PRENATAL DIAGNOSIS

Prenatal diagnosis refers to all the tests that take place during pregnancy, in order to uncover serious, mainly genetic disorders, in the fetus, occurring with an overall frequency of about 1/20 pregnancies. Prenatal testing includes:

• monitoring the proper progression of pregnancy in terms of the mother and the fetus and undertaking the necessary tests for this purpose
• diagnosis of serious malformations of the fetus with the aid of ultrasound
• risk assessment for possible genetic defects, mainly chromosomal abnormalities, such as Down syndrome, especially in the 1^st trimester of pregnancy, through the combined use of sonographic markers, such as nuchal translucency – NT and biochemical markers in maternal blood (PAPP-a and β-hCG in the 1^st trimester)
• diagnosis of chromosomal abnormalities, such as Down syndrome and other chromosomal abnormalities
• diagnosis of genetic diseases due to gene mutations, by direct analysis of fetal genetic material

InterGenetics has been active in prenatal diagnosis from its inception 39 years ago and this filed involves the largest part of our work, having successfully completed more than 95,000 prenatal cases.

In every case we have applied the most comprehensive testing available at the time, with many options and in-depth analysis through the latest molecular genetics and cytogenetics techniques. Where necessary, genetic testing was accompanied with in-depth genetic counseling.

Several innovative applications include the extended prenatal panel – EPP and later, the application of molecular karyotype (array CGH) as a routine standalone test in prenatal chromosomal testing.

We should also remember that there are more than 8000 registered diseases caused by gene mutations, such as thalassemia and cystic fibrosis.

Currently, there is no mass genetic screening test (as in the case of karyotype analysis for chromosomal abnormalities) which may detect prenatally all of the above single gene disorders.

Prenatal testing is therefore currently applied only families where we already know that one or both parents are carriers of mutations or are affected and thus a prerequisite is the prior knowledge of the pathogenic mutations in the parent/parents, before we can proceed with diagnosis in the fetus.

Genetic testing requires a fetal DNA sample, usually obtained from chorionic villi or amniotic fluid cells, either directly or after culture. In all cases involving prenatal diagnosis for gene disorders, we apply parallel analysis of DNA polymorphisms from a maternal blood sample, in order to exclude the possible presence of maternal cell contamination in the fetal sample.

For this reason, the fetal sample should always be accompanied with 1-2ml peripheral blood of the mother.

Prenatal diagnosis involves both chromosomal and gene disorders and may be performed in any trimester, from the 10^th week of pregnancy onwards.

A first assessment of the course of pregnancy is usually performed done by the obstetrician through an ultrasound examination in the sixth week of pregnancy.

At the same time, a general check-up is performed, which includes: complete blood count, blood group, rhesus, glucose, urea, urine analysis, hemoglobin electrophoresis and HbA2 quantitation for β-thalassemia, serum iron and ferritin, Toxoplasma (Toxo-IgG & Toxo-IgM), Cytomegalovirus (CMV-IgG & CMV-IgM), Rubella (Rubella-IgG & rubella-IgM), Hepatitis B antigen (HBsAg), Hepatitis C (HCV), AIDS virus (HIV 1 + 2), thyroid hormones, etc..

According to established international guidelines, carrier detection for cystic fibrosis (for at least 85% of pathogenic classic CF mutations in the Greek population) should also be performed in a parent.

This test, together with the assessment for thalassemias, is performed once in the first pregnancy of the couple and ensures that they are not at risk of having affected children with these common genetic diseases.

At the same time, the obstetrician will investigate the family history, with the aim of identifying any troubling issues associated with congenital defects, mental retardation, or other genetic diseases in the families of both parents.

In case any problems are identified, the obstetrician and/or the parents should consult a clinical geneticist for further evaluation and testing, such as chromosomal analysis, investigation of gene mutations in the affected or other members of the family.

These test may be necessary for preventing the birth of an affected child and should be performed early in pregnancy or, even better, before pregnancy.

Following the first evaluation by the obstetrician, at about 10 to 11 weeks of pregnancy, a screening test is applied in order to evaluate the statistical risk for chromosomal abnormalities and other genetic diseases of the fetus as well as for other detectable congenital malformations and for NT measurement, through a specialized ultrasound examination in combination with measurement of biochemical markers in maternal blood (placental protein – PAPP-a and beta-chorionic gonadotrophin – βhCG).

