ked - society of clinical embryologists 1 st symposium, istanbul alan thornhill the bridge centre,...
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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Wells and Fragouli, unpublished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.
23
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