be detectable by karyotype, but most require chromosomal microarray to be diagnosed. Although small when compared to trisomy of an entire chromosome, CNVs can be associated with significant medical and intellectual disabilities. Unlike the common trisomies, the rate of significant CNVs does not increase with maternal age and is estimated at about 0.5–1% of pregnancies in the mid‐trimester. Therefore, these are more common than Down syndrome and the common aneuploidies in women under age 35.
Diagnosis and screening protocols
Prenatal screening and diagnostic testing for detection of chromosomal abnormalities should be offered to all pregnant women, regardless of maternal age. Prenatal diagnostic testing involves direct analysis of fetal tissue, with collection through chorionic villus sampling (CVS) and amniocentesis being the most commonly performed prenatal procedures for diagnostic genetic testing. In contrast, prenatal screening provides a risk of chromosomal abnormality, with the most common current approaches being combinations of first‐ and second‐trimester serum and sonographic screening, and cell‐free DNA (cfDNA) screening, also referred to as noninvasive prenatal testing (NIPT) or noninvasive prenatal screening (NIPS).
Prenatal diagnostic testing
Prenatal diagnostic testing can be performed on fetal tissue collected by first‐trimester CVS or second‐trimester amniocentesis. CVS is typically performed between 10 and 14 weeks of gestation, although a later placental biopsy is also possible and may be required under some clinical circumstances. Two approaches are commonly used to access the placenta under sonographic guidance; with the transabdominal approach, a 20 gauge spinal needle traverses the maternal abdominal and uterine walls, while with the transcervical approach, a plastic cannula or biopsy forceps traverses the vagina and cervix. Both transabdominal and transcervical CVS are associated with an overall pregnancy loss rate of approximately 1 in 455 or 0.22%; this is not statistically different from the risk associated with amniocentesis. CVS performed prior to 10 weeks of gestation has been associated with a risk of fetal limb reduction defects and is not recommended; this risk is not increased with later procedures (10 weeks and later).
Genetic amniocentesis is most commonly performed between 15 and 20 weeks of gestation, although can also be performed later. Sonographically directed placement of a 22 gauge spinal needle into the amniotic cavity is a very safe procedure, with a reported loss rate of 1 in 900 pregnancies, or 0.11%. Recent data indicate that when compared to patients with the same risk profile, the loss rate of CVS and amniocentesis is negligible.
Fetal tissue obtained with CVS or amniocentesis can be cultured for karyotype analysis, or DNA can be extracted from chorionic villi, amniotic fluid, or cultured fetal cells for chromosomal microarray analysis (CMA) or other specialized genetic testing. When indicated, fluorescence in situ hybridization can be done on interphase cells for rapid aneuploidy testing or on metaphase cells for identification of microdeletions or duplications.
Cell‐free DNA screening
In 2011, cell‐free DNA screening (also known as noninvasive prenatal testing or noninvasive prenatal screening) became clinically available as a screening test for aneuploidy. This screening test relies on the analysis of cell‐free DNA (cfDNA) fragments in the maternal circulation. After 10 weeks of gestation, approximately 10–15% of the cfDNA in the maternal serum is of placental origin and therefore reflects the fetal DNA. Clinical testing measures the chromosomal contribution of the cfDNA in the maternal circulation to determine whether there is over‐ or underrepresentation of targeted chromosomes. Different laboratories use different approaches, including massively parallel shotgun sequencing (MPSS), a targeted microarray approach, or targeted sequencing using single nucleotide polymorphisms (SNPs); performance for aneuploidy screening is generally comparable between platforms. Standard cfDNA screening tests for trisomies 13, 18, and 21, and can also assess the sex chromosomes to determine fetal sex and, in some cases, screen for sex chromosomal aneuploidy.
The accurate performance of cfDNA screening depends on the presence of adequate fetal (placental) cfDNA, referred to as the “fetal fraction.” In some laboratories, a result is not provided when the fetal fraction falls below a prespecified level; this cut‐off is typically about 4%. Early gestational age, increasing maternal body mass index, and fetal aneuploidy are associated with a lower fetal fraction and increase the chances of a failed test.
Studies of test performance for cfDNA screening report a >99% detection rate for fetal trisomy 21 and 98% detection for trisomy 18 with a combined false‐positive rate (FPR) of 0.13–0.25%. Because trisomy 13 is a rare disorder, data are far more limited but reported detection rates vary from 40% to 100% in individual studies. The detection rate of sex chromosome aneuploidy is also difficult to determine due to limited data.
Importantly, these data were calculated for patients with a reported result, and as many as 3–4% of samples result in test failure. Test failure, particularly in the setting of low fetal fraction, is associated with an increased risk of aneuploidy and patients should be counseled accordingly and offered follow‐up testing.
While cfDNA screening has excellent performance in detection of trisomy 21, both false‐positive and false‐negative results can occur, particularly with low fetal fraction. The presence of mosaicism or a vanishing twin may result in false‐positive cfDNA results. Standard cfDNA screening tests do not provide risk assessment for other chromosomal, genetic, or structural disorders. Some laboratories offer expanded cfDNA panels to test for chromosomal microdeletions, rare autosomal trisomies, or genome‐wide copy number variants. Such tests have not been clinically validated, performance characteristics are unknown, and these are generally not recommended at the present time.
First‐trimester combined screening
The ability to provide an accurate, patient‐specific, risk assessment for fetal trisomy 21 during the first trimester is an established part of routine clinical practice. This allows patients the option of CVS to confirm or exclude fetal aneuploidy, and the possibility of pregnancy termination earlier in gestation. Such patient‐specific risk estimation is currently most commonly performed using a combination of maternal age, sonographic measurement of nuchal translucency (NT), and assay of two maternal serum markers – pregnancy‐associated plasma protein A (PAPP‐A) and either the free beta‐subunit (fβ) or the intact molecule of human chorionic gonadotrophin (hCG).
Nuchal translucency sonography
Nuchal translucency