An Update on PGD: Where we are today

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1 An Update on PGD: Where we are today Joyce Harper UCL Centre for PG&D and CRGH Institute for Womens Health University College London

2 Overview What is PGD/PGS How we do it Disadvantages and advantages Future use of these procedures ESHRE world data Ethics

3 What is Preimplantation Genetic Diagnosis? Selection of genetically normal or carrier embryos from couples with known inherited mutations (nuclear DNA) chromosomally balanced embryos from couples with known chromosomal rearrangements Aim to transfer an embryo that will develop into a healthy child Couples Mostly fertile Diagnosis Specific Undiagnosed embryos never transferred Ethics / controversy Disorders / predispositions / traits New techniques will test for many things

4 PGD COUPLE CARRY DISEASE Have affected child Other family members affected Diagnosed as an adult OPTIONS Remain childless Reproductive roulette Prenatal Diagnosis PGD Gamete donation Adoption

5 Indications for PGD Single gene disorders Recessive, dominant, X-linked Triplet repeat disorders Late onset disorders Key male female carrier affected Chromosome abnormalities Translocations Other inherited chromosome abnormalities Sexing for X linked disease Specific diagnosis Sexing Key male female Carrier Normal chromosome 7 p.f508del mutation? mutation Normal Normal Carrier Carrier Normal Carrier Carrier Cystic fibrosis (CF)

6 What is Preimplantation Genetic Screening? Selection of Euploid embryo (oocyte) Aim To improve IVF delivery rate Couples Infertile or subfertile Diagnosis Screening of few / many / all chromosomes Undiagnosed embryos can be transferred Ethics / Controversy Randomised controlled trials have not proved efficacy Best approach still debated Controversial technique (Harper et al, 2008, 2010a) and many others

7 Indications for PGS Advanced maternal age (>35/38) Recurrent IVF failure (2 or more) Recurrent miscarriage (karyotypes normal) Severe male factor

8 Stages of PGD/PGS 1. Removal of some of embryo - biopsy 2. Diagnosis from 1-5 cells

9 Biopsy 1. Polar Body 2. Cleavage Stage 3. Blastocyst

10 Polar Body Biopsy a) b) c) d)

11 Cleavage stage biopsy Generated by Foxit PDF Creator Foxit Software

12 Blastocyst biopsy McArthur et al., Fertil Steril 2005

13 Blastocyst biopsy Generated by Foxit PDF Creator Foxit Software

14 Diagnosis Polymerase chain reaction (PCR) Analyse specific mutation Single-gene defects Fluorescent in situ hybridisation (FISH) Analyse chromosomes Sexing for X-linked disease Chromosome abnormalities PGS Arrays Analyse chromosomes and/or genes Array CGH SNP arrays

15 Mosaicism Present at all stages of preimplantation development 50% cleavage stage embryos Growing evidence in blastocysts Wells et al 33% Problematic for PGD and PGS

16 Misdiagnosis Wilton et al, 2009, Human Reproduction Allele dropout Contamination Using wrong test Mixing up embryos Unprotected sex

17 F-PCR for myotonic dystrophy Mother - marker (18) Mother normal myotonic allele Abnormal allele refractory to PCR Father - marker (4) 170 (19) Father two normal myotonic alleles

18 PGD for myotonic dystrophy marker Cell 1 normal DM alleles marker Cell 2 normal DM alleles

19 Sexing embryos for PGD FISH analysis of Interphase nuclei Chromosome X Chromosome Y Chromosome 16 Normal Female Normal Male

20 PGD of Chromosome Abnormalities: Robertsonian Translocation Carrier 45,XX,der(13q;21q)(q10;q10) Chromosome 21 Chromosome 13 Normal for Chromosomes 13 & 21 Monosomy 13 & 21

21 PGS X (gold), Y (blue), 13 (green), 18 (aqua) and 21 (red) 13(red), 16(aqua), 18(blue), 21(green) and 22(gold) Courtesy of Santiago Munne

22 Disadvantages of PGD/PGS Patients have to go through IVF Cost All embryos may be affected Making diagnosis from 1-2 cells Have been misdiagnosis Success rate lower than IVF (even for PGS) Typical case 12 oocytes 10 fertilised 8 for biopsy 7 diagnosis result 5 abnormal 2 normal 1 good embryo 1 poor embryo

23 Advantages of PGD/PGS PGD Alternative is prenatal diagnosis so PGD avoids TOP Start pregnancy knowing embryo does not carry disease PGS No advantage until show improves delivery rates Closure after many failed IVFs Data for repeated IVF failure, recurrent miscarriage not available

controlled trials to see if valid procedure with clinical significance NOT cleavage stage

24 Future of PGD/PGS PGD New technology to allow diagnosis of more disorders Whole genome amplification SNP arrays and array-cgh Blastocyst biopsy and vitrification PGS Randomised controlled trials to see if valid procedure with clinical significance NOT cleavage stage biopsy, NOT FISH Try polar body or trophectoderm Try arrays (either SNP or array-cgh) Blastocyst biopsy and vitrification

25 ESHRE PGD Consortium - Aims To survey the availability of PGD To collect prospectively and retrospectively data on the accuracy, reliability and effectiveness of PGD To initiate follow-up studies of embryos, pregnancies,... To produce guidelines and recommended PGD protocols To formulate a consensus on the use of PGD To educate in the science of genetics and reproduction

26 Consortium guidelines All published in Human Reproduction Harton, G, Braude, P, Lashwood, A, Schmutzler, A, Wilton, L, and Harper, JC. ESHRE PGD Consortium-Best Practice Guidelines for Organization of a PGD Center for Preimplantation Genetic Diagnosis/Screening (PGD/PGS) Harton, G, Coonen, E, Pehlivan, T, Vesela, K, Harper, JC and Wilton, L. ESHRE PGD Consortium Best Practice Guidelines for FISH-Based Preimplantation Genetic Diagnosis (PGD) Harton, G, DeRycke, M, Fiorentino, F, Moutou, C, SenGupta, S, Traeger- Synodinos, J, Sermon, K, and Harper, JC ESHRE PGD Consortium Best Practice Guidelines for DNA amplification based Preimplantation Genetic Diagnosis (PGD) Harton, G, Cristina Magli, Kersti Lundin, Markus Montag, Lemmen, J and Harper, JC. ESHRE PGD Consortium/Embryology Special Interest Group-Best Practice Guidelines for Polar Body and Embryo Biopsy for Preimplantation Genetic Diagnosis/Screening (PGD/PGS).

