KED - Society of Clinical Embryologists1st Symposium, Istanbul

Alan Thornhill

The Bridge Centre, London , UK

December 5th 2012

Assessment of Gamete/Embryo Viability by the Utilization of new PGS (Preimplantation

Genetic Screening) Techniques or

PGS: an Embryologist’s Perspective

OR

I am doing PGS anyway Can you give me any tips?

0102030405060708090

100

30-34 35-38 39-42 43-50

Wells and Fragouli, unpublished

Evidence: Clinical Value of ACGH?

Bridge Success Rates LIVE BIRTH RATE post PGD ANEUPLOIDY screening

Using ARRAY CGH  in 40 – 45 yr Maternal Age Group

 Year PGD-A array CGH

 2010  22%

 2011  22%

*Equivalent UK national rates for this age group are 10-13%

New Technologies – The Rise of array CGH

• First randomised trial* of blastocyst PGS with aCGH

• Women under 35 years (good prognosis)

• eSET

• Day-5 (blastocyst) biopsy

• All cycles had embryo transfer (day 6)

• No PGS: embryo transfer on morphology (n=48)

Pregnancy rate**: 41.7%

• PGS (aCGH): chromosomally normal embryos transferred (n=55)

Pregnancy rate**: 69.1%

*Yang et al (2012) **On-going pregnancy rate ≥20 weeks/cycle started

Evidence At Last?

• Retain all embryos• No harm to embryo• No upfront cost to patient• Transfer all embryos sequentially

Why Not?• Risk of abnormality NOT accepted• Miscarriage NOT Trivial• Repeated failure NOT Accepted• Multiple cycles (inc. FET) costs Time and Money

Why PGS? Do Nothing Option

• 1st polar body - Single cell (small)• No role in further development• Fragmented

Biopsy sample

• 2nd polar body - Single cell (small)• Haploid, Incomplete extrusion• No role in further development

• Nucleated blastomere – single cell (large)• Representative of embryo? mosaicism• critical role in further development

• Trophectoderm – 6-10 cells• Representative of FUTURE BABY?• Mosaicism

• Laser or mechanical• Timing• ICSI required

Biopsy Procedure

• Laser or mechanical• Timing (Sequential or simultaneous)

• Chemical, Laser or mechanical• Safety? Reduced implantation potential

• Laser or mechanical• Need to see AND AVOID ICM• Laser effects on sample?• Need for vitrification post-biopsy

• Entire MII cohort analyzed• High Biopsy & diagnostic workload/costs• Proportion will not fertilize

Biopsy – Practical Issues

• Entire 2pn cohort analyzed• High Biopsy & diagnostic workload/costs• Proportion will not develop well

• Good quality ‘day 3’ embryos only• Chance of no biopsy• Medium Biopsy & diagnostic workload/costs

• Good quality blastocysts d5/6 only• Moderate chance of no biopsy• Lower Biopsy & diagnostic workload/costs

• High result rate (>90%)• No paternal/meiosis II/post-zygotic information

Biopsy – Diagnostic Issues

• High result rate (>90%)• No paternal/post-zygotic information

• High result rate (>90%)• Chromosomal mosaicism common

• Extremely high result rate (>>95%)• SIGNIFICANCE OF MOSAICISM?

What is the aim of PGS?….to transfer embryos with

the correct amount of genetic material to increase the chance of having a healthy

baby &reduce the risk

of having a pregnancy with a chromosomal abnormality

..identify patients with high risk of aneuploid gametes/embryos to inform future treatment

And Neither Is PGS…..

So What Happens In The Real World?

Life is never that simple…..

