In-vitro maturation: In-vitro maturation: patient selection and patient selection and resultsresults

Aygul DemirolAygul DemirolAssoc Prof, Medical DirectorAssoc Prof, Medical Director

GURGAN CLINIC IVF Center, Ankara-GURGAN CLINIC IVF Center, Ankara-TurkeyTurkey

In-vitro maturation In-vitro maturation (IVM)(IVM)

Immature oocyte retrieval and Immature oocyte retrieval and subsequent oocyte subsequent oocyte maturation in vitro without maturation in vitro without any ovarian stimulationany ovarian stimulation

IVM (in-vitro IVM (in-vitro maturation)maturation)

In 1991, Cha et al. reported a pregnancy from IVF with oocytes obtained from ovariectomy specimens and matured in culture (healthy triplet girls) (Cha KY, Fertil Steril 1991)

Trounson et al. reported the birth of a normal baby with IVM of immature oocytes from a polycystic ovary syndrome (Trounson Fertil Steril 1994)

IVM (in-vitro IVM (in-vitro maturation)maturation)

IVM (in-vitro IVM (in-vitro maturation)maturation) There have been more than 1000

births of babies with IVM procedures, including in patients with PCOS

(Chian RC)

BUT, IVM has not become mainstream in IVF, with ovulation induction cycles with oocyte retrieval of mature (MII) oocytes still the highly favored protocol.

Why IVM ?Why IVM ?

Reduced costReduced cost Avoiding OHSSAvoiding OHSS Simplification of treatment

compared with conventional IVF-ET

IVM IndicationsIVM Indications PCOSPCOS High respondersHigh responders IVF/IVMIVF/IVM Fertility preservationFertility preservation Donation cyclesDonation cycles Delayed respondersDelayed responders Male infertilityMale infertility Poor respondersPoor responders

2-12 mm follicles

In generalIn general

clinical pregnancy rate 30-35%clinical pregnancy rate 30-35% implantation rate 10-15%implantation rate 10-15%

(R.C. Chian RBM Online, 2004)(R.C. Chian RBM Online, 2004)

IVMIVM

low implantation rates when low implantation rates when compared to conventional compared to conventional stimulated cyclesstimulated cycles..

– asynchrony in the cytoplasmic and asynchrony in the cytoplasmic and nuclear maturation of the oocyte nuclear maturation of the oocyte

– asynchrony in the endometriumasynchrony in the endometrium– culture conditions culture conditions

IVM is based on the treatment of

Two main groups of patientsTwo main groups of patients– With PCO (With PCO (have irregular, mostly

anovulatory cycles and are at increased risk for OHSS because of their higher sensitivity to gonadotropins)

– With normal ovaries (With normal ovaries (may wish to avoid the side-effects of hormone injections)

How to maximize IVM results byoptimizing clinical management

Patient selection criteria-best Patient selection criteria-best candidatescandidates

– Under 35 years of ageUnder 35 years of age– PCO/PCOSPCO/PCOS

Patient management-IPatient management-I

Baseline TV-USG (day 2, 3)Baseline TV-USG (day 2, 3)

Second USG (between day 6 and day Second USG (between day 6 and day 9)9)

(for follicular and endometrial assessment)(for follicular and endometrial assessment)

HCG priming 36 hours prior to egg HCG priming 36 hours prior to egg collectioncollection

Patient management-IIPatient management-II

IVM ovum aspiration needle, IVM ovum aspiration needle, single or double lumen, 19 G, 35 single or double lumen, 19 G, 35 cmcm

Aspiration pressure 85-100 Aspiration pressure 85-100 mmHgmmHg

All visible follicles are aspiratedAll visible follicles are aspirated

Patient management-Patient management-IIIIII

Priming with FSH or HMG ?Priming with FSH or HMG ? Priming with HCG ?Priming with HCG ?

