The prevalence and risk of Down Syndrome in monozygotic and dizygotic multiple pregnancies in Europe: implications for prenatal screening B. Boyle, J.K. Morris, R. McConkey, E. Garne M. Loane, MC. Addor, M. Gatt, M. Haeusler, A. Latos-Bielenska, N. Lelong, R. McDonnell 1, C. Mullaney, M. O’Mahony and H Dolk.

Background Both dizygotic twinning and Down Syndrome are associated with older maternal age Prenatal screening uses maternal age, along with ultrasonic and biochemical tests to estimate risk of Down syndrome and so aid the decision to proceed to more invasive tests Lack of precise age specific risks in multiple pregnancies may potentially lead to inaccurate estimates of risk of Down Syndrome The rate of fetal loss after amniocentesis is twice as high in multiple as in singleton pregnancies Technically more difficult to carry out an amniocentesis on a multiple pregnancy

Epidemiological Literature comparing risk of Down Syndrome in multiple compared to singleton pregnancies: per baby, analysis carried out on live births Author Measures of association Type of multiple birth Number of DS cases from multiple births Estimate (95% CI) Adjusted for maternal age Hay 1970Odds RatioUnlike sex230.81no Odds RatioLike sex330.56no Kallen 1986*Relative RiskAll twins ( )no Doyle 1970Relative RiskAll twins ( )Yes Tang 2006Relative RiskAll Twins ( )Yea Kallen included stillbirths

Conflicting advice available for prenatal screening Risk per fetus Similar, but in monozygotic pregnancy both fetuses will be affected Gonce (2005), Sebire (2006) Risk per pregnancy Similar as the risk per fetus is lower Wald (2005) Cuckle (1998) UK (NICE) guidelines Risk per pregnancy is higher, but they do not state by how much French guidelines Use singleton risk estimates, but do not state if this is per fetus or per pregnancy

Aim of the Study To calculate, using a large European birth population, the maternal age-specific prevalence of DS in monozygotic and in dizygotic pregnancies and to explore the risk for each relative to singleton pregnancies

Methods Data from 10 EUROCAT congenital anomaly registries in 8 countries, including the National Down Syndrome Cytogenetic Register, a UK based registry affiliated to EUROCAT which provided case data especially for this study The total study population was 14, 849,746 births between 1990 and 2009, of which 2.89% were from multiple births. EUROCAT DQI used to select registries with good ascertainment of Down Syndrome Included all livebirths, stillbirths and terminations for fetal anomaly (TOPFA)

Methods Correction factor was used to correct TOPFA cases for probability of survival to 20 weeks Cases from like sex concordant pairs were considered to be monozygotic (3 unlike sex concordant pairs) Denominators were available by maternal age, multiple birth and like and unlike sex status Weinberg’s rule: total twin pairs – 2(unlike sex pairs) = monozygotic twins was used to estimate zygosity by maternal age in the population Statistical methods: Poisson regression adjusting for country and maternal age where appropriate

Prevalence per 10,000 births of DS cases from singleton and multiple pregnancies, and by 5 years of maternal age for England and Wales and for the rest of Europe separately

Main findings per fetus The risk of Down Syndrome for a fetus from any multiple birth relative to a singleton birth when adjusted for country and maternal age was: 0.58 (95%CI ). Only one fetus of a mother aged over 44 years was diagnosed with Down Syndrome. The expected number was 55.

Risk of Down Syndrome, per pregnancy, for monozygotic and dyzygotic pregnancies relative to singleton pregnancies, corrected for survival to 20 weeks’ gestation, by maternal age

Main findings per pregnancy The risk of Down Syndrome for a monozygotic pregnancy relative to a singleton pregnancy when adjusted for country and maternal age was: 0.34 (95% CI 0.25 – 0.44) Both fetuses will be affected The risk of Down Syndrome for a dizygotic pregnancy relative to a singleton pregnancy when adjusted for country and maternal age was: 1.34 (95%CI 1.23 – 1.46) If the risk per fetus was the same as for a singleton pregnancy this risk would be 2 as each fetus has an independent risk of Down Syndrome

Discussion Twin, and mainly monochorionic pregnancies are more fragile than singleton pregnancies. When combined with a embryo / fetus with Down Syndrome losses in pregnancy increase. Prenatal screening and TOPFA are less common for Down Syndrome cases from multiple pregnancies than for singletons at similar maternal ages Assisted reproductive therapies: With frozen embryos the egg was harvested from the mother or from a donor at an age which is less than that of the mother at the index pregnancy Embryos with Down Syndrome, even when there is not pre-implantation diagnosis may be more fragile than other embryos and therefore are not selected for implantation

Conclusions Fetuses from twin and higher order multiple pregnancies in mothers who are over the age of 44 are very unlikely to be diagnosed as Down Syndrome The risk of Down syndrome per fetus is lower in multiple than in singleton pregnancies at all maternal ages The risk of Down Syndrome in monozygotic and dizygotic relative to singleton pregnancies described here should be considered in both genetic counselling and prenatal diagnosis

Acknowledgements: Co-authors: B. Boyle, J.K. Morris, R. McConkey, E. Garne M. Loane, MC. Addor, M. Gatt, M. Haeusler, A. Latos- Bielenska, N. Lelong, R. McDonnell 1, C. Mullaney, M. O’Mahony and H Dolk. B Boyle was funded through a Northern Ireland Research and Development Studentship through the Northern Ireland Public Health Agency.. EUROCAT is co ‐ funded by the EC, under the framework of the EU Health Programme 2008 ‐ 2013, Grant Agreement (Executive Agency for Health & Consumers).

Boyle B, Morris J, McConkey R, Garne E, Loane M, Addor M-C, Gatt M, Haeusler M, Latos- Bielenska A, Lelong N, McDonnell R, Mullaney C, O'Mahony M and Dolk H Prevalence and risk of Down syndrome in monozygotic and dizygotic multiple pregnancies in Europe: implications for prenatal screening. British Journal of Obstetrics and Gynaecology. 121(7):809-19; discussion 820, 2014 Jun.