1. Pre-implantation Genetic Testing
Pre-implantation Genetic Diagnosis
Pre-implantation Genetic Testing
Yacoub Khalaf MD, FRCOG
Director of Assisted Conception Unit & Centre for
Pre-implantation Genetic Diagnosis
Guy’s & St Thomas’ Hospital and King’s College, London, UK
Centre for Preimplantation Genetic Diagnosis 1

2. Pre-implantation Genetic Testing
To diagnose/exclude embryos that are affected by serious genetic disorders (PGD).
To screen IVF embryos to exclude aneuploid embryos from transfer with a view to improving chances of pregnancy (PGS).

3. PGD

4. Health legacy of genetic disorders
Recurrent pregnancy loss
Neonatal and childhood death
Structural abnormality
Mental disability
Chronic disease
Late onset disease
Centre for Preimplantation Genetic Diagnosis

5. Reproductive options for those with serious genetic risk
Reproductive roulette
Gamete donation
Adoption
Remain childless
Prenatal diagnosis and termination of pregnancy
Centre for Preimplantation Genetic Diagnosis

6. Centre for Preimplantation Genetic Diagnosis
There is another way!
Pre-implantation Genetic Diagnosis
Centre for Preimplantation Genetic Diagnosis

7. Pre-implantation Genetic Diagnosis
Detection of genetic information in an embryo at risk of a
specific genetic disease
by examining a representative sample taken at a preimplantation stage of development
What is PGT
Centre for Preimplantation Genetic Diagnosis 7

8. Centre for Preimplantation Genetic Diagnosis
The principle of PGD
affected
affected
affected
Transfer only unaffected embryos to the patient
Centre for Preimplantation Genetic Diagnosis

9. Preimplantation Genetic Diagnosis9

10. Centre for Preimplantation Genetic Diagnosis
Tissues for Preimplantation Biopsy
egg
cleavage stage
blastocyst
Polar Body
Blastomere
Trophectoderm
Centre for Preimplantation Genetic Diagnosis

11. Assisted Conception Unit Fertilisation in vitro
Genetics Centre
Fertilisation in vitro
(IVF or ICSI)
Accurate genetic diagnosis
Appropriate
Genetic Counselling
Embryo biopsy
Diagnosis by
FISH
PCR
Transfer
2 unaffected
embryos
Steps to PGD

12. Types of genetic disorders
Single Gene disorders (PCR)
Spinal Muscular Atrophy
Cystic Fibrosis
Huntingdon’s Disease
Sickle cell disease EB
Chromosome rearrangements (Arrays)
64 Reciprocal translocations 14 Robertsonian 6 Inversions
Sex Linked disorders e.g. OTC, Hunter’s, ALD etc
Centre for Preimplantation Genetic Diagnosis

13. Diseases detectable by PCR on single cells
Ornithine transcarbamylase deficiency
X-linked hydrocephalus
Crouzon syndrome
Stickler syndrome
Tuberous sclerosis
Central core disease
Gaucher's disease
Hyperinsulinaemic hypoglycaemia
Hunter's syndrome
Agammaglobulinaemia
Alzheimers disease
HLA typing
Cystic Fibrosis
Sickle cell disease
Haemophilia A and B
Thalassaemia
Fanconi Anaemia
Spinal and bulbar muscular atrophy
Retinitis Pigmentosa
Alport Syndrome
Phenylketonuria
Tay Sachs disease
Alpha 1 antitrypsin deficiency
Myotonic Dystrophy
Huntington's disease
Adenine deaminase deficiency
Fragile X syndrome
Marfan disease
Lesch Nyhan syndrome
P53 mutations
Epidermolysis bullosa
Plakophilin deficiency
Kennedy disease
Long chain acyl CoA dehydrogenase deficiency
Charcot Marie tooth disease type 1a, 2a
Spinocerebellar ataxia type 7
Osteogenesis imperfecta type I and IV
Duchenne muscular dystrophy
Congenital adrenal hyperlasia

14. PGH/HLA

15. Outline Experience at GSTFT PGD Centre
Compare data with PGD Consortium Data
Factors affecting likelihood of conception
Recent developments - Elective single blastocyst transfer - Freezing of PGD Blastocysts
Centre for Preimplantation Genetic Diagnosis

16. Outline Experience at GSTFT PGD Centre
Compare data with PGD Consortium Data
Factors affecting likelihood of conception
New developments - Elective single blastocyst transfer - Freezing of PGD Blastocysts
Centre for Preimplantation Genetic Diagnosis

17. Number of PGD cycles started by year

18. UK PGD cycles HFEA 3 year aggregate data
ACU, Guy’s Hospital
UCH, London
CARE, Nottingham
The Bridge Centre, London
Glasgow Royal Infirmary
IVF Hammersmith, London
Oxford Fertility Unit
Edinburgh ACU
ARGC, London 18 18

19. Main conditions in
2009
2010
2011
2012
2013 CF 35 28 39 29 HD 17 31 32 26
DMD 11 8 5 9 16
Fragile X 10
Hbopathy 6 4 MD 7 12 3

