Clinical experience of PB biopsy and CGH micro-array in poor prognosis IVF patients

1. Clinical experience of PB biopsy and CGH micro-array in poor prognosis IVF patients Stuart Lavery Consultant Gynaecologist Director IVF Hammersmith Hammersmith and Queen Charlotte’s Hospitals Imperial College London

2. Clinical experience of PB biopsy and CGH micro-array in poor prognosis IVF patients A new chapter in PGS? Stuart Lavery Consultant Gynaecologist Director IVF Hammersmith Hammersmith and Queen Charlotte’s Hospitals Imperial College London

3. Background What is the evidence? Clinical experience Ethical challenges

4. Preimplantation genetic diagnosis (PGD) is a form of very early prenatal diagnosis. The technique combines assisted reproductive technology with molecular and cyto-genetics to allow the identification of abnormalities in embryos prior to implantation. Couples at risk of having children with serious genetic diseases can have unaffected embryos transferred to the uterus, eliminating the need for prenatal diagnosis and termination of pregnancy.Screening of preimplantation embryos for aneuploidy (PGS) may ameliorate the effect of female age on reproduction: improving pregnancy rates and decreasing the chance of multiple birth and miscarriage.

5. Background • 20% of all UK births are to women >35 • 2009: 706 248 live births in England and Wales, 114 288 women aged 35-39, and 26 976 births to women over 40, 71 over 50 • Background of media sensationalism • Under-explored in serious discussion

6. Background • Pregnancy and live birth rates decline rapidly with advancing age • Main factor is oocyte quality: • Age <35: 32% embryos abnormal • Age 35-37: 42%, >37:53% • IVF failure x3: 54% embryos abnormal • IVF failure x5: 63%

7. Hypothesis: screening oocytes and preimplantation embryos should increase pregnancy rates, decrease miscarriage and multiple pregnancy rates and help prevent the birth of affected children

8. Hypothesis: screening oocytes and preimplantation embryos should increase pregnancy rates, decrease miscarriage and multiple pregnancy rates and help prevent the birth of affected children Lavery S, El-Shawarby SA, Moissidou M, Taylor D, Turner C, Lavender B, Trew G, Margara R, Winston R. Live birth following preimplantation genetic screening for trisomy 21. Should aneuploidy screening be offered to all older patients undergoing IVF? Hum Fertil

9. What is the evidence? • Sound, self-evident, easily understood hypothesis • Which patients: advanced maternal age, recurrent implantation failure, recurrent miscarriage, previous aneuploidy, MESA/TESE ? • Different biopsy technique:1 or 2 cells? • Which chromosomes and how many? • Multiple case series, retrospective case controlled, prospective randomised controlled trials, meta-analysis

10. What is the evidence? To date there is insufficient data to determine whether PGS is an effective intervention in IVF/ICSI for improving live birth rates. Available data on PGS for advanced maternal age showed no difference in live birth rate and ongoing pregnancy rate. However, only two randomised trials were found, of which one included only 39 patients. For both studies comments on their methodological quality can be made. Therefore more properly conducted randomised controlled trials are needed. Until such trials have been performed PGS should not be used in routine patient care.

11. Is PGS dead? Yes, for FISH on cleavage stage embryos

12. Is PGS dead? Yes, for FISH on cleavage stage embryos What about for alternative genetic techniques looking at different stages of biopsy?

