Cell-free fetal DNA and non-invasive prenatal diagnosis.

Currently in the UK, prenatal diagnosis of genetic conditions and Down’s syndrome requires invasive diagnostic tests such as amniocentesis and chorionic villus sampling (CVS). Procedural related miscarriage rates of about 1% have been quoted for these tests which are not usually done before 11 weeks’ gestation. Annually in the UK, 32 000 women have an invasive diagnostic test as a result of other screening tests, indicating that they are at increased risk for their children having Down’s syndrome. Around a further 1500 pregnant women have an invasive test because the fetus is at high risk of a genetic condition. An area that has generated much interest in recent years has been in isolating fetal genetic material present in maternal blood as a target for non-invasive prenatal diagnosis (NIPD). In 1997 Lo et al identified cell-free fetal DNA in the maternal circulation. This finding has generated research in the development of clinical applications based on analysing this fetal genetic material for NIPD. Cell-free fetal DNA represents extracellular DNA which originates from trophoblastic cells. However, the vast majority of cell-free DNA in maternal blood originates from the mother, with cell-free fetal DNA representing only 3% of the total cell-free circulating DNA in early pregnancy rising to 6% in late pregnancy. After delivery, cell-free fetal DNA is rapidly cleared from the maternal circulation, making it specific to that pregnancy. As the fetal DNA is swamped by the presence of cell-free maternal DNA, the challenge has been to separate the fetal from the maternal cell-free DNA. Various methods have been used for this, including using methods based on the fact that the fetal DNA is shorter than maternal DNA. However, it is still not possible to extract pure fetal DNA and so currently prenatal diagnosis using cell-free fetal DNA is limited to the detection or exclusion of genetic sequences that are not present in the mother, that is, determination of fetal sex, fetal rhesus D status in D-negative supportive studies have demonstrated its diagnostic sensitivity which ranges from 95–100% with specificities of over 99%. The analysis of cell-free fetal DNA is now the method of choice in the management of pregnancies at high risk of haemolytic disease of the newborn. For several years now the management of mothers with a history of haemolytic disease of the newborn or antibodies to rhesus D has involved non-invasive prenatal diagnosis to determine the fetal rhesus D status. If the fetus is predicted to be D-negative no further monitoring is required, but if it is Dpositive close monitoring in a fetal medicine unit is needed. These tests have been done using labour intensive laboratory methods but, if high throughput technology can be developed, there is potential for use routinely. Consider that in the UK anti-D prophylaxis is given to all D-negative women usually in the third trimester, and after feto-maternal haemorrhage as well as after birth if the baby is D-positive. However, around 40% of D-negative women carry a D-negative fetus and so do not require anti-D prophylaxis. Anti-D is a human blood product and thus reducing its use in women who do not need it will reduce potential exposure to hepatitis C and prion type diseases as well as NHS expenditure. A high throughput method for reliable analysis of fetal rhesus D status at 28 weeks has been described recently and the challenge now is to refine the technology for routine use earlier in pregnancy and thus avoid unnecessary administration of anti-D.