FSH PrimingFSH Priming

Results are conflictingResults are conflicting

Potential benefits:Potential benefits:– Larger ovarian sizeLarger ovarian size– Easier retrievalEasier retrieval– Higher E2 levelsHigher E2 levels– More maturational competenceMore maturational competence

May lead to improved May lead to improved endometrial primingendometrial priming

HCG PrimingHCG Priming

Theoretically HCG primingTheoretically HCG priming

– Promote invitro maturationPromote invitro maturation– Improve pregnancy ratesImprove pregnancy rates

However the exact mechanism of HCG However the exact mechanism of HCG on small follicles is still unclearon small follicles is still unclear

HCG PrimingHCG Priming

First prospective studyFirst prospective study

Chian et al. (2000)Chian et al. (2000)

• Increased oocyte maturationIncreased oocyte maturation

• High clinical pregnancy rateHigh clinical pregnancy rate( 36 %)( 36 %)

hCG PriminghCG Priming In-vitro maturation rate is faster in oocytes In-vitro maturation rate is faster in oocytes

obtained from hCG primed IVM oocytes. obtained from hCG primed IVM oocytes.

(Chian et al.,Human Reprod ,2000 ; Son et al., (Chian et al.,Human Reprod ,2000 ; Son et al., RBM Online,2006)RBM Online,2006)

- Hastens the the oocyte maturation in-vitro- Hastens the the oocyte maturation in-vitro

- Makes the oocyte retrieval easier- Makes the oocyte retrieval easier Mature oocyte on the day of retrieval is higher Mature oocyte on the day of retrieval is higher

(Son et al.,Hum.Reprod,2002)(Son et al.,Hum.Reprod,2002) Higher fertilization,cleavage and blastocyt Higher fertilization,cleavage and blastocyt

development rates in IVM cyclesdevelopment rates in IVM cycles Number of good quality blastocysts higher (40% Number of good quality blastocysts higher (40%

vs 23.3%) (Son et al.,RBM Online,2008)vs 23.3%) (Son et al.,RBM Online,2008)

Lab management-ILab management-I

Determination of cumulus-oocyte Determination of cumulus-oocyte complexes (COCs) (special sliding complexes (COCs) (special sliding technique-after using cell strainer)technique-after using cell strainer)

The immature COCs are incubated The immature COCs are incubated in culture dish containing 1 ml in culture dish containing 1 ml oocyte maturation medium oocyte maturation medium supplemented with a final supplemented with a final concentration of 75 mIU/ml FSH and concentration of 75 mIU/ml FSH and 75 mIU/ml LH 75 mIU/ml LH

for SAGE mediumfor SAGE medium

For MediCult medium, For MediCult medium, preincubation in LAG medium (2-3 preincubation in LAG medium (2-3 hours)hours)

Transferring into IVM final Transferring into IVM final maturation medium (9 ml IVM maturation medium (9 ml IVM medium is added 1 ml medium is added 1 ml patient patient serumserum, 10 , 10 µµl pregnyl, 100 l pregnyl, 100 µµl FSH)l FSH)

Lab management-IILab management-II

Stripping oocytes 24 hours after Stripping oocytes 24 hours after cultureculture

Twenty for hours after maturation Twenty for hours after maturation additional 24 hours for immature additional 24 hours for immature COCsCOCs

Mature oocytes are subjected to ICSIMature oocytes are subjected to ICSI Embryo maintenance medium for Embryo maintenance medium for

SAGESAGE ISM1 for MediCultISM1 for MediCult

Lab management-IIILab management-III

Endometrial primingEndometrial priming

17-17-ββ-oestradiol starts on the day -oestradiol starts on the day of OPU (2 mg orally, three times of OPU (2 mg orally, three times daily and continue until daily and continue until pregnancy test)pregnancy test)

Two days after OPU, intravaginal Two days after OPU, intravaginal progesterone suppositories 600-progesterone suppositories 600-800 mg, daily and continue until 800 mg, daily and continue until pregnancy test)pregnancy test)

Clinical outcome for Clinical outcome for PCO/PCOSPCO/PCOS

Jurema MW. Fertil Steril 2006;86:1277–91.

Clinical outcome for normal Clinical outcome for normal ovariesovaries

Jurema MW. Fertil Steril 2006;86:1277–91.

