ENDOMETRIAL PREPARATION

IN FROZEN EMBRYO

TRANSFER CYCLES

Aboubakr Elnashar Benha university, Egypt

Aboubakr Elnashar

CONTENTS

INTRODUCTION

I. NATURAL CYCLE

II. ARTIFICIAL (HORMONE

REPLACEMENT) CYCLE

III. OVULATION INDUCTION

CONCLUSION

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INTRODUCTION

1st successful pregnancy following FET: 1983 Trounson and Mohr

(Trounson, 1983).

Cryopreservation of embryos has become an integral part of ART programs.

Increased dramatically

1.Ttrend towards transferring fewer embryos after a fresh IVF cycle

2. Improved laboratory techniques (Skovmand 1997; Diniz, 2002; Fineschi et al., 2005; Gordts et

al., 2005; Thompson, 2005; Le Lannou et al., 2006; JOINT SOGC-CFAS, 2008; Min et al., 2010).

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Success= keypoints in performing FET

1. Selecting proper embryos used for FET

•Cleavaged embryos with grade I or II, and present

≥6 blastomeres at D3 are the right kind.

•Embryos with 4 blastomere: continue in vitro

culture to reach the stage of 8 blastomere then

perform vitrification cryopreservation.

•Non-high quality embryos: continue to culture

them into blasocyst and select the valid blastocyst

for cryopreservation.

This step is important for settling the appropriate

time for ET

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2. Accurate synchronization of endometrium and

embyos is the key of performing FET.

Important for implantation

Age of the embryos after thawing corresponds to

age of the endometrium on the day of ET

3. Sufficient luteal support

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Methods:

Regular ovulatory Irregular or unovulatory

I. Natural II. Artificial=Hormone

replacement

III. Ovulation induction

True Modified E and P Gna, E and P GnT Letrozole Nolvadex

Functioning ovaries:

Any method

Quiescent Ovaries (e.g. donor oocyte recepient

with ovarian failure)

Only HRT with E and P

Many infertility units use a mixture of protocols for

FET.

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Best method: Little agreement in ovulatory women (Ghobara and Vandekerckhove, 2008;Weissman et al., 2009).

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I. NATURAL CYCLE NC-FET

Indication

It is only feasible for women with regular cycles and

proven ovulation.

In normally ovulating women: protocol of choice

The simplest method

Endocrine preparation of the endometrium is

achieved by endogenous sex steroid production

from a developing follicle.

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A. True Natural cycle Timing of ET is determined by detecting the

spontaneous LH surge

Method:

1. D10-12 (3-5 d prior to estimated ovulation day)

Serial US: E thickness, follicular development

and to time the commencement of testing for LH

LH (urine or blood) for detection of the LH surge ,

P levels When a rise in serum LH levels is observed, it is assumed that ovulation will occur

36–40 h later (Andersen et al., 1995).

LH surges in urine lag up to 21 h behind the appearance of the surge in blood (Hoff et al., 1983; Frydman et al., 1984; Miller and Soules 1996).

The day when LH exceeds 180% of baseline (calculated as the mean of the 3

previous morning samples) corresponds to a day prior to OPU/ovulation.

2. US for evidence of ovulation.

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3. FET

3–5 days after ovulation depending on the stage

of the embryo when frozen

The day of ovulation corresponds to the day of

egg retrieval.

If embryos were frozen at 72h, ovulation day+3 is

the right time to transfer. (Nawroth and Ludwig, 2005; Paulson,2011).

4. LPS:

progesterone

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Advantage

no medications are used: preferable to many

women.

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Disadvantages:

1. Even in women having regular menstrual cycles,

ovulation may not always occur

2. Problem associated with the detection of

spontaneous LH surges

A. variation in timing of its occurrence between

cycles and between patients (Park et al., 2007).

B. In order to assess the LH levels correctly,

determination should be performed at least daily,

and preferably twice a day.

C. LH urine kits have a large variation in thresholds,

which involve the risk of up to 30% of false-

negative testing, and are often reported by

patients as being difficult to interpret (Miller and Soules, 1996; Guermandi et al., 2001; O’Connor et al., 2006).

