This document discusses mild ovarian stimulation protocols for ovulation induction and in vitro fertilization (IVF). It outlines important factors to consider like ovarian reserve, previous response, and hormone profiles. It compares protocols using clomiphene citrate, aromatase inhibitors, and gonadotropins alone or in combination to induce ovulation of 1-3 follicles. Premature luteinization during ovarian stimulation is also discussed. The document aims to develop cost-effective low-dose IVF procedures suitable for developing countries like India.

1. Mild Ovulation Induction

2. What are important factors to be
considered important
• Ovarian reserve
• Previous ovarian response
• Basic hormone profile
• Role of LH
• Trigger
• Luteal phase support
• Pregnancy rate/cycle

3. After effects of the stimulation
• Oocyte quality
• Endo quality
• Embryo quality comparision between
conventional Vs minimal stimulation
• Difficulties with this approach
• Can we make it truly costeffective …to make
low cost IVF procedure in our developing
country..INDIA

4. Ovarian Reserve Assessment

5. Antral follicle count (AFC)
• Number of antral follicles (early follicular phase)
• Low numbers of antral follicles - sign of DOR and observable
earlier than a rise in FSH serum level
• But - cycle–cycle variation
• Intra-observer variation
• AFC ~ AMH
• Scheffer et al Fertil Steril 1999
• Broekmans et al Fertil Steril March 2008 Epub

8. Aims…
1. Induction of a single dominant
follicle…natural
2. Induction of small number of follicles
(1-3)…mild stimulation…IUI
3. Multiple follicular development (IVF&ICSI)

9. Role of FSH and LH in follicular development and
maturation
“Two–cell, two-gonadotrophin Concept”
• FSH works on granulosa cells-stimulating their
proliferation and growth
• FSH is active in early follicular phase thus stimulates
small follicles
• LH-acts on theca cells.. to produce androgens –
• Importance lies in the mid-to late follicular phase
stimulates large–follicle growth
• Causing atresia of small follicles- prevent OHSS
Marco Filicori

10. Important concepts in Folliculogenesis.
• FSH threshold.
The lowest dose that induces
growth of the single most
sensitive follicle
• FSH window.
– The time frame for which
this level of FSH needs to be
maintained for follic.
recruitment and growth
(mono/multifollicular
recruitment )
• LH window/ ceiling.
– Concentration of LH essential
for follicular growth &
selection of DF

12. 0
500
1000
1500
2000
2500
3000
3500
0 5 10 15 20
Endometrium (mm)
E2
(pmol/L)
Endometrial Thickness vs. Estradiol
Barash et al, Fertil Steril 1998

13. • Oocytes retrived in high LH concentrations during
the later stages of follicular development and in
the peri-ovulatory phase were of poor quality
and were associated with a reduction in the rates
of fertilization and cleavage .
• Embryos obtained from such eggs were of
inferior quality and displayed excessive
fragmentation and asymmetric alignment of
blastomeres
Mishra S Lin PC Lei ZM Carrillo AJ Rao CV 2000
Gynecologic Investigation 7 (suppl 1) abstract 576.

14. How to induce a single dominant
follicle?
Ovulation induction protocol which mimic
more closely the FSH threshold and window of
the natural cycle.

16. How to Obtain Small Number of
Follicles (1-3)

17. 1. CC.
2. Aromatase Inhibitor..step up,extended
3. CC ± FSH or ± HMG.
4. Gn. Standard step-up protocol.
5. Gn. Low dose step-up protocol.
6. Gn. Low dose step-up, step-down
protocol.
Protocols….