13. Is PGS dead? Yes, for FISH on cleavage stage embryos What about for alternative genetic techniques looking at different stages of biopsy? We just don’t know…

14. New Scientific Developments • Whole Genome Amplification (WGA) • Comparative Genomic Hybridisation (CGH) • Metaphase spread • Micro-array

15. Euploid -4, +5, +7, -9, +17, -22, +X -8p, -15 +3, +15

16. CGH Aneuploidy ScreeningPreliminary Evidence • Blastomere biopsy, CGH metaphase and cryopreservation Wilton 2003 • PB biopsy CGH microarray Obradors 2008 • 45 couples, mean age 38, 2.4 previous failed cycles • Blastocyst biopsy: CGH 24 chromosomes • 94% embryos diagnosed • 51% aneuploid • Implantation rate69% • Pregnancy rate 82% Schoolcraft Nov 2009 • ESHRE call for proof of concept trial of PB biopsy and CGH microarray Geraedts Dec 2009

17. ESHRE trial of PB biopsy and CGH microarray ‘Proof of principle’ • 2 centres: 226 eggs, 41 couples in 42 cycles, average maternal age 40 • 177 eggs analysed: 34 euploid, 122 aneuploid (69%) • 19 cycles, all eggs aneuploid (45%) • 37 embryos transferred in 23 cycles, 8 clinical pregnancies, 27% implantation rate/ET • PB1 72% aneuploidy in eggs • PB1+2 89 % aneuploidy in eggs • Reliable and accurate Geraraedts 2010

18. Recent Work: why not just culture to blastocyst? • Aneuploid embryos cannot be distinguished from euploid embryos by culturing to blastocyst Cater 2011 • 554 blastocysts: 57% were aneuploid, 41% were mosaic, 72% of most advanced were male Alfarawati 2011

19. CGH Aneuploidy ScreeningIVF Hammersmith experience • 93 cycles started • 20 patients age 36 and under • 61 patients 37 and above • 55 patients had >3 failed implantations • all 93 got to egg collection • 89 had polar body 1 biopsied (96%) • 12 patients had non-fertilisation (13%)

20. CGH Aneuploidy ScreeningIVF Hammersmith experience • 629 eggs injected • 421 fertilised (67%) • 437 PB analysed • No result 56 (13%) • Euploid 84 (19%) • Aneuploid 297 (68%) • All PB aneuploid in 14 cycles (15%)

21. CGH Aneuploidy ScreeningIVF Hammersmith experience • 85 embryos transferred in 53 cycles 57% • Clinical pregnancy rate/cycle started 16/93 17% • Clinical pregnancy rate/ET 16/53 30% <37 years CPR/ET 5/14 36% >37 years CPR/ET 11/39 28% • 1 ectopic, 2 miscarriages, 5 livebirths, 8 ongoing pregnancies

22. Comparison with matched controls CGH Controls Age 40 40 40 Previous cycles 2.8 - 2.5 CPR/cycle started 17% 25% 16% CPR/ET 30% 31% 20%

23. Impressions • High attrition rate, low success rate • Strong patient demand despite high cost and unproven efficacy • What to do when no diagnosis? • What is significance of all eggs aneuploid?

24. Impressions • Move towards PB1 and 2 Handyside 2011 • Are there two groups of patients? • Older patients offer PB 1 and 2 biopsy • Young patients with recurrent implantation failure offer blastocyst biopsy and CGH • Blastocyst biopsy? • 2 cases of blastocyst biopsy, vitrification, thaw and transfer in drug-assisted cycle, no pregnancy • 2 cases analysis performed with ET on morning of day 6, 1 pregnancy

25. Ethical challenges • Is it ethical to offer this treatment in absence of robust evidence from prospective RCT? • Is it ethical to deny this treatment to a fully informed patient who has a poor prognosis? • Could we do any harm? • High non fertilisation rate • ? Discarding normal undiagnosed eggs • Possibility of self-correction • CGH does not diagnose triploidy • Comparisons against SNP arrays

26. In Conclusion • Efficacy of PGS remains hotly disputed • PB biopsy and CGH micro-array can predict chromosomal abnormality in 89% of cases • Initial clinical data looks promising, adds to our understanding • Await ESHRE’s prospective multicentre RCT for confirmation

27. Thanks to Tony Gordon at Bluegnome and Dagan Wells, ElpedaFragouli at Reprogenetics IVF Hammersmith Team