UNDERSTANDING DOWN REGULATION GnRH Agonists and Antagonists In ART by Dr. Narendra Malhotra, Dr. Jaideep Malhotra

1. narendra malhotra
jaideep malhotra
www.malhotrahospitals.com
www.rainbowhospitals.org
UNDERSTANDING DOWN REGULATION
GnRH Agonists and Antagonists In ART

2. • The IVF story is today more
than 35 years old
• Acceptance to this method of
procreation is now widely
acceptable
• More than 4.3million babies
have been born by IVF
• Protocols have been changing
everyday for better patient
comfort and convenience
The IVF Story 2010
Congrats Bob Edwards
For the Nobel prize 2010
And Indira Hinduja for
Padamshree 2011

3. CHANGING APPROACH
YESTERDAY
TODAY
TOMORROW
CONVENTIONAL
MILD/MINIMAL
INDIVIUALISED
More Oocytes
Supraphysiological E2
Agonist protocols
More embryos/OHSS
Multiple pregnancies
Cryopreservation
Less Oocytes
Optimal E2
Antagonist protocols
No OHSS
No multiple pregnancies
Lower cost
Optimal Oocytes/Embryos
No OHSS
No Multiple pregnancies
Lower cost
PATIENT FRIENDLY

4. Problems of Ovarian Stimulation
• Poor responders
• Hyper responders: OHSS & multiple pregnancies
• Premature LH surge

5. Controlled ovarian stimulation with
recombinant FSH and GnRHa has been the
key factor in success of IVF.
Orvieto.R 2004

6. agonists
• The main aim of introducing the agonists
to IVF protocol was to prevent the
premature LH surge
TO START USING AGONISTS FOR ART
WE NEED TO UNDERSTAND DOWNREGULATION

7. understanding downregulation

8. GnRH
• is produced & secreted by a
group of neurons located in
the arcuate nucleus of medial
basal hypothalamus &
preoptic area of ventral
hypothalamus.
• These molecules are
transported to pituitary by
intricate network of
capillaries.

9. GnRH
• Is a decapeptide with a short half life.
• regulates trophic function of anterior
pituitary gland.
• Plays a key role in functioning of
hypothalamic-pituitary axis
• Regulates production & release of both
LH & FSH in the gonadotrophs of ant.
Pituitary.

10. PHYSIOLOGY OF GnRH ACTION
• Gonadotrophs of ant.pituitary release FSH and LH
rhythmically in response to rhythmic release of GnRH.
• This secretion of GnRH is governed by GnRH pulse
generators which resides in mediobasal hypothalamus.
• This pulse generator is operative during fetal life and
in early infancy but is suppressed by intrinsic
inhibitory control

11. Puberty starts when GnRH pulse generator is
released from suppressive effects of gonadostats
at around 8.5 –12.5 yrs in girls and 9.5-13.5 yrs in
boys.

12. • GnRH secretions are
negatively modulated by
gonadal steroids.
• However under certain
circumstances estradiol
can have positive
influence,on GnRH
secretion like prior to
ovulation.
• Prolonged strenuous
exercise and severe weight
loss can inhibit GnRH
secretion.

13. The number of receptor in the membrane
of gonadotrophs change with altered
physiological conditions and are
subjected to up & down regulation.

14. ••It is possible to readily synthesise GnRH agonists by
using solid peptide synthesis.
It is possible to introduce amino acid substitution to
prolong the half life .
Position 6 of amino acid sequence is important for
inactivation.
2 and 3 for gonadotropin release
1,6 and 10 for binding
1 2 3 4 5 6 7 8 9 10
GnRH shows two phases with half lives of 2-8 mins
for fast and 15-60 mins for slower phase

15. GnRH AGONISTS
SYNTHETIC AGONISTS
• INCREASE STABILITY AND BINDING AFFINITY
TO ITS RECEPTORS.
• 100-200 TIMES HIGHER.
What is the need to synthesise agonists?

16. GnRH agonists
• First synthesised in 1972
• produced by altering the amino acids at
position 6 and /or 10,resulting in compounds
with higher affinity for the GnRH receptors
and a long half-life due to