18. Unripe
follicle
Ripening
follicle
Ovulation Corpus
luteum
Regression of
Corpus luteum
Clomiphene
100 mg day2
for 5 days
Gonadotrophin
stimulation from
day 4 to day of
HCG
HCG
Leading follicle > 18mm
Oocyte mature
38 hrs

19. • A novel protocol of ovulation induction with delayed
gonadotropin-releasing hormone antagonist administration
combined with high-dose recombinant follicle-stimulating
hormone and clomiphene citrate for poor responders and women
over 35 years
• 85 patients received clomiphene citrate, high-dose recombinant
human FSH, and a delayed, multidose GnRH antagonist, whereas
60 patients underwent a standard long protocol.
• Conclusion
The proposed protocol of ovulation induction can be usefully
administered in poor responders as well as in aged woman,
probably because the delayed administration of GnRH antagonist
prevents its adverse effects on ovarian paracrine activity and on
oocyte maturation.
• Fertility and Sterility
Volume 81, Issue 6 , Pages 1572-1577, June 2004

20. Difference … CC & Aromatase inhibitor

21. Letrozole Step up Protocol
• A novel protocol for multifollicular
development
• Dose : 1,2,3,4 tablets of letrozole (2.5mg) daily
on menstrual cycle days 2,3,4 and 5
respectively

23. Highlights :
• Extends FSH window by prolonging the suppression of
estrogen levels
• Prevents rising estrogen from suppressing endogenous FSH
• Control break through estrogen production due to
proliferation of granulose cells
• Increases the duration of elevated FSH, resulting in
multifollicular development
• A higher clinical pregnancy rate/ treatment cycle (27.3% vs
11.8%)

24. Letrozole step up CC
No of follicles
15mm on the 2.2±1.5 1.5±0.6
day of hcg
ET on day of hcg 9.8±2.1 9.9±2.1
(Mitwally et al, Fertil Steril, April 2008, Suppl-2)

25. A novel protocol for CC resistant PCOS
• Dose : 2.5mg of letrozole daily starting from day 1 of the
menses for 10 days
• Results in continuous production of FSH for a longer duration
• Maintains FSH levels above the threshold
• These FSH levels allow a greater cohort of small follicles to
reach maturity
• Results in multifollicular development (3.0 follicles >/=
18mm) & higher pregnancy rate (17.4% vs. 12.4% with short
letrozole group i.e. 5 days)
• Fertility and Sterility July 2009; Vol.92(1): 236-9

27. Short letrozole Long letrozole
No of follicle > 14mm 2.1 3.7
No of follicle >18mm 1.8 3.0
Endometrial thickness 10.4 11.2
Serum E2 315.5 338
pregnancy/cycle 12.4 % 17.4%
Miscarraige/ patient 17.9% 18.4%
(Badawy et al, Fertil Steril, 2008)

28. Highlights
• Ten day letrozole protocol extends FSH window
• Higher number of patients ovulated
• No of dominant follicles were more
• Pregnancy rates significantly greater
• No extra cost

30. • 308 IVF cycles
IVF via the mild ovarian stimulation protocol
Clomiphene citrate onD3 and recombinant FSH
on D 5 GnRH antagonist when dominant
follicle14mm.
• Serum LH was measured at the time of GnRH
antagonist administration and at the time of hCG
inj.

31. Comparison of
letrozole
with continuous gonadotropins
and clomiphene-gonadotropin combination
for ovulation induction in 1387 PCOS women
after clomiphene citrate failure: a randomized
prospective clinical trial.
Ganesh A, et al. J.Asst.Reprod.Genet 2009.

32. Aim: compare the efficacy of letrozole. with that of
rFSH and CC /rFSH for OS in IUI cycles.
METHODS: Randomized, prospective, single-blinded
clinical trial. 1387 PCOS pts after CC failure
randomized into 3grps.
• Group A - letrozole,
• Group B -CC with two doses rFSH
• Group C -continuous rFSH day 2 onwards until
hCG injection.

33. Ganesh A, et al. J.Asst.Reprod.Genet 2009.
RESULTS:
Group A(let) B(CC+G) C (gonad)
OR 79.30%, 56.95% 89.89%
Cycle cancel 20.70% 43.05% 10.11%,
PR 23.39% 14.35% 17.92%,
MR 13.80% 16.67% 14.52%,
CONCLUSION:
Letrozole appears to be a suitable ovulation inducing
agent in PCOS women with CC failure and is found to
be most effective when baseline estradiol level >60
pg/ml