Pregnancies and deliveries after transfer of humanPregnancies and deliveries after transfer of humanblastocysts derived from in vitro matured oocytes inblastocysts derived from in vitro matured oocytes inIVM ( PCO(S) ) (Blastocyst vs cleavage ET)IVM ( PCO(S) ) (Blastocyst vs cleavage ET)

The abortion rate, gestational age and birth weight at delivery, and obstetric complications of pregnancies conceived by IVM-ET in women with PCOS were comparable with those of other women with PCOS being treated by conventional IVF-ET(Fertil Steril, 2005)

CONCLUSION: Compared with IVF and ICSI, IVM is not associated with any additional risk.(Obstet Gynecol 2007;110:885–91)

Outcome of the IVM Outcome of the IVM cycles (sept 2005-jan cycles (sept 2005-jan 2010)2010)

GURGAN CLINIC IVF CenterGURGAN CLINIC IVF Center

Ankara, TurkeyAnkara, Turkey

n % mean

Mean age 30.2Cycles 321Cycles with oocytes 321 100Oocytes retrieved 27258.724 h maturationOocytes reaching MII 1253 47.1 4.2Oocytes fertilized(2PN) 989 79.2 3.08Embryos cleaved 890 90.148 h maturationOocytes reaching MII 408 15.3 1.2Oocytes fertilized (2PN) 301 74Embryos cleaved 210 70.2

n % mean

transfer and outcome

Cycles with embryo transfer 288 90.4Embryos transferred 806 2.8Biochemical pregnancies/transfer 103 36.1Clinical pregnancies/transfer 78 27.1Implantation rate 13.4

Ongoing pregnancies 41Live birth 16Abortion rate 21 26.9

Immature oocyte after oocyte Immature oocyte after oocyte pick-uppick-up

Mature oocytes after Mature oocytes after denudation & ICSIdenudation & ICSI

Fertilised (2 Fertilised (2 PN)oocytesPN)oocytes

Cleavage stage (Day 2) Cleavage stage (Day 2) embryosembryos

Blastocyst stage Blastocyst stage (Day5)(Day5)

PREGNANCIES IN TURKEY PREGNANCIES IN TURKEY FOLLOWING IN VITRO OOCTYE FOLLOWING IN VITRO OOCTYE MATURATIONMATURATION

Aygul Demirol, Tamer Sari, Bagdagul Girgin, Aygul Demirol, Tamer Sari, Bagdagul Girgin, Erkin Kent, Suleyman Guven, Timur GurganErkin Kent, Suleyman Guven, Timur Gurgan2007, GORM2007, GORM**Two women with history of infertility and Two women with history of infertility and PCOS underwent in vitro maturation (IVM) PCOS underwent in vitro maturation (IVM) program without controlled ovarian program without controlled ovarian hyperstimulation. The patients were primed hyperstimulation. The patients were primed with 10.000 IU HCG 36 h before oocyte with 10.000 IU HCG 36 h before oocyte retrieval. Oocytes-cumulus masses were retrieval. Oocytes-cumulus masses were matured in IVM medium. The matured matured in IVM medium. The matured oocytes were fertilized by ICSI and embryo oocytes were fertilized by ICSI and embryo transfer was performed on day 3transfer was performed on day 3

accepted

IVM for a Second ChanceIVM for a Second Chance Recent reports:Recent reports:

Risk of OHSSRisk of OHSS

Immature oocyte retriavalImmature oocyte retriaval ++ IVM instead of cancellationIVM instead of cancellation

47 % CLINICAL PREGNANCY47 % CLINICAL PREGNANCY

No OHSSNo OHSS

Lim Lim et al. et al. Fertil Steril 2002Fertil Steril 2002

10,000 IU HCG10,000 IU HCG

Leading follicle = 12-14 mmLeading follicle = 12-14 mm

Natural Cycle + IVMNatural Cycle + IVM

But natural cycle yields only 1 But natural cycle yields only 1 single folliclesingle follicle

For other non dominant For other non dominant folliclesfollicles

IVM may be an optionIVM may be an option

Because IVM is possible Because IVM is possible even if the dominant even if the dominant follicle is selectedfollicle is selected

Thornton 1998 Fertil SterilThornton 1998 Fertil Steril

Thank you for their Thank you for their help and supporthelp and support

Ri-Cheng Chian, McGill Ri-Cheng Chian, McGill Reproductive Center, Montreal, Reproductive Center, Montreal, CanadaCanada

Mette Munk, Jyllinge, DenmarkMette Munk, Jyllinge, Denmark and others…and others…