The risk assessment through combination of ultrasound – NT, biochemical markers and the mother’s age, may lead to a high risk mainly for chromosomal abnormalities or other genetic diseases or congenital malformations in the fetus.

In the case of a high risk, it is recommended to proceed to invasive testing, by obtaining chorionic villi in the 1^st trimester of pregnancy (11 to 13 wks) or amniotic fluid in the 2^nd trimester of pregnancy (17 to 23 wks), in order to perform classic karyotype either through morphological analysis of the chromosomes (report in 14 days), or with the more recent method of molecular karyotype-aCGH, which detects many more chromosomal abnormalities involving hundreds of known syndromes and has also the advantage of a shorter report time (4-5 days).

In the case of pregnancies at ‘medium’ risk following 1^st trimester screening, there are those who currently recommend the option of performing a more accurate screening test, known as Non-Invasive Prenatal Testing – NIPT, which is a non-diagnostic test practically for Down syndrome only, which will not detect all possible chromosomal abnormalities in the fetus.

This test is performed from a maternal peripheral blood sample, thus avoiding invasive testing.

Currently, a widely accepted position in prenatal chromosomal diagnosis is to recommend testing through prenatal molecular karyotype-aCGH, since the risk for pregnancy loss though amniocentesis is nowadays internationally quoted as being less than 1 in 500, while molecular karyotype will diagnose approximately 3% (1 in 30) embryos as being affected (Konialis C., Pangalos C., 2015).

In cases were ultrasound examination will reveal mild or severe structural malformations in the fetus, mainly in the 2^nd trimester of pregnancy, and following the exclusion of a fetal chromosomal abnormality by prenatal molecular karyotype, InterGenetics is currently in a unique position to perform in-depth fetal genomic testing for mutations in ~760 genes, which have been implicated in congenital abnormalities such as: severe skeletal malformations, heart and renal defects, structural brain anomalies, ambiguous genitalia, and several other abnormalities.

This test is called Fetalis^® and may lead to the discovery of a potentially serious genetic mutation, linking the observed fetal malformations to a recognized genetic disorder/syndrome, which may also include intellectual disability.

Therefore, the Fetalis^® test could be proposed whenever there are troubling ultrasound findings in the fetus and following a normal prenatal molecular karyotype result.

If, as already mentioned, the family history reveals the possible presence of a genetic disease in the family, then it is necessary to detect the underlying gene mutations in the patient and/or other members-carriers.

This is deemed necessary in order to counsel the couple regarding the options of prenatal diagnosis during pregnancy or preimplantation genetic diagnosis (PGD) following IVF.

In such cases, prenatal testing is preferably performed in the 1^st trimester by obtaining chorionic villi and subsequent analysis of fetal DNA, since the risk for the fetus being affected in these cases is high (25-50%) and pregnancy may be terminated, if necessary, at an early stage. However, genetic testing may be also performed through an amniotic fluid sample drawn in the 2^nd trimester of pregnancy.

It is highly recommended that all the above procedures are accompanied by proper genetic counseling by a qualified clinical geneticist, both before and after genetic testing.

It is estimated that approximately 1 in 50 newborns are affected with a severe genetic disease and the vast majority of these are due to chromosomal abnormalities or single-gene disorders.

These genetic diseases may be the result of a new genetic aberration in the fetus, i.e. not inherited from a parent, or the result of genetic mutations inherited from unsuspecting carrier parents.

InterGenetics is currently in a position to offer a newly developed comprehensive prenatal test, the FetalSafe^® test, complementing prenatal chromosomal diagnosis through molecular karyotype-aCGH with parallel genomic testing via Next Generation Sequencing – NGS of approximately 300 genes, associated with several severe gene disorders, occurring with a combined frequency of approximately 1 in 300 to 1 in 400 births, not detectable by molecular karyotype-aCGH.

Therefore, the FetalSafe^® genomic test, in combination with molecular karyotype-aCGH, constitutes an expanded, comprehensive and rapid (1 week) prenatal genetic test, detecting more than 400 severe chromosomal and gene disorders and may be applied in all pregnancies requiring prenatal diagnosis.

InterGenetics performs all invasive and non-invasive prenatal tests relating to all requirements of prenatal testing and the tests are detailed below. We wish to remind that invasive testing in the 1^st trimester relies on obtaining embryonic cells through chorionic villi sampling, while in the 2^nd and 3^rd trimester through amniotic fluid sampling-amniocentesis. In selected cases, there is also the possibility of obtaining fetal blood or other fetal tissues, e.g. skin.