34. Letrozole for ovulation induction and controlled ovarian
hyperstimulation. Curr Opin Obstet Gynecol. 2010 Pritts
EA
A review of studies:
• Letrozole has fewer side effects, and a shorter half-life
than CC. Safety profile better than CC.
• No demonstrable effect upon E R.
• Effective in - chr.anovulation, POR, UI and patients with
Ca desirous of childbearing.
• PR equal or superior to CC - bio-equivalent doses.
• Congenital malformation rate lower than CC .
Letrozole is associated with a > chance of uncomplicated
preg. & a < risk of multiple pregnancy than CC.
Ekerhovd E. 2009

35. CC resistance - Role of LOD
Rapid < secretion of all ov. hormones - < of the follicle excess
& (intra-ovarian) androgen levels – restores feedback H-P
axis – appropriate gonadotrophin secretion.
• LOD is effective in restoring ovulation in 70 % CC resistant
women.
• Should be resorted to in special circumstances. >LH, when
monitoring an issue.

36. Low dose Step-up regimen
It allows the FSH threshold to be
reached gradually, minimizing
excessive stimulation decreasing the
risk of multifollicular response

37. Low dose Step-up regimen
Starting dose = 37.5-75 IU/day
If
Follicle > 12 mm
E2 > 400US
Continue
1 FSH/day
no response  1.5 FSH/day
for 1 more week (max. 3 amp.)
Endocrine Rev. 1997; 18: 71
37.5-75 FSH/hMG/day
Day 3 Day 75 days

38. Low dose Step-down regimen
Day 3Day 3
2 FSH/d 1½ FSH/d 1 FSH/d
3-4 amp.3-4 amp.
U/S & E2
Foll >11 mm
U/S & E2
Foll >11 mm
2-3 days2-3 days
U/SU/S
hCG
D7
FSH dose may be high or low:
• Need to dose.
•Need to dose by one ampoule.

40. Low dose Step-up Step-down
regimen
one FSH/day
Day 3
step-up till 14 mm foll.
step-down
hCG

41. Multiple Follicular
Development

42. • Rationale of COH:
To disturb the normal relationship between
FSH&Eby increase FSH available to follicles
other than the dominant follicleincrease
total number of follicles that reach the pre
ovulatory stage.

43. Terminologies

46. Special situation…PCOS

47. • Problems in ovulation induction & COS.
–Failure of stimulation
–Hyperstimulation
–Premature Lutenization

48. PREMATURE LUTEINIZATION

49. Pre-ovulatory Progesterone
Peri-ovulatory progesterone plays an
indispensable role in ovulation and
luteinization
controls tissue remodelling of the ovulatory
follicle.
anti-atretic and pro-differentiative actions.
Chaffin et al HR2000.
Facilitates positive feedback response of E2 on LH.
Induces second FSH surge ensuring completion of FSH
action in the follicle.

50. Premature Luteinization
• P elevation >1- 1.5 ng/ml on or before HCG
administration during COH & the concomitant
differentiation of GC to luteal cells.
• P:E ratio>1: P ng/ml X1000 / E2 pg/ml
• It is unrelated to pre-ovulatory LH rise.
• Prevalence rate : 13% – 61% (Younis et al FS
1998). Even in D/R cycles.

51. PL - pathogenesis
• LH content of HMG. However it is seen to
occur in cycles using rFSH.
• Dose of exogenous FSH administered - PL
reflects the mature GC response to high FSH
exposure. Ubaldi et al. & Filicori et al.
• Cumulative P secretion by a cohort of
maturing follicles. ( Younis 2001, Segal et al
FS 2008).

52. PL - pathogenesis
• Gene involvement.
– A series of genes expressed differentially in PL
GCs have been identified. These genes were vital
for steroidogenesis, translation, cell adhesion,
cellular metabolism, lipid metabolism, and
apoptosis. Xingyo long et al 2009
• PL could reflect low ovarian reserve. (Younis
1998 .study in pt.’s with UI).

53. Can we prevent PL?
• Use of GnRH analog.
– GnRH agonist.
– GnRH antagonist.
• Use lower dose of gonadotrophins for COS.
• Progesterone antagonist . – mifepristone
effective in < PL, effect on ER a concern.
Escuredo et al. spain.

