WORLD IVM EXPERIENCE

Milton K. H. Leong, M.D.

IVF Centre

Hong Kong Sanatorium & Hospital, China

LEARNING OBJECTIVES

At the conclusion of this presentation, participants should be able to:

1. Describe the indications IVM

2. Outline the various IVM approaches undertaken currently.

3. Evaluate the IVM outcomes with regard to the treatment success rates and the babies born as a result of IVM treatment.

DISCLOSURE

Milton K. H. Leong, MD

None

Development of IVM

• It was first demonstrated in 1935 that the immature oocytes have the ability to resume meiosis spontaneously when removed from the follicle.

–Pincus G, Enzmann EV. J. Exp. Med. 62, 665-675 (1935)

• Edwards showed that in-vitro matured human oocytes could be fertilized.

–Edwards RG, Bavister BD, Steptoe PC. Nature. 221(5181), 632-5 (1969).

• the immature human oocytes retrieved during gynecologic surgery in an oocyte donation program resulted in the first IVM pregnancy in 1991.

–Cha et al., Fertil Steril 55; 109-13 (1991).

• 1994-first IVM pregnancy with a patient’s own oocytes.

– Trounson A, Wood C, Kausche A. Fertil Steril 62; 353-62 (1994)

Development of the follicleStage Follicle size (mm)

Primordial 0.03 - 0.04Primary 0.05 - 0.06Secondary 0.07 - 0.11Preantral 0.12 - 0.20Early antral (*) 0.21 - 0.40Antral (* +) 0.41 - 16.00Preovulatory (+) 16.10 - 20.00

+ IVF * IVM

Gougeon, Hum Reprod 1986;1:81-7

Target patient group

• Women with high AFC;– PCOS– PCO with regular cycles

• The most significant factor which determines the success of IVM treatment is the AFC of the woman

(Tan, 2002. Am. J. Obstet. Gynecol. 186; 684-9)

Patient selection for IVM

Suikkari 2007; Best Practice & Res Clin Obstet Gynecol 21; 145-155

promising outcomes are also reported in “regular cycling” women

Suikkari 2007; Best Practice & Res Clin Obstet Gynecol 21; 145-155

Better prognosis if AF basale count > 7Better prognosis if AF basale count > 7

Common Indications for IVM

• failure after > 6 cycles of ovulation induction

• women having IVF with high AFC

• repeated poor embryo quality in previous IVF cycles for no obvious reason

• repeated poor responders to ovarian stimulation

however

• low implantation rates when compared to conventional stimulated cycles.

– asynchrony in the cytoplasmic and nuclear maturation of the oocyte

– asynchrony in the endometrium– culture conditions

Various approaches to improve implantation rates in IVM

• Gonadotropin priming – None– hCG– FSH / FSH+hCG

• Metformin• IVF / ICSI

• Culture conditions

Clinical Laboratory

HCG Priming

• Theoretically;

– Promote invitro maturation– Improve pregnancy rates

IVM following hCG priming

• Cycles of IVM 25• Age (yrs) 35.4 4.7• Oocytes retrieved 10.3 5.4• Maturation rate (%) 84• Fertilization rate (%) 87• Cleavage rate (%) 95• Embryos transferred 2.9 0.6• Clinical pregnancies - no (%) 10 (40)

Chian et al

New Engl J Med 1999; 341:1624-6

0

10

20

30

40

50

60

70

80

90

0 12 24 36 48

+ HCG

- HCG

hours of culture

% o

f m

etap

has

e II

*p < 0.05

Chian et alHum Reprod 2000; 165-170

Response to LH in granulosa cells from follicles < 8 mm from ovulatory women (with normal ovaries or PCO compared to anovulatory women with PCO)

Fold increase in steroid accumulation in response to LH

above control

Patients Estradiol Progesterone

Ovulatory (normal and ovPCO) 1.0 (5.0 - 3.9);

(n = 46)a

1.0 (0.3 - 2.5);

(n = 42)c

Anovulatory (anovPCO) 1.4 (0.7 - 25.4);

(n = 17)b

1.3 (0.3 - 7.0);

(n = 20)d

a vs b, P<0.0003c vs d, P<0.03

Willis et al., Journal of Clinical Endocrinology and Metabolism 1998; 83:3984-91

Duration between HCG administration and oocyte retrieval

• When the durations of 35 hours vs. 38 hours between hCG administration and the oocyte retrieval were compared, the 38 h group yielded significantly higher number of mature oocytes.