27 Data Collection 13 years ( ESHRE Preimplantation Genetic Diagnosis (PGD) Consortium (1999). Preliminary assessment of data from January 1997 to September 1998 ESHRE PGD Consortium Steering Committee. Human Reproduction, 14: ESHRE PGD Consortium Steering Committee (2000) ESHRE Preimplantation Genetic Diagnosis (PGD) Consortium: data collection II (May 2000). Hum. Reprod. 15, ESHRE PGD Consortium Steering Committee (2002) ESHRE Preimplantation Genetic Diagnosis (PGD) Consortium: data collection III (May 2001), Hum Reprod., 17, Sermon, K., Moutou, C., Harper, J., Geraedts, J., Scriven, P., Wilton, L., Magli, M.-C., Michiels A, Viville, S., De Die, C. (2005) ESHRE PGD Consortium data collection IV: May-December 2001, Human Reproduction, 20(1): Harper, J.C, Boelaert, K, Geraedts, J., Harton, G., Kearns, WG, Moutou, C., Muntjewerff, N., Repping, S, SenGupta, S, Scriven, P.N., Traeger-Synodinos, J, Vesela, K, Wilton, L, Sermon, K.D. (2006) ESHRE PGD Consortium data collection V: Cycles from January to December 2002 with pregnancy follow-up to October 2003, Human Reproduction, 21, 3-21 Sermon, K.D, Michiels, A, Harton, G, Moutou, C, Repping, S, Scriven, P.N, SenGupta, S,Traeger-Synodinos, J, Vesela, K, Viville, S, Wilton, L, Harper, J.C (2007) ESHRE PGD Consortium data collection VI: Cycles from January to December 2003 with pregnancy follow-up to October 2004, Hum Reprod. 22(2): Harper, JC, De Die, C, Goosens, V, Harton, G, Moutou, C, Repping, S, Scriven, P., SenGupta, S.,Traeger- Synodinos, J., Viville, S., Wilton, L., Sermon, K.D. (2007) ESHRE PGD Consortium data collection VII Cycles from January to December 2004 with pregnancy follow-up to October Hum Reprod Goossens, V, Harton, G, Moutou, C, Scriven, PN, Traeger-Synodinos. J, Sermon, K, Harper, JC (2008) ESHRE PGD Consortium data collection VIII: Cycles from January to December 2005 with pregnancy follow-up to October 2006, Human Reproduction, 23(12): Goossens, V, Harton, G, Moutou, C, Traeger-Synodinos, J, Van Rij, M and Harper, JC (2009) ESHRE PGD Consortium data collection IX: cycles from January to December 2006 with pregnancy follow-up to October 2007 Harper, JC Coonen, E, De Rycke, M, Harton, G, Moutou, C, Pehlivan, T, Traeger-Synodinos, J, Van Rij, M, Goossens, V, ESHRE PGD Consortium data collection X: Cycles from January to December 2007 with pregnancy follow-up to October 2008

Evolution of cycle data 6000 5000 4000 3000 2000 PGS social sexing sexing for

28 Evolution of cycle data PGS social sexing sexing for X-linked disorders chromosomal abnormalities monogenic disease Harper et al, HR, 2010

29 Pregnancy rate by centre Data X % clinical pregnancy OR %/OR ,73% 10 - >500 OR OR OR 2-20 OR 1 OR Success rates tend to be higher in more active centres But some active centres below 22%

Top 10 monogenic disorders Recessive Cystic Fibrosis β-thalassaemia Spinal muscular atrophy Sickle cell disease Dominant Myotonic dystrophy

30 Top 10 monogenic disorders Recessive Cystic Fibrosis β-thalassaemia Spinal muscular atrophy Sickle cell disease Dominant Myotonic dystrophy Huntington s disease Charcot-Marie-Tooth disease Sex linked (specific diagnosis) Duchenne muscular dystrophy Fragile X syndrome Haemophilia

31 Most commonly tested disorders

32 Consortium working groups Misdiagnosis monitoring and audit: Chair Joanne Traeger-Synodinos Accreditation: Chair Katerina Vesela Database: Chair Celine Moutou Guidelines: Chair Gary Harton Molecular methods: Chair Francesco Fiorentino New - arrays

33 Generated by Foxit PDF Creator Foxit Software Accreditation External quality assessment ISO UK NEQAS

34 Consortium update Tens years of PGD data collection Harper et al Human Reproduction Update In press Check

35 Ethical dilemmas in PGD Saviour sibling Sex selection Non life threatening who decides? SNP arrays can test for everything in one go

36 UCL Centre for PG&D and CRGH

The role of the PGD Consortium. Overview. ESHRE PGD Consortium - Aims

The role of the PGD Consortium Joyce Harper Chair of the ESHRE PGD Consortium Overview Aims Membership Data Pregnancies Guidelines Training courses/education/post congress workshops Working groups PGS