20. Type of chromosomal rearrangement

21. Cycle Outcome (%)

22. % transferable = 42% (2.6 embryos)
48%
51%
27%

23. 1741 sequential cycles of PGD
GSTT Centre for Pre-implantation Genetic Diagnosis
(Sept Dec 2013)
Stimulated
Cycles to OR
Cycles to biopsy
Cycles to ET 726(79%) 92(75%)437(63%) (72%)
Clinical Preg
Per egg collection 36% 31% 26% %
Per Transfer 42% 36 % 39% %
Rearrange
X-linked
Total
Single gene
Centre for Preimplantation Genetic Diagnosis

24. Outline Experience at GSTFT PGD Centre
Compare data with PGD Consortium Data
Factors affecting likelihood of conception
New developments - Elective single blastocyst transfer - Freezing of PGD Blastocysts
Centre for Pre-implantation Genetic Diagnosis

25. PGD outcome at GSTT (1997–2011) cf ESHRE PGD consortium (1997-2007)
SGD
XL FISH
Chromosome
Total
Cycles to OR
561 (4733)
95 (1167)
495 (4253)
1151 (10153)
Cycles to ET
480 (3727)
80 (880)
341 (2731)
901 (7338)
Mean no. replaced
1.4 (1.9)
1.4 (1.8)
1.4 (1.7)
CPR/OR
36% (22%)
33% (19%)
27% (17%)
32% (19%)
CPR/ET
42% (29%)
39% (26%)
40% (27%)
Centre for Preimplantation Genetic Diagnosis

26. Outline Experience at GSTFT PGD Centre
Compare data with PGD Consortium Data
Factors affecting likelihood of conception
New developments - Elective single blastocyst transfer - Freezing of PGD Blastocysts
Centre for Preimplantation Genetic Diagnosis

27. Factors affecting likelihood of conception in couples having PGD
Condition being tested
Female age
No. of fertilised oocytes
No. of embryos suitable for transfer
Centre for Preimplantation Genetic Diagnosis

28. Clinical pregnancy rate and type of genetic condition
Centre for Preimplantation Genetic Diagnosis

29. Effect of age on likelihood of pregnancy
≤38 years 3x more likely to get pregnant per cycle

30. No. of fertilised eggs and chance of transfer & pregnancy
OR 2.4
P<0.05 %
OR 2.9
P<0.003
Centre for Preimplantation Genetic Diagnosis

31. Clinical pregnancy rate and embryo availability
Centre for Preimplantation Genetic Diagnosis

32. Clinical pregnancy rate and freezing
Centre for Preimplantation Genetic Diagnosis

33. Centre for Preimplantation Genetic Diagnosis
Adverse factors
Age >38 years
<8 fertilised oocytes
<3 transferable embryos No freezing
Centre for Preimplantation Genetic Diagnosis

34. Outline Experience at GSTFT PGD Centre
Compare data with PGD Consortium Data
Factors affecting likelihood of conception
Recent developments - Elective single blastocyst transfer - Trophectoderm biopsy and freezing of PGD Blastocysts
Centre for Preimplantation Genetic Diagnosis

35. PGS

36. Does screening for aneuploidy make a difference to ART outcome?

38. PGS Outcome – implantation
Gianaroli 1997, Gianaroli 1999, Munne 1999, Pellicer 1999, Kahraman 2000, Obasaju 2001, Munne 2003,
Rubio 2003, Pehlivan 2003, Oter 2004, Montag 2004, Wilding 2004, Platteau 2005, Rubio 2005
Non-randomised observational studies 38 38

40. Outcomes in Women Who Underwent Preimplantation Genetic Screening and in Controls
Table 2. Outcomes in Women Who Underwent Preimplantation Genetic Screening and in Controls.

41. It’s not all about the technology –
It’s the biology!
We have to take a step back and realise that it isnt all about the technology – we need to look at the biology

42. 10 whole chromosome mosaicism Structural mosaicism in 12 cells
Chromosome complements of the blastomeres analysed by array-CGH
( in 9 young patients whose previous transfer resulted in live birth)
Microarray analysis reveals abnormal chromosome complements in over 70% of 14 normally developing human embryos.
Mertzanidou A, Wilton L, Cheng J, Spits C, Vanneste E, Moreau Y, Vermeesch J, Sermon K. Hum. Reprod. 2012
4 uniformly diploid;
10 whole chromosome mosaicism
Structural mosaicism in 12 cells

44. ICM T1 T3 T2

45. ICM / trophoblast results may conflict
Is this a sufficiently robust test to discard patients’ embryos

46. For an embryo selection technique to be adopted ethically
Have clear established indications
Repeatable and reliable in any reasonably competent professional hands
A low false negative or false positive rate
Must make a significant clinical difference shown in properly constructed clinical trials
Cost effective from the patient / health provider perspective
PGS using FISH has failed all of these tests.

47. Conclusions
PGD is an important option that should be offered to couples at risk of having a child with serious genetic disease.
PGS is appealing in theory but has so far failed to show improvement in the chances of pregnancy per started cycle and should not be offered for IVF patients.