Case studies: PB1 only

Extensive PRE- AND POST-TEST counsellingSuggested egg donation Last chance/diagnostic closureFOLLOW-UP OF EMBRYOS CONFIRMED PB1 RESULTS

• 49 year old • 2ND attempt IVF/ICSI• FAVOURABLE AMH/AFC• Results: 11 OOCYTES ALL WITH MULTIPLE

ANEUPLOIDIES

New IVF test – Array CGH – produces baby

Oliver, offering hope to infertile

13 previous failed IVF cycles

7/9 first polar bodies aneuploid

September 2nd 2009

Summary of Net Gains and Losses inThe Aneuploid Zygotes

Source of error Gains Losses Total (%)

Meiosis I 44 33 77 (34)

Meiosis II 55 47 102 (45)

Paternal or chromosome loss

18 30 48 (21)

ESHRE data 2010

Case studies: PB1+2

*Reciprocal gain and loss (chr 9 + 13) Reported ‘normal’ (no plots/correction?)Counsel for risk of patau’s syndromeNeed for data on outcomes of G/LNeed for truth data

• 41 year old (AMA)• 2nd attempt (1st PB1 – LB)• 11 x oocytes/7 x 2pns• Results: 6 x abnormal, 1 x normal* 

RECIPROCAL GAIN/LOSS – PB1+2

Case studies: PB1+2

Reported as abnormalMosaicism in TE/ICM?Counsel for risk of cri-du-chat Need for data on mosaicism

• 40 year old (AMA)• 1st attempt (top – trisomy 21)• 6 x oocytes/5 x 2pns• Results:• 1x Normal, 2 x Abnormal, 2 x Normal (but

no result for PB2) – D5 rebiopsy• Partial deletion in chr 5  

Case studies: Trophectoderm

Reliability of aCGH?*5AB – discard/re-biopsy/et?Question diagnostic labRepeat test? Follow-up

• 38 year old • RIF (4 x IVF) • Frozen d3 (5) + D5 (1) thawed for TE BIOPSY• Results: • DELAYED DEVELOPMENT (no Fresh ET)• 2 x Euploid, 2 x Aneuploid• 1 x multiple aneuploid* (every chromosome!)

Single blastomere analysis:Complex pattern of gains and losses

Algorithms and reporting

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Automated calling

Estimation of confidence in aneuploidy call

Karyomapping and 24sure from a single cell SurePlex amplification

• Cells disaggregated from the remainder of embryo. Data supplied with courtesy of Professor Alan Handyside, London Bridge Fertility Centre.

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FISH probes 13, 16, 18, 21, 22 • Trisomy 17 • Monosomy 18, 21, 22

FISH probes X, Y, 21 • Trisomy X • Monosomy 21

• Advanced maternal age (>35 yr)• Most embryos aneuploid• No embryo transfer in most cycles• Prognostic for chance of pregnancy with the patient’s own

eggs• Optimal age range for embryos selection 37 to 43 years of

age

• eSBT (<35 yr) good prognosis patients at high risk of twins• Moderate incidence of aneuploidy• All cycles with ≥1 aneuploid blastocyst• All cycles with ≥1 euploid blastocyst for transfer

• Severe male factor infertility• Idiopathic repeat IVF failure• Miscarriage risk/previous abnormal/TOP

(NEW) Indications - 24 chromosome test

FACTS•Aneuploidy INCIDENCE c.50% affects all ages•Aneuploidy main cause of failed implantation•Aneuploidy testing CANNOT change the cumulative pregnancy rate

POTENTIAL BENEFITS•Prevent transfer or storage of aneuploid/non-viable eggs/embryos•Identify AMA women with good/poor prognosis for a livebirth using their own eggs•Increase pregnancy/live birth rate/ET (eSBT)•Reduce time to live birth or resolution

Summary i – What patients/providers need to know

• Genetic Counsellor provides Face to face, telephone,

email support for clinicians/ embryologists/patients

throughout process

• Single point of contact for patients

• Excellent patient information & consent forms

• Secondary reports (combine embryology and diagnostic

information)

• Time-lapse incubation?

• Treat every egg/embryo as important

• Question diagnostic team

Summary ii – how to provide good PGS

The Bridge Centre•Alan Handyside•Michael Summers•Karen Sage•Shaun Rogers

Bluegnome•Tony Gordon

Reprogenetics uk•Dagan Wells

Acknowledgements