Preimplantation Genetic Diagnosis (PGD) for Huntington’s Disease using Grand Parental Exclusion

James Marshall, Sydney IVF, 4 O’Connell St, Sydney 2000

Preimplantation Genetic Diagnosis (P.G.D.), (the genetic screening of the early embryo before it implants and in the uterus), is available to couples who have a family history of Huntington Disease (HD). The HD gene is on chromosome 4 and HD is an autosomal dominant disorder; meaning you only require the HD gene on one of the two chromosome 4s to be affected in order to get the disorder.

The PGD screening is available in two forms:

1.      Disclosure, where one of the couple is aware they carry the Huntington Disease expansion, or

2.      Undisclosure, (or Exclusion), where any embryos who inherit the HD gene from a known affected grandparent are excluded, without knowing whether it is the affected or unaffected copy of the HD gene from the affected grandparent. Undisclosure is used when a patient whose parent is affected with HD does not wish to know their own HD status, but does wish to have a child that will not be affected.

What is PGD? PGD is a group of tests used with in-vitro fertilisation (IVF) to diagnose and exclude genetic abnormalities in the early preimplantation embryo. Embryos created using IVF are cultured to five days of development, the blastocyst stage. Where 3-5 cells are removed from the outer layer (the trophectoderm - following transfer and implantation this layer forms the placenta).

The DNA is extracted from these placental stem cells and analysed to determine the absence or presence of the chromosomal region containing the genetic disorder. The analysis usually included tests for both the mutation itself and for linkage markers, which are short identifying DNA sequences close to the gene or affected region. In some disorders, and HD Undisclosure is an example, the mutation itself is not tested for, only linkage marker testing is used, utilising linkage to the affected grandparent. In either case the reliability of the tests is usually about 99%.

After analysis an ‘unaffected’ embryo, that is an embryo with the desired genetics, is transferred to the uterus, and any spare ‘unaffected’ embryos are frozen for use in subsequent pregnancies.

The analysis for HD Undisclosure used two to four polymorphic linkage markers (PLMs) situated close to the HD gene. PLMs are small lengths of DNA along the chromosome which may differ in size between chromosomes and between people.

They are used in forensics for DNA fingerprinting of a person’s whole genome; we used them to build a mini DNA fingerprint around or close to a mutation or affected regions. PLMs used must be informative for both of the chromosome 4 regions containing the HD gene from the affected grandparent.

The test for each PLM is multiplexed (that is made to work together) in a single test. That test is used on the extracted DNA from the 3-5 cells removed from each embryo and the absence or presence of the chromosome from the affected grandparent can be determined. On average one in two embryos will carry the chromosome from the affected grandparent and will therefore be excluded. The test takes about 4-5 hours to complete, unlike the standard diagnostic test for HD which is more complex and takes 1-3 days to complete, and an embryo can be transferred to the uterus the same day as testing occurs.

At Sydney IVF six couples have undergone stimulated IVF cycles with PGD to exclude Huntington Disease, with five having embryo transfers and four pregnancies. Three couples have given birth to five babies with one couple soon to give birth.

Further information can be obtained by contacting:

Mail                  Sydney IVF, Level 11

                        4 O’Connell Street, SYDNEY, 2000

Telephone:             (02) 9221-5964

Email:                jim.marshall@sivf.co.au

“Newsletter”

Australian Huntington’s Disease Association (Qld) Inc

April 2004