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B. Modified natural cycle To overcome the disadvantages of LH monitoring:

Administering hCG to initiate luteinization

Method:

1. Regular US:

2. HCG

5000 IU when dominant follicle (16 or 17or 18 mm):

ovulation 36–38 h later (Andersen et al., 1995). Combined with E2 levels of (450-550 pmol/L), 600-700 or 800-900 respectively

HCG administration correspond to the HCG

administration day of the source cycle

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Zheng et al, 2014 Aboubakr Elnashar

Advantages:

less US evaluation due to the hCG administration

compared with true NC-FET: less burden on

patients and doctors

(Weissman et al., 2011).

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Risks

1. unexpected ovulation

: accurate planning of embryo thawing and transfer

is not possible: cycle cancellation (7–12%)

(Fatemi et al., 2010; Hill et al., 2010).

2. Difficulty in ensuring timely thawing and ET.

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LPS in NC-FET.

Not required (Kyrou et al, 2010)

similar result in patients receiving true NC-FET (Lee et al., 2013).

{CL formation is not hampered by inadequate LH

secretion}

higher LBR in patients undergoing true NC-FET

with LPS. (Bjuresten et al. 2011)

Too little evidence supporting a positive effect of

LPS in patients undergoing NC-FET (Groenewoud et al, 2013, MA)

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True NC-FET Vs modified NC-FET:

no difference in outcome

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II. ARTIFICIAL (HORMONE

REPLACEMENT) CYCLE

In patients with irregular cycles HRT is best used.

Aims

To mimic the endocrine exposure of the

endometrium in the normal cycle.

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Key points:

1.An endometrial thickness 6 and showing triple line is

favorable for ET

2.Endometrial development is unaffected by the length

of the follicular phase. No adverse effects of reduction

the duration of exposure to E to 6 days or an increase

up to 35 days. However receptivity is best preserved

when the follicular phase is kept between 12 and 19

days

3.The endometrium is affected by either incremental or

fixed E levels , even in supraphysiologic range

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A. Estrogens and progesterones without

GnRHa. used in women with remaining ovarian function (Jaroudi 1991; Lelaidier 1992).

Method:

1. Oestrogen

Effect: proliferation of the endometrium, while suppressing the

development of the dominant follicle

Form and dose Oral E2 (progynova or Trisequence (Blue tab) or Cycloprogenova (White tab))

4-6 mg/d or 2 mg/d with an increasing doses

Transdermal patches (estrofem/estrace)

subcutaneous implants

vaginal rings or tablets. Aboubakr Elnashar

When:

D1 of cycle: prevents follicular recruitment by

suppressing FSH: spontaneous ovulation is

avoided.

D3

before D4.

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2. Progesterone

Aim:

initiate secretory changes

Form

oral tablets

IM 40-60 mg in oil

vaginal suppositories, gel or rings (Devroey 1998).

When

Once the lining endometrium: 7–9 mm on US

Not duration of E2 supplementation but the

endometrium thickness should be the leading factor

in determining the start of progesterone (Nawroth and Ludwig, 2005; El-Toukhy et al., 2008).

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Route:

No difference in PR between vaginal and IM

progesterone. (Glujovsky et al., 2010, SR)

based on patient and doctor preference.

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Timing of embryo thawing and ET:

Planned according to the moment of progesterone

supplementation.

The day P is started is considered ovulation/opu

day, and the time of thaw/transfer is determined

accordingly.

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1. E2

From D2 or 3 of cycle:

3 or 4 tab (acc to wt of pt) (for 7 or 8 days)

2. US:

When endometrial thickness 9 mm

3. Prontogest 400 mg twice daily or

duphaston 10 mg tds

4. ET

If you are freezing

Blastocyst: transfer on 6th or 7th day of progesterone

D3 or Morula: transfer on day 4 of progesterone

(some say D 5 for all, no difference)

4. LPS

Estrogen and progesterone

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Cases of amenorrhea (after induction by lutone

or lutofolon or cycloprognova or pills)

1. E2

1 Tab x 3 day from D3 of the cycle

2 Tab x 3 day

3 Tab x (2 – 5 day )

2. TVS monitoring

till endometrial lining is 9 – 10 mm.

3. Uterogestan1 tab of utrogestan/d

E2: 2 Tab/d till pregnancy test

4. ET

5. LPS:

if preg test positive: utrogestan 2X3

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1. Oral E2

2 mg/d from cycle day 1-4

4 mg/d from day 5-8

6 mg/d from day 9-12.

2. TVS: assess endometrial-thickness and ovulation

from D13 and E2 dosage was adjusted based on

the endometrial-thickness.