54. Impact on Endometrium
• The impact of ovarian stimulation on the integrity and function of the
endometrium the duration of the window of implantation is dependent on
levels of estrogen administered in the luteal phase (Ma et al 2003).
• The evidence for a detrimental effect of ovarian stimulation on
endometrial receptivity is now clear, and raises questions as how best to
ameliorate this effect. The move towards
• milder stimulation regimens, increasing use of cryopreservation of
embryos and transfer in a natural cycle are addressing this issue. However,
adjunctive therapies aimed at improving endometrial receptivity in IVF
cycles have as yet not been shown to be beneficial.
• Ma et al. PNAS 2003 100:2963
• Horcajadas JA et al. J Clin Endocrinol Metab. 2008 Nov;93(11):4500-10
• Boomsma CM, et al, Fertil Steril 2010 (e-pub ahead of print)
• AB-07

55. Endometrial Biopsies D 25
Left : Normal Histology : Patchy areas of stromal decidualization,
Scattered stromal large granulated lymphocytes, Glands without
cytoplasmic vacuoles.
Right : Patchy areas of stromal decidualization and scattered
stromal large granulated lymphocytes, but the glands are highly
vacuolated, typical of D18 glands.

56. Luteal Phase defect in stimulated cycles

59. •Thank you
•Welcome…..Women’s Health Congress,IMA
E mail - bdhorepatil@gmail.com
Website – www.punefertilitycenter.com

60. • What happens when, after three cycles of Clomid,
there is no ovulation? The initial step would be to add
something to the routine. Perhaps the physician will
decide to add intrauterine insemination (IUI) to the
Clomid regimen for the next three cycles. Another
option may be to try direct stimulation of the ovaries
with an injectable drug. There are several injectables to
choose from, for instance, Bravelle, Ovidrelle, or
Follistim - all of which come with their own specific list
of side effects which are less than desirable. Of primary
importance is the need for careful monitoring as all of
these drugs, including Clomid, can cause birth defects
if they are taken after conception.

61. Shortcomings of COH
• Multifollicular recruitment bring about a sudden increase in
estradiol (E2) serum levels that is enough to induce an LH
surge while follicular growth is still in progress.
• 24% of IUI cycles suffer from premature LH surge and this
can result in undesired premature luteinization leading to IUI
procedure cancellation.
• An increased risk of multiple pregnancies and the occurrence
of ovarian hyperstimulation syndrome (OHSS).

62. Rationale for using GnRH antagonists
• The premature LH surge
20%Cycle cancellation
Loumaye 1990
• The need both to control LH surge as well as to avoid
complex,high cost stimulation regimens involving
prolonged agonist treatment,led to the development
of newer antagonistic analogues
(Albano et al 2000, Borm and Mannaerts 2000,
Olivennes et al 2000, Fluker et al 2001, European
and Middle East Orgalutran Study Group 2001)

63. Mechanism of action
• Antagonist treatment is highly dose dependent,
• relying on the balance between endogenous GnRH present
and antagonist administered
(Felberbaum et al 1995)
• Almost directly after GnRH antagonists enter circulation
any growing follicle or corpus luteum present will be adversely
affected,while uterine bleeding is expected to occur within
48h
• Within 6-8 hours of administration, any imminent LH surge is
blocked
(Klingmuler at al 1993)

65. • What happens when, after three cycles of Clomid,
there is no ovulation? The initial step would be to add
something to the routine. Perhaps the physician will
decide to add intrauterine insemination (IUI) to the
Clomid regimen for the next three cycles. Another
option may be to try direct stimulation of the ovaries
with an injectable drug. There are several injectables to
choose from, for instance, Bravelle, Ovidrelle, or
Follistim - all of which come with their own specific list
of side effects which are less than desirable. Of primary
importance is the need for careful monitoring as all of
these drugs, including Clomid, can cause birth defects
if they are taken after conception.

66. Shortcomings of COH
• Multifollicular recruitment bring about a sudden increase in
estradiol (E2) serum levels that is enough to induce an LH
surge while follicular growth is still in progress.
• 24% of IUI cycles suffer from premature LH surge and this
can result in undesired premature luteinization leading to IUI
procedure cancellation.
• An increased risk of multiple pregnancies and the occurrence
of ovarian hyperstimulation syndrome (OHSS).
A prospective randomized trial. Hum Reprod. 2007 Jan;22(1):101-8.