• In-vitro maturation rate after 24 h in the culture was significantly higher, and the clinical pregnancy rate in the 38 h group was higher compared to the 35 h group in the unstimulated cycles, 40.9% vs. 25%.

Son et al. Fertil Steril 88(Suppl. 1), S24-S25 (2007).

Clinical outcome in hCG-primed IVM cycles with (Group 1) and without (Group 2) MII-stage oocytes on the day of retrieval

Groups Group 1 (n=48) Group 2 (n=46) P

No. of oocytes collected (mean + SEM)

922 (19.2 + 8.4) 854 (18.6 + 9.9) NS

No. of MII-stage oocytes collected (%)

135 (14.6) 0

No. of oocytes cultured 787 854 NS

No. of oocytes matured in vitro (%) 500 (63.5) 535 (62.6) NS

Total no. of oocytes matured (%) 635 (68.8) 535 (62.6) NS

No. of oocytes fertilized (%) 456 (71.8) 419 (78.3) NS

No. of oocytes cleaved (%) 396 (86.8) 377 (90.0) NS

No. of oocytes transferred (mean) 178 (3.7) 173 (3.8) NS

No. of pregnancies (%) 23 (47.9) 13 (28.3) <0.05

Son WY et al. RBM Online. (2008), in press

Hormonal Priming

Regular cycling PCOS

• BeneficialBeneficial•Wynn 1998Wynn 1998

• No differenceNo difference•Trounson 1998Trounson 1998•Suikkari 2000Suikkari 2000•Mikkelsen 2005Mikkelsen 2005

• BeneficialBeneficial•Mikkelsen 2001Mikkelsen 2001

• No differenceNo difference•Lin 2003Lin 2003•Chian 2000Chian 2000

FSH Priming

• Results are conflicting

• Potential benefits:– Larger ovarian size– Easier retrieval– Higher E2 levels– More maturational competence

May May improve endometriumimprove endometrium

Overview of IVM treatment cycle

• Withdrawal bleed• U/S scan day 2-4 to identify if PCO and

measure AFC• Repeat u/s scan on day of hCG to measure

endometrial thickness• s/c hCG 10,000 IU when ET 6-8 mm, largest

follicle 10-12 mm and oocyte retrieval 38 hours later

Transvaginal U/S-guided oocyte retrieval

• vaginal vault cleansed with sterile water• i.v. sedation sedation with fentanyl and L.A. • 19 G single single-lumen needle • reduced aspiration pressure (7.5 kPa)• multiple punctures• 10 ml culture tubes with 2ml warm 0.9% saline

with 2 IU heparin

In-vitro maturation of oocytes

• GV oocytes cultured in IVM medium supplemented with 75mIU/ml FSH + LH for 24 - 48 hrs, checked every 12 hours all MII oocytes undergo ICSI

• ET day 2 or 3 following ICSI

• Patients receive estradiol-17ß (micronized) immediately following OR and progesteron following ICSI

Endometrial Priming

Endometrium is Endometrium is

exposed to lowerexposed to lower

E2 levelsE2 levels

Dyssynchrony between phase Dyssynchrony between phase of endometrium – matured of endometrium – matured

oocyteoocyte

Endometrial preparation is necessaryEndometrial preparation is necessary

Endometrial preparation

Endometrial thickness on day of oocyte retrieval

<6 mm 10 - 12 mg estradiol-17ß (micronized)

6 - 8 mm 8 - 10 mg estradiol-17ß (micronized)

>8 mm 6 mg estradiol-17ß (micronized)

Progesterone support (50 mg I/M or 200mg tid, pv) started following ICSI

Timing of Oocyte Retrieval

Early atreticEarly atreticfolliclesfollicles

Dominant follicleDominant follicle

Still competent toStill competent toEmbryonic developmentEmbryonic development

Can be used in IVMCan be used in IVM

But; TIMING ?But; TIMING ?

