Preimplantation genetic diagnosis : evaluation of results and experiences

Preimplantation genetic diagnosis (PGD) is an established alternative for couples at high risk of having an affected child, with the advantage that the genetic testing is performed at the embryo stage and the couple can thereby avoid a pregnancy termination of an affected foetus. The disadvantage is that an IVF treatment is required, which can be a stressful experience. The main indications for PGD are monogenic disorders and chromosome abnormalities and there is an increasing demand for PGD each year. The PGD process can be divided into three steps 1) The IVF treatment, 2) The biopsy and 3) The genetic analysis. The aim with this thesis was to identify factors of importance for an optimal PGD and to learn more about patient’s experience of PGD in order to improve the advisory procedure and care of these patients. Another aim was to gain more knowledge regarding the segregation of different reciprocal translocations and their influence on fertility.

Carriers of reciprocal translocations are usually healthy but have an increased risk of producing sperm or oocytes with an unbalanced chromosome content which gives them a high risk of repeated miscarriages, infertility and an increased risk to have an affected child. The unbalance arises during meiosis when the sperm and oocytes are formed and are present in every cell in the body in the offspring. However, some abnormalities arise after conception during the early embryo development resulting in mosaicism where some cells have the abnormality and some do not. This was the case in Paper I where germline mosaicism was demonstrated to be the cause of repeated pregnancies with the same unbalanced chromosome abnormality, although karyotypes from both parents initially were interpreted to be normal. Extended investigations with microsatellite markers and FISH analysis revealed the same abnormality in 4-6% of the mother’s fibroblasts. The couple went through four PGD cycles and the abnormality was found in 35% of the embryos. The low level mosaicism in the fibroblasts gave no phenotypic symptoms but since the abnormality seemed to be present at a higher frequency in her gonads, there was a high” hidden” recurrence risk for affected offspring or repeated miscarriages.

In Paper II a linear regression analysis was performed of data from all 569 PGD cycles performed between 1996 and 2009. We found two factors of significant importance for the PGD outcome. Firstly, the age of the woman at stimulation start where women under 36 years were three times more likely to achieve a pregnancy P = 0.003 and odds ratio 3.1 [95% confidence interval (CI) 1.5-6.5]. Secondly, the number of biopsied cells from each embryo where PGD cycles with one cell removal were twice as likely to result in a pregnancy compared to those cycles were two cells had been removed P = 0.0013 and odds ratio 2.55 (95% CI 1.44 – 4.52). Accordingly, we have now introduced an age limit of 40 years at stimulation start for the woman and changed policy to one cell biopsy for almost all indications since 2009.

Paper III was based on statistical analyses of data from a survey study concerning the experience of PGD between June 2005 and 2011. A questionnaire was sent to 222 couples and 146 answered, of which 20% had the experience from a pregnancy termination of an affected foetus, one third had the experience of previous traditional prenatal diagnosis and 35% had given birth to an affected child. The results showed that couples with monogenic disorders choose PGD because of an objection to pregnancy termination for psychological reasons while carriers of chromosome abnormalities opted for PGD because of previous miscarriages. We could confirm that there is an extensive stress associated with PGD and that the couples seemed to have been less prepared for the psychological stress than for the physical stress.

It has previously been suggested that a sperm FISH analysis could predict the PGD outcome and that there is a linear correlation between the proportion of abnormal sperm and the proportion of abnormal embryos. In Paper IV sperm FISH analyses from ten male carriers of different reciprocal translocations was performed in connection with their first PGD and the result from sperm and embryos were compared. We found a difference with an increase of unbalanced embryos compared to sperm with no linear correlation. We could confirm that the number of balanced embryos available for transfer correlates to the pregnancy rate.

In conclusion, PGD is a valuable and preferred reproductive alternative for couples at high risk of having a child with a severe genetic disorder. It is a rapidly developing field worldwide and new techniques as well as new indications are continuously announced. It is of great importance that medical and ethical aspects are considered and up to date before the introduction of new methods and that new techniques are constantly evaluated regarding accuracy and safety, in order to optimise the PGD program.

List of scientific papers

I. Haapaniemi Kouru K, Malmgren H, White I, Blennow E. Hidden mosaicism for a structural chromosome rearrangement: a rare explanation for recurrent miscarriages and affected offspring? Fertil Steril. 2011; 95: 806-808.
https://doi.org/10.1016/j.fertnstert.2010.09.022

II. Haapaniemi Kouru K, Malmgren H, Nordenskjöld M, Fridström M, Csemiczky G, Blennow E. One-cell biopsy significantly improves the outcome of preimplantation genetic diagnosis (PGD) treatment: retrospective analysis of 569 PGD cycles at the Stockholm PGD centre. Hum Reprod. 2012; 27: 2843-2849.
https://doi.org/10.1093/humrep/des235

III. Haapaniemi Kouru K, Syk Lundberg E, Malmgren H, Ingvoldstad C. Preimplantation genetic diagnosis in Sweden: patient’s experience and attitudes. [Manuscript]

IV. Haapaniemi Kouru K, Malmgren H, White I, Rodriguez Sanchez A, Syk Lundberg E. Meiotic segregation analyses of reciprocal translocations in sperm and embryos: no support for predictive value regarding PGD outcome. [Manuscript]