67. Rationale for using GnRH antagonists
• The premature LH surge
20%Cycle cancellation
Loumaye 1990
• The need both to control LH surge as well as to avoid
complex,high cost stimulation regimens involving
prolonged agonist treatment,led to the development
of newer antagonistic analogues
(Albano et al 2000, Borm and Mannaerts 2000,
Olivennes et al 2000, Fluker et al 2001, European
and Middle East Orgalutran Study Group 2001)

68. Mechanism of action
• Antagonist treatment is highly dose dependent,
• relying on the balance between endogenous GnRH present
and antagonist administered
(Felberbaum et al 1995)
• Almost directly after GnRH antagonists enter circulation
any growing follicle or corpus luteum present will be adversely
affected,while uterine bleeding is expected to occur within
48h
• Within 6-8 hours of administration, any imminent LH surge is
blocked
(Klingmuler at al 1993)

69. Two Principle differences….

76. Letrozole should be the
drug of first choice in
CC resistance and CC failures.
ISA help in IR PCOS
Gonadotrophins should be the last resort
LOD can be used in special situations

77. PREMATURE LUTEINIZATION

78. Pre-ovulatory Progesterone
Peri-ovulatory progesterone plays an
indispensable role in ovulation and
luteinization
controls tissue remodelling of the ovulatory
follicle.
anti-atretic and pro-differentiative actions.
Chaffin et al HR2000.
Facilitates positive feedback response of E2 on LH.
Induces second FSH surge ensuring completion of FSH
action in the follicle.

79. Premature Luteinization
• < PR due to
– lowered oocyte quality
– premature resumption of meiosis
– impaired endometrial receptivity –
hastening of secretory changes.

80. Endometrial Biopsies D 25
Left : Normal Histology : Patchy areas of stromal decidualization,
Scattered stromal large granulated lymphocytes, Glands without
cytoplasmic vacuoles.
Right : Patchy areas of stromal decidualization and scattered stromal
large granulated lymphocytes, but the glands are highly vacuolated,
typical of D18 glands.

81. Luteal Phase defect in stimulated cycles

82. • In prediction of the outcome ovarian response two issues are keynote. One is that the relation between FSH dose
• level and the response of the ovaries in terms of number of dominant follicles growing, has not been clearly
• established. So far, evidence has shown that the dose response curve is very steep, indicating that with only small
• increases in FSH level single follicle growth is turned into maximal ovarian response. This implies that in the vast
• majority of patients the dosage administered (150 IU and over) will provide maximal stimulation of the ovaries.
Only
• factors like high BMI or the presence of specific FSH receptor variants may contribute to the creation of FSH levels
• that lead to submaximal stimulation of the ovaries.
• Second, the use of maximal stimulation dosages as a rule implies that the ovarian response is not principally
dependent
• upon the dose of FSH applied, but on the size of the cohort of the FSH sensitive antral follicles present in the
ovaries
• at the time of stimulation. The antral follicle cohort size is fully determined by the ovarian reserve status of the
• individual woman and expressed by female age and possibly by ovarian reserve tests. These two factors are
therefore
• the most important tools for response prediction.

83. a
• the impact of ovarian stimulation on the integrity and function of the
endometrium
• the duration of the window of implantation is dependent on levels of estrogen
administered in the luteal phase (Ma et al 2003).
• The evidence for a detrimental effect of ovarian stimulation on endometrial
receptivity is now clear, and raises
• questions as how best to ameliorate this effect. The move towards milder
stimulation regimens, increasing use of
• cryopreservation of embryos and transfer in a natural cycle are addressing this
issue. However, adjunctive therapies
• aimed at improving endometrial receptivity in IVF cycles have as yet not been
shown to be beneficial.
• Ma et al. PNAS 2003 100:2963
• Horcajadas JA et al. J Clin Endocrinol Metab. 2008 Nov;93(11):4500-10
• Boomsma CM, et al, Fertil Steril 2010 (e-pub ahead of print)
• AB-07

84. Different protocols and endocrinology

85. 85