Timing of Oocyte Collection

• Russell et al. (1999)

When the leading follicle > 13 mm

• Less oocytesLess oocytes

• Less fertilizLess fertilizationation

• Fewer embryosFewer embryos

Timing of Oocyte Collection

• Cobo et al. (1999)

When the leading follicle < 10 mm

Higher blastocyst formationHigher blastocyst formation

Metformin in IVM

• 56 women, 70 cycles • Metformin, 500 mg bid for 12 weeks before the IVM

treatment • HMG for 5 days and hCG 10,000 IU, 36 h prior to OPU• number of immature oocytes, oocyte maturation,

fertilization and cleavage rates in were comparable to the control group

• significantly higher implantation and clinical pregnancy rates were obtained in the metformin-treated group (15.3% and 38.2% respectively) compared to the controls (6.2% and 16.7%)

Wei Z et al. Fertil Steril 2007 Nov 15

IVM outcomes

Authors (year) No. of cycles

Indication No. of ET cycles

at cleavage stage

GnPriming

MaturationRate (%)

FertilizationRate (%)

Implantation

Rate (%)

Pregnancy Rate/ET

(%)

Miscarriage Rate (%)

Chian et al (1999) 25 PCOS 25 HCG 84 87 32 40 20

Cha et al (2000) 94 PCOS 85 None 75.1 67.9 6.9 27.1 26.1

Chian et al (2000) 1113

PCOS 1113

NoneHCG

69.184.3

83.990.7

24.816.6

27.338.5

040

Mikkelsen and Lindenberg (2001)

1224

PCOS 921

NoneFSH

44.059.0

69.070.0

021.6

033.3

57.1

Child et al (2002) 107 PCO/PCOS

107 HCG 76.0 78.0 9.5 26.2 26.1

Lin et al (2003) 3533

PCOS 3533

FSH+HCGHCG

76.571.9

75.869.5

9.511.3

31.436.4

13.0

Chian (2004) 254 PCO/PCOS

NA HCG 78.8 69.2 11.1 24.0 NA

Soderstrom-Anttila et al (2005)

PCO: 13 7

PCOS: 18 10

PCO: 13 7

PCOS: 18 10

9 (IVF)5 (ICSI)17 (IVF) 9 (ICSI)

None 60.649.254.353.2

35.072.482.570.0

13.30

34.512.5

22.20

52.922.2

0-

33.350.0

Cha et al (2005) 203 PCOS 187 None NA NA 5.5 21.9 36.8

Torre et al (2007) 138 PCOS NA HCG 61.7 62 10.9 24.5* 42.3

Son et al (2007) 415106

PCO/PCOS

415106

(blastocyst)

HCGHCG

74.078.2

80.180.5

9.726.8

28.451.9

NA21.8

Outcome of IVM cycles from literature in women with PCO/PCOS.

Outcome of IVM cycles from the literature in women with normal ovaries and regular cycles.

Authors (year)

No. of cycles No. of ET cycles

at cleavage

stage

GnPriming

Mean no. of oocytes retrieved

Maturation

Rate (%)

FertilizationRate (%)

Implantation

Rate (%)

CPR/ET (%)

M/CRate (%)

Child et al. (2001)

56 (normal)53 (PCO)

68 (PCOS)

505267

HCGHCGHCG

5.1 ± 3.710 ± 5.111.3 ± 9

79.575.9*

67.771.6*

1.58.99.6

423.129.9

502550

Mikkelsen et al. (2001)

132 83 None 3.9 60.1 72.9 NA 18 NA

Soderstrom-Anttila et al. (2005)

92 (IVF)100 (ICSI)

58 (IVF)86 (ICSI)

None 6.3 ± 3.46.5 ± 3.6

66.954.5

35.967.1

22.615

3121

33.316.7

* PCO and PCOS groups pooled together.

IVM for other indications

IVM oocyte donation

• 12 oocyte donors (29.7 yrs; AFC 29.7) • oocyte retrieval days 9-18 of unstimulated cycle• mean of 12.8 GV oocytes retrieved• 8.67 mature oocytes and 5.9 fertilized oocytes• 3.9 embryos transferred• implantation rate 19.1%; 6/12 clinical pregnancy

– 4 delivered Holzer et al

Fertil Steril 2007; 88: 62-67

IVM +/- natural cycle IVF and PGD

• 35 yr old with RM failed 2 IUI and 2 IVF • IVM offered because of PCO; 1 M II and 14 GV oocytes;

ICSI performed• 8 embryos, 6 biopsied, 1 embryo from MII oocyte and 1

from GV oocyte chromosomally normal for 6 autosomes and X and Y chromosome

• 2 ET – one blastocyst from MII oocyte and one morula from GV oocyte

• ß-hCG 399 IU 14 days after ET and livebirth in May 2005

Ao et al

Fertil Steril 2006;85:1510-12

IVM as a Rescue

• Some cycles are cancelled due to– Risk of OHSS– Poor pesponse

Can IVM be a Can IVM be a rescuerescue ? ?

these oocytes can be matured in-vitrothese oocytes can be matured in-vitro

IVM as a rescue

Risk of OHSS

Immature oocyte retriavalImmature oocyte retriaval ++ IVMIVM

47 % CLINICAL PREGNANCY47 % CLINICAL PREGNANCY

No OHSSNo OHSSLim 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

IVM as a rescue

• In POOR RESPONSE = E2 < 1000 pg/ml

< 4 mature oocytes

Poor responders no HCG Poor responders no HCG

Immature oocyte retrieval + IVMImmature oocyte retrieval + IVM

37,5 % Pregnancy rate37,5 % Pregnancy rate

Liu Liu et al. et al. Fertil Steril 2003Fertil Steril 2003

IVM for Fertility Preservation

Fertility preservation for young women

• Best option; embryo cryopreservation, after ovarian stimulation followed by oocyte retrieval and fertilization of oocytes by sperm; IVF or ICSI

• Probably second best;

oocyte cryopreservation after ovarian stimulation followed by oocyte retrieval

Ovarian stimulation is not suitable for certain cancer patients; no sufficient time and/or ovarian stimulation contraindicated

Solution ?

Trial: Retrieval of immature oocytes from unstimulated ovaries, and maturation in-vitro followed by cryopreservation of oocytes by vitrification

Viability and pregnancy outcome of vitrified IVM oocytes

No. of patients 20

Mean age 30.8 + 0.9

No. of mature oocytes retrieved 6

No. of immature oocytes retrieved 290

Mean oocyte maturation rate 67.3 + 4.9

No. of oocytes vitrified and thawed 215

No. of oocytes survived (mean % + SEM; range) 148 (67.5 + 5.8; range 23.5 -100.0)

No. of oocytes fertilized (mean % + SEM) 96 (64.2 + 4.5)

No. of embryos transferred (median; range) 64 (4; range 1 - 6)

No. of implantations (mean % + SEM) 4 (9.6 + 5.4)

No. of pregnancies (%) 4 (20.0)

No. of clinical pregnancies (%) 4 (20.0)

No. of ongoing pregnancies (%) 0 (0)

No. of live births (%) 4 (20.0)

Mean birth weight (grams) 3486

Chian et al, 2008, Fertil Steril, in press

Fertility preservation strategies offered for women

at MRC with cancer

Chemotherapy cannot be delayed and/or hormonal

stimulation contraindicated

Immature oocyte retrieval

IVM

Male partner available

(ICSI)

No male partner available

Male partner available

No male partner

Embryo cryopreservation

Ooycte vitrification

Embryo cryopreservation

Ooycte vitrification

Chemotherapy can be delayed and hormonal

stimulation not contraindicated

Ovarian stimulation mature oocyte

retrieval

Ovarian wedge resection or

oophorectomy

Immature oocyte retrieval from ovarian tissue

Ovarian tissue cryopreservation

Obstetric and perinatal outcomes of the IVM

pregnancies

Outcome of IVM, IVF, ICSI and normal pregnancies

• obstetrical and perinatal outcome of 432 babies (55 IVM, 217 IVF, 160 ICSI) compared with 1,296 age-matched spontaneous pregnancies (controls) delivered at a single hospital (MUHC)

Buckett et al.Obstet Gynecol 2007; 110:885-91

Perinatal outcome

IVM IVF ICSI Controls p-value

Twin pregnancy rate 12.0% 16.0% 14.0% 1.3% p<0.001

Triplet pregnancy rate 4.0% 2.0% 3.0% 0 p<0.001

Mean birthweight (g) 2,812 2,826 2,801 3,289 p<0.001

Mean gestational age (wks) 37 37 36 39 p<0.001

Mean Apgar scores at 1 min 8 8 8 8 n/s

Mean Apgar scores at 5 min 9 9 9 9 n/s

Mean cord pH 7.29 7.30 7.30 7.29 n/s

Congenital abnormalities following IVM (n=55)

Major malformations 2

• ompalocele• small ventricuoloseptal defect

1

1

Minor malformations 3

• patent ductus arteriosus 1

• congenital hip dislocation 2

Relative risk for any congenital abnormality compared with controls

RR 95% CI

IVM 1.19 0.35 – 3.25

IVF 1.01 0.52 – 1.90

ICSI 1.41 0.72 – 2.68

Pregnancy outcomes per clinical pregnancy after IVM, IVF and ICSI

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

Live Birth (p<0.05) Miscarriage(p<0.005)

Ectopic Late PregnancyLoss

Miscarriage inPCOS (NS)

IVM IVF ICSI Buckett et alFertil Steril 2007

Pregnancy Outcome in IVM

• Mikkelsen et al. (2005) ----- 47 IVM babies– 2 twins– 1 NT Normal karyotype– 2 preterm deliveries– 1 stillbirth (42 weeks)– 1 chromozomal abnormality

Pregnancy Outcome in IVM

• Malformation:– Cha, Fertil. Steril. 2005 5,3% major

malformation rate

• Later neuromotor development:– Soderstrom-Anttila, Hum. Reprod. 2006

))) Minor developmental delay at first year

))) No Difference in the second year

Deliveries and ongoing pregnancies(facts and educated guesses)

Countries Deliveries and ongoing pregnancies

Scandinavia 150

Italy 77

France 40

Germany 20

Rest of Europe 33

Total Europe 320

Deliveries and ongoing pregnancies(facts and educated guesses)

Countries Deliveries and ongoing pregnancies

Middle East 21

Japan 100

Vietnam 26

China (incl. HK) 60

Korea (Cha Hosp.) 57

Korea (Maria Cl.) ≈ 400

Rest of Asia 15

Total Asia 679

Deliveries and ongoing pregnancies(facts and educated guesses)

Countries Deliveries and ongoing pregnancies

Canada 120

USA 5

Australia 5

Total 130

Deliveries and ongoing pregnancies(facts and educated guesses)

Countries Deliveries and ongoing pregnancies

Asia 679

Europe 320

North America 125

Australia 5

Grand Total 1129

- one year ago !

Korea 455

Taiwan 20

Colombia 7

Canada 131

Finland 52

Turkey 8

China 58

Japan 51

Vietnam 42

Hong Kong 18

Denmark 34

Italy 56

UK 8

Total 930

Conclusions

• IVM simplifies treatment, reduces costs and eliminates OHSS

• IVM successful in women with high AFC

• hCG increases final number of MII oocytes and rate of maturation

• IVM may be helpful in women with repeated poor embryo quality in previous IVF cycles for no obvious reason, or repeated poor responders to ovarian stimulation

Conclusions

• IVM produces CPR/C of 35%, and up to 48% in selected cases, in women up to 35 .

• obstetric and perinatal outcomes of IVM pregnancies comparable with IVF and ICSI

• IVM may be useful for oocyte donation or PGD

• IVM may offer a chance for fertility preservation to young women with cancer and undergoing cytotoxic treatment.

• IVM may not replace standard IVF but appears to play increasingly important role in ART

Acknolwedge

Dr. Ezgi Demirtas

Reproductive Centre

McGill University