Prenatalsafe® screening test limits
A non-invasive prenatal test that analyses circulating cell-free fetal DNA isolated from a maternal blood sample is a screening test rather a diagnostic test. Although the Prenatal SAFE® screening test is very accurate, the test carries a residual risk of false positive and false negative results (0.1%) and is neither intended nor validated for diagnosis.
The results should be evaluated in the context of the pregnant patient's clinical and family history. Furthermore, the test is not a substitute for invasive prenatal diagnosis (CVS or Amniocentesis). Conclusions about the foetal condition and pregnancy management decisions should not be based on the results of the NIPT screening alone.
This test is not validated for pregnancies with more than two fetuses and is not designed to detect chromosomal mosaicisms or polyploidy, such as triploidy. The test was validated on single or twin, monozygotic or dizygotic pregnancies, with at least 10 weeks of gestation. The test can not rule out the presence of all fetal chromosomal abnormalities.
The result of this test does not exclude the possibility that the chromosomes harbour abnormalities other than those included in the test, and it does not detect abnormalities of untested chromosomes, genetic disorders, birth defects or congenital complications of other origin. The test result may not reflect the real state of the fetus, as it can detect chromosomal abnormalities arising from confined placental mosaicisms (i.e. the presence of two cell lines with different chromosome arrangement), balanced chromosomal rearrangements, vanishing twin or maternal condition
The Prenatalsafe® screening test is not able to detect point mutations, methylation defects, polyploidies. The partial alterations of the analysed chromosomes and the structural chromosomal alterations can be highlighted only with the Prenatalsafe® Karyo screening tests. The estimated resolution limit of the test is superimposable to that of the cytogenetic (traditional) cytotoxicity with 400 bands (about 7-10 Mb). The Prenatalsafe® Karyo Plus highlights structural chromosomal alterations at a resolution of about 3 Mb, at the level of the chromosomal regions associated with the microdeletion syndromes investigated.
Prenatalsafe® 3 evaluates only the aneuploidy of chromosomes 13, 18, and 21. Prenatalsafe® 5 also evaluates the aneuploidy of the sex chromosomes (X and Y)
The results of the test can be confounded by certain maternal and fetal factors including but not limited to: recent maternal blood transfusion; maternal organ transplant; maternal surgical procedure; maternal immunotherapy or stem cell therapy; maternal malignancy; maternal mosaicism; fetal placental mosaicism; fetal demise; nonviable twin.
In twin dizygotic pregnancies it is not possible to distinguish the condition of the single fetus or to evaluate the aneuploidy of the sex chromosomes. It is, however, possible to find the presence/absence of the Y chromosome. If the presence of the Y chromosome is detected, it is not possible to discern if only one or both fetuses are male. In pregnancies which started as twins or multiple, followed by the spontaneous abortion of one or more fetuses with reabsorption of the gestational chamber (vanishing twin), cell-free fetal DNA of the aborted fetus may also be present in the maternal blood. This could interfere with the quality of the results, causing false positives if the cause of the abortion was due to the presence in the aforementioned fetus of chromosomal aneuploidies affecting one of the investigated chromosomes. Likewise, an inconsistency in the reporting of sex could result (eg male diagnosis, in which the presence of the Y chromosome originates from the aborted fetal DNA). The existence of a tumor condition (metastasis) in the pregnant woman could result in false positive test results. The test is based on the quantification of the cell-free fetal DNA fragments circulating in the maternal blood, which are of placental origin. Therefore, due to conditions of chromosomal mosaicism (frequency: 1-2%), there may be discrepancies in the results (false positive or false negative) that justify the sensitivity and specificity of the test <100%. In particular, the test could produce a positive result (aneuploidy detected), but this chromosomal anomaly could be confined to the placenta due to the chromosomal mosaicism, and therefore the fetus could eventually result with normal karyotype to control in invasive prenatal diagnosis (false positive). Vice versa, the test could produce a negative result (aneuploidy not detected), but due to the chromosomal mosaicism fetal DNA without aneuploidy could be confined to the placenta, and therefore the fetus could eventually result with aneuploid karyotype to control in invasive prenatal diagnosis (false negative). Fetal sex is referred to as masculine or feminine, based on the presence or absence of the Y chromosome, but does not provide information on the presence or absence of the SRY gene. Pregnancies with ultrasound findings suggestive of fetal pathology should be studied with other types of prenatal studies, such as the molecular fetal karyotype on chorionic villi or amniotic fluid, in consideration of the higher detection rate. There is a possibility of identifying with this test abnormality of the sex chromosomes present in the mother (homogeneous or mosaic) that can interfere with the accuracy of the results concerning the fetal sex chromosomes. A result "NEGATIVE - Aneuploidy or undetected structural chromosomal alteration" greatly reduces the chances that the fetus has an aneuploidy or a structural chromosomal alteration at the level of the examined chromosomes, but can not guarantee that the chromosomes are actually normal or that the fetus is healthy. This test can not be performed on women who are carriers of aneuploidy. For the limits outlined above, in case of a positive result, it is recommended to carry out an consultation with a geneticist and confirm the result by analysing the karyotype on amniotic fluid.
In twin pregnancies, it is not possible to distinguish the condition of the single fetus. In pregnancies that started as twins or multiple, followed by the spontaneous abortion of one or more fetuses with reabsorption of the gestational chamber (vanishing twin), free fetal DNA of the aborted fetus may also be present in the maternal blood. This could interfere with the quality of the results, resulting in false-positives. The existence of a tumor condition (metastasis) in the gestant could result in false positive test results due to mutations of the circulating tumor DNA (ctDNA) at the level of genes involved in the carcinogenesis process (eg BRAF, KRAS, NRAS). A "NEGATIVE - Low risk for genetic disease" result greatly reduces the chances that the fetus has the genetic diseases examined, but it can not guarantee that the fetus is healthy. The GeneSAFE™ screening test only identifies mutations with a known pathological significance. The test does not look for variants with benign meaning, i.e. those found in normal individuals and are devoid of pathological significance, and variants with uncertain clinical significance, i.e. those not yet known or characterised by the medical-scientific community. The interpretation of genetic variants is based on the latest knowledge available at the time of analysis. This interpretation could change in the future with the acquisition of new scientific and medical information on the structure of the genome and influence the evaluation of the variants.
