Non-invasive prenatal diagnosis of Down Syndrome

7 October 2008

A new paper in the journal PNAS from a team at Stanford University in the US has been in the news; the paper reports on the identification of Down Syndrome (DS) in a fetus using a maternal blood sample (see BBC news article). Down Syndrome (DS), also called Trisomy 21, is the most common form of aneuploidy – the presence of an abnormal number of chromosomes – that is compatible with life, and is caused by the presence of an additional (third) chromosome 21.

This non-invasive form of testing analyses cell-free DNA or RNA from the fetus, which is found in the mother’s blood during pregnancy from as early as 5 weeks gestation, which was first described in 1997 by Dennis Lo [Lo YMD et al. (1997) Lancet 350: 485-487]. Since then, research and developmental work has been undertaken on a number of potential applications including prenatal testing for certain rare genetic (inherited) disorders, identifying the sex of the fetus (which is relevant for certain sex-linked genetic disorders), and testing for blood group Rhesus D status, which is important for the management of some high-risk pregnancies. In addition, late last month another group in the USA announced early findings on a different method for non-invasive prenatal Down Syndrome testing based on a method described by Lo and co-workers in 2007 that uses mass spectrometry to detect fetal RNA [Lo YMD et al (2007) PNAS 104:13116-13121].

Currently, definitive diagnosis of genetic or chromosomal disorders requires an invasive procedure (amniocentesis or chorionic villus sampling) to sample fetal DNA, which carries a risk of miscarriage of 1-2%. Non-invasive testing not only removes this risk, but can also be performed much earlier in pregnancy. Many groups are working to develop the technique for different applications, including several in the UK.

Hitherto, one of the major technical barriers to testing using fetal DNA in the maternal blood is reliably distinguishing the small amounts of fetal DNA from the large background of maternal DNA present. To date, this problem has been solved primarily by only detecting paternally inherited sequences not otherwise present in the mother, such as DNA from the Y chromosome in the case of male fetuses. However, diagnosis of DS and other aneuploidies requires the detection of an increase or decrease of fetal chromosomal sequences, caused by the presence or absence of a specific chromosome.

Importantly, the approach described in the most recent paper does not require differentiation of fetal versus maternal DNA, and therefore has the advantage that it could work in all pregnant women. It uses a previously characterised technique known as shotgun sequencing to achieve simultaneous high-throughput sequencing of millions of short DNA sequences [Fan HC et al. (2008) PNAS October 6, doi:10.1073/pnas.0808319105]. The researchers then compared the amount of sequences produced from different chromosomes to detect any over- or under-representation caused by aneuploidy in the fetus.

Testing samples from a group of eighteen pregnant women, the researchers were able to correctly diagnose nine cases of trisomy 21 (DS), two cases of trisomy 18 (Edwards Syndrome) and one case of trisomy 13 (Patau Syndrome). The technique could theoretically also detect the rarer forms of trisomy where there is only part of a chromosome lost or gained, although it is noted that this would be technically more challenging because it would require detection of a significantly smaller increase or decrease in the amount of DNA sequences present.

It is notable that the costs of diagnosis using this technique would be quite high, with the authors’ estimating the cost of sequencing at around $700 per sample; however, they correctly observe that the cost of sequencing is likely to drop significantly as newer and faster techniques emerge in the next few years. For example, a US company has just announced that it will offer whole genome sequencing for just $5000 (see Technology Review news article). Although whole genome sequencing of fetal DNA obtained from maternal blood is not feasible, as fetal DNA in the maternal blood is present in short fragments, this twenty-fold reduction from current prices for sequencing is indicative of rapidly falling prices. Moreover, the cost of diagnosis of DS by current invasive methods is also quite high, requiring the expertise of highly trained specialists for invasive sampling and molecular analysis or karyotyping of the fetal DNA.

Comment: This new technique requires assessment in a much larger cohort of women to determine just how effective it is, but the results are an exciting development in the area of non-invasive analysis of fetal DNA. Non-invasive prenatal testing is progressing rapidly, and for this reason an expert Working Group has been convened in the UK to produce a strategy for the implementation of cell-free fetal nucleic acids for different applications within UK clinical services. This includes evaluation of the current status of the technology, consideration of the wider implications of the technique, and determination of what action needs to be taken by the NHS in order to keep track of developments and anticipate how and when it might enter routine antenatal practice and specialist clinical services in the UK.

The PHG Foundation is leading this project (see Our Current Work), and the report and recommendations of this group to the Joint Committee for Medical Genetics of the British Society of Human Genetics, the Royal College of Physicians and the Royal College of Pathologists will be released by the Foundation in January 2009.

Bodies represented in the expert group include the British Maternal & Fetal Medicine Society, the Royal College of Obstetricians and Gynaecologists, the Royal College of Midwives, the UK National Screening Committee, the Human Genetics Commission and the Genetic Interest Group and Antenatal Results and Choices charities, as well as expert scientists and clinicians, NHS managers and policy makers, and experts in law and ethics.

The group’s preliminary conclusion is that before any of these techniques can be used in routine care a number of things need to happen:

  • Techniques need to be thoroughly evaluated and validated in a large number of patients to determine clinical accuracy, and consequently are unlikely to replace invasive testing for some time.
  • Technological development and laboratory standardisation is required to ensure reliable and accurate results.
  • Non-invasive techniques have the potential to replace current multistep Down syndrome screening tests with a single diagnostic test, and both women and healthcare professionals must be fully informed about the implications of these changes.
  • Careful consideration should be given to safeguarding patient autonomy and providing for informed consent.

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