3. P: 40 mg IM when the endometrium reached a

thickness of 8 mm or maximum.

60 mg, 80 mg, and 80 mg progesterone were used

respectively in the following 3 days.

4. ET after 4 days of progesterone administration.

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1. Oral E2: 2 mg three times daily.

2. TVS: After 11, 12 or 13 days

A. If no leading follicle is present and the endometrial thickness is

≥ 8 mm: micronized progesterone (utrogestan) is added and

thawing and transferring is commenced 4 or 5 days later

according to the stage of cryopreservation

B. If the endometrial thickness is less than 8 mm: progynova

dose is raised to 2 mg 4 times daily for 7 days. After a week the endometrium is checked once again.

When the endometrium thickness is >8 mm and no dominant follicle (≥

14 mm) is present: utrogestan can be added and thawing and

transferring is performed according to local protocols.

If a follicle is visible during ultrasound: LH and P levels are determined.

• If these are raised, (serum LH ≥ 13 E/l or progesterone ≥ 15

nmol/l) luteinization of the follicle is considered to have taken place

and because of the associated diminished pregnancy rates,

thawing and transferring will not be performed.

• If serum levels are below the above mentioned levels thawing and

transferring can be performed according to local protocol

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Zheng et al, 2014 Aboubakr Elnashar

Advantages

1. Greater control and flexibility in the timing of

transfer. cycles are easier to plan: popular

among many patients and their doctors.

2. The length of the follicular phase can be varied

without detriment to IR or PR (Leeton 1991; Navot 1989)

3. Cycle cancellation rate is low.

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4. Higher IR and PR than patients with natural

cycles

IR and PR was higher in patients without ovulation

than ovulatory patients

ovulation in HRT cycle has a detrimental effect on

pregnancy.

HRT should be used in FET cycles, and ovulation of

patients should be evaluated during the treatment.

Zheng et al, 2014

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Risks:

1. The administration of E and P does not guarantee

complete pituitary suppression: a dominant follicle

may occur. luteinization may occur in 5% (El Toukhy et al., 2004).

2. Should the follicle undergo spontaneous

luteinization: endometrium may be exposed to

progesterone earlier: incorrect timing of thawing and

transferring.

From a cost-efficiency perspective, this aspect is of

great importance and should be taken into account

GnRHa co-treatment may be used to down-

regulate the pituitary and prevent follicular growth

(AC-FET with GnRH-FET). Aboubakr Elnashar

3. Both of these AC-FET approaches require

medication and are therefore less ‘physiological’

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LPS:

If a pregnancy occurs, E and P must be continued

until placental autonomy is established to replace

the absent corpus luteum.

No difference in PR between different methods of

LPS in patients undergoing ‘fresh’ IVF or ICSI (Chocrane SR, van der Linden et al., 2011).

The use of synthetic progesterone in patients

undergoing IVF or ICSI did lead to higher PR

compared with natural progesterone.

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B. Estrogens and progesterones with

GnRHa. Indication: In women with remaining ovarian function

Aim: GnRHa is used to suppress temporarily ovarian function and

render the patient functionally agonadal prior to inducing an

artificial cycle with E and P.

Method: 1. GnRHa (Decapeptyl CR, 3.75 mg)

in regularly menstruating it is given on D21 of the cycle

in oligomenorrhic from D1 of the cycle.

2. After 14 days:

E2 is determined

E2 is started as before

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Disadvantages:

1. More expensive

2. GnRHa can have side effects

3. Delay the resumption of spontaneous ovulation

if FET fails.

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AC-FET Vs AC-FET with GnRH

Cost is less

Cancellation due to luteinization occurs in 5% is

more

No significant difference in PR

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NC-FET Vs AC-FET.

No difference in PR.

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NC-FET Vs AC-FET with GnRHa:

No significant difference in PR or LBR. (Ghobara and Vandekerckhove,2008)

Significantly higher cancellation rates in

NC-FET (17.4%) Vs AC-FET with GnRH (4.5%)

{preventive effect of down-regulation with regard to

luteinization and ovulation}. (Hill et al., 2010).

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III. OVULATION INDUCTION

Ovary is functional but anovulatory and irregular cycles, ovulation may be induced by CC, Nolvadex, letrozole or hMG or or a combination (Van der Auwera;1994).

