PGD – The good, bad and ugly

Dr Aniruddha Malpani, MDwww.drmalpani.com

Questions• What is the difference between PGD

and PGS ?• Should we be doing PGD for

everyone ?• Does PGD help ? Or harm ?

Will this embryo become a baby ?

Pre-implantation Genetic Diagnosis

• Handyside, Winston, and Hughes • 1990 for sexing embryos for preventing X linked

disorder– Used PCR for Y chromosome and transferred XX embryos

• Definition: Biopsy of a single cell per embryo, followed by its genetic diagnosis through different techniques (FISH, PCR, aCGH, NGS ), and the subsequent transfer of those embryos classified as genetically normal

PGD vs PGS • Preimplantation Genetic Diagnosis (PGD) • Refers to diagnosis of a definitive genetic condition in

an embryo • AIM: To provide disease free offspring

• Preimplantation Genetic Screening (PGS)• Refers to screening of embryos for known genetic

defects (mainly chromosome aneuploidies) • Aim: To improve the IVF outcome

IndicationsPGD

• Single gene disorders – Thalassemia, DMD,

• Carriers of chromosome abnormalities – Translocation carriers

PGS• At risk for aneuploidy – maternal age > 35 yrs– Prior trisomic

conception• Recurrent pregnancy losses• Failed IVF cycles – >3 prior embryo transfers with

high quality, morphologically normal embryos

PGS – old vs PGS – new

Polar body: Not recommended by ESHRE

Day 3 . Blastomere

Disadvantages- Chance of damaging the embryo- Only single cell sampled- MOSAICISM- Embryo is alive – autocorrection of genetic abnormalities

Advantages- Permits collection of many cells

Day 5 . Blastocyst

Techniques to choose for PGD/PGS Depends on the disorder !

• Chromosomal • FISH • Array CGH• SNP array• NGS

• Molecular (those that require PCR)• PCR- Sequencing • PCR variants• NGS

Problem : Mosaicism• Not all the cells of a given embryo are

genotypically identical

46XY

46XY

47XYY

46XY

47XY

+13

45XO

47XY

+1846XY

46XY

46XY

47XY

+1847X

Y +18

46XY

47XY

+18

Simple mosaicism Chaotic

10-25 % of embryos are mosaics

Cause of misdiagnosis

Solution: Trophectoderm biopsy (Blastocyst stage)

• Advantages– More accurate. Permits collection of

more than 2 cells– Reduces risk of misdiagnosis because of

mosaicism– Reduces number of embryos to be screened

• Disadvantages– Need to freeze embryos and then transfer them in

the next cycle after thawing– May not get enough blastocysts to biopsy,

especially in the older woman

PGS cannot diagnose all genetic problems – only what you look

for !

Words of caution• Counseling is a very crucial component of PGD/PGS• Patients understand the

• risk for having an affected child • the impact of the disease• the multiple technical limitations including the possibility of an erroneous

result• Discuss alternative options, including the use of donor

eggs and adoption• Prenatal Diagnosis of patients undergoing PGD (not

required for PGS)• Field of genetics advances rapidly. Need to work closely

with the embryologist and the geneticist

The embryo is alive !

• Embryo has the amazing ability to self-correct and repair itself• Abnormal cells die by apoptosis• They are eliminated and replaced by

healthy cells, repairing - and in many cases completely fixing - the embryo.

Mouse model of chromosome mosaicism reveals lineage-specific depletion of aneuploid cells and normal developmental potential

• Helen Bolton, et al• Nature Communications . Published 29

March 2016

Limitations • Miscarriages can occur after transfer of “

normal “ PGS tested embryos . • Time-consuming and expensive. • Lots of coordination and expertise. Best

offered in a few referral centers• Large number of embryos may be abnormal,

thus leaving few or no healthy embryos for transfer. Older patients produce few embryos. PGS can actually reduce pregnancy rates

The American Society for Reproductive Medicine (2007) position on PGS

• Available evidence does NOT support the use of PGS as currently performed :

• to improve live-birth rates in patients with advanced maternal age ( AMA)

• to improve live-birth rates in patients with previous implantation failures ( RIF).

• to improve live-birth rates in patients with recurrent pregnancy loss ( RPL) .

• to reduce miscarriage rates in patients with recurrent pregnancy loss related to aneuploidy

PGS for aneuploidy(Schattman, UptoDate review, 2014) • “PGS using day 3 blastomere biopsy and FISH decreases the

chances of live birth. • In couples with recurrent pregnancy loss, PGS is unlikely to benefit

those with proven karyotypically normal miscarriages. • In patients with karyotypically abnormal miscarriages, PGS may

decrease the risk of subsequent miscarriage, although there is no evidence that it will increase the probability of having a child.

• In young, good prognosis couples, PGS using blastocyst biopsy and 23 chromosome analysis may improve the chance of conception with single embryo transfer; however, the chance of live birth after including frozen embryo transfers is not increased”.

An adverse past and an uncertain future

• In 2007, Mastenbroek et al. published a RCT (n=206 patients in PGS and n=202 in the control group for older women.

• PGS (day 3 biopsy + fluorescence in situ-hybridization-FISH) reduced the rates of ongoing pregnancies and live births (PGS = 24%; Control = 35%). This was the first time in which the use of PGS was criticized.

• Advocates of PGS blamed negative outcomes on:• imperfect FISH technique• biopsy carried out on the third day (embryo cleavage stage) would carry a high

frequency of mosaicism.

• Solution = PGS-new . Biopsy on Day 5/6 !• Use new genetic techniques (Comparative genomic hybridization-CGH; Array-

CGH-aCGH; Real-time quantitative polymerase chain-reaction-qPCR; Next generation sequencing-NGS)

New wine in old bottles ?• Poor quality biased RCTs• Selects good prognosis patients only ! • Restricted to special populations (a

mean of 16 eggs collected and 6 blastocysts produced) Rare in the universe of older patients who require ART.

Indication creep• Good example – ICSI

Do it now for all IVF patients. However a bad IVF lab can kills lots more eggs when doing ICSI as compared to doing IVF !

• When you invest all that time, money and expertise in learning how to do PGD, what do you do with that expertise if there aren’t enough patients? The answer is simple - create more patients !

Routine PGS for all IVF patients• Why not do PGS for all older women ? Makes logical sense • Next step - Why not just offer it to everyone who is doing IVF

? • Which patient is going to want genetically abnormal embryos

transferred? • If you are spending so much for IVF, why not spend a little

more ?• However, in real life , we forget that all these additional

technical procedures add to risk; cost, and time

Why PGS has become “ hot”• World leading clinics present their data at conferences

Other IVF clinics want to start copying them• Patients demand this “new “ technique which reporters write about• IVF doctors like doing new stuff ; especially when this increases our

income• PGD was supposed to revolutionise IVF treatment and improve

pregnancy rates. Ended up reducing pregnancy rates for patients who never needed it in the first place.

• Steep learning curve with technically complex procedures • Patients serve as uninformed guinea pigs in these clinics.

Medical reversal = more is not always better

• Too much technology can cause unintended harm• IVF doctors don’t seem to learn. Repeat the same mistakes every

decade. • “New “ set of technologies come with the promise that they are

better! • IVF treatment has a limited success rate. Human reproduction has its

own biological limitations. • Papers published in medical journals quote high success rates. This

can be very misleading . Most IVF clinics can never match those rates because they simply don't have the experience or expertise of these researchers.