For synchronization the day of HCG administration

should correspond to the day HCG administration of

the source cycle in which the embryos were retrieved

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A. Letrozole, GNT In patients with irregular cycles Letrozole is 1st

choice for endometrium preparation for FET

1. Letrozole:

2.5-5mg on cycle D3 to 7, in order to promote monofollicular development

2. TVS: if follicles growth is not ideal: mild stimulation (HMG 150 IU)

3. HCG:

10000: when the follicles reach the criteria of mature

4. ET

Embryos were frozen at 72h: ETAfter 4 or 5 days

Blastocyst transfer is performed after 7 days.

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1. Letrozole

2.5mg,once daily from D3 to D7 of cycle

2. TVS

D10 of cycle: IM Gnt 37.5~75 IU/d until there is

LH surge or ovulation

3. HCG

On the day appearing LH surge: IM HCG10000

IU.

If there is sill no LH surge when the follicle is 20-

24mm: ovulation is induced by HCG.

4. ET

3 days after ovulation is observed.

4. LPS:

HCG 2000-2500 IU/3d.

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B. Letrozole or nolvadex, E2 1. Letrozole: 1x1 x 5 or

Nolvadex 1x3x5 from D3 of the period

2. E2 1x2x3 from D 8 - 9th day of period (follicle 12-14mm)

1x3x3

3. TVS: from D10-11 till endometrial lining 9-11 mm

4. HCG

4. ET after 3.5 Days for the 2 days frozen embryo or after 4.5 Days for the 3 days

frozen embryo

5. LPS: E2: 1x2x12 w of pregnancy.

Cyclogest 200mg 2 days before E.T

Then Cyclogest 400mg after E.T. till 12w

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C. CC, E2 1.CC

Start D 3 to 5 of menstruation or withdrawal for 5

days

2. E2

2mg/d until D of HCG administration to overcome

detrimental effect of CC on endometrium.

3. HCG

Repeated testing of P with US

P within follicular levels

adequate follicular size (20-24 mm) and

E thickness ≥8mm: HCG 10000

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D. Chronic low dose GnT Anovulatory patients resistant to CC:

1.FSH

37.5 -75 U /d for up to 14 days

2. TVS

if no dominant F is recruited , the dose is increased

at increments 37.5 U every week to a maximum

225 U/d

When single dominant F of 10-12 mm the dose is

maintained

3. Monitor by US and E2 and P

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CONCLUSION It is not possible, to recommend one endometrial

preparation method in FET over another

Future RCTs should not only address PR but also

consider convenience and cost efficiency.

No significant advantage of one specific approach

to prepare the endometrium for FET in terms of PR

or LBR. (Ghobara and Vandekerckhove, 2008; Groenewoud et al, 2013, MA)

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Choice for either NC or AC should be made based

upon other factors:

1. number of canceled cycles

2. number of hospital visits to plan FET

3. possible serious adverse events and side-effects

4. hospital and IVF laboratory logistics

5. patient or doctor preference.

No publications that address these factors

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Thank you Aboubakr Elnashar

For the patients with thin endometrium or multiple

ET failures, HRT

1. Oral E2

(ethinyloestradiol) is given 75ug/day for 14-28 days.

Once the lining is greater than 8

mm, Femonston (estace and dydrogesterone)

8mg/d can be started.

1. The day femonston is started is considered

ovulation/opu day, and the time of thaw/transfer is

determined accordingly.

2. If oral E2 is not sufficient, E2 can also be given

through vaginal routes. Femonston (estrace)

1mg/d by vaginal route has good effects for the

patients with thin endometrium.

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For the patients with menstrual cycle more than 40

days: dose of letrozole is 5mg qd for 5 days.

for the patients with cycle 35-40 days, the dose of

letrozole commonly use 5mg for 3 days.

for the patients with cycle less than 28 days, the

dose of letrozole is recommended 2.5mg for 3-5

days.

For purpose of synchronization day of HCG

administration correspond to the HCG administration

day of the source cycle in which the Embryos were

retrived

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For the patients can not build the lining with the

above methods

1. office hysterscopy before FET.

2. if the endometrium is present pale mucous and

insufficiency bloold flow, docotors will scissor

endometrium gently.

3. Intrauterine scar tissue canbe removed with hyste

roscopy, intrauterine device will be placed for a

few months to prevent further adhesion formation

when necessary.

(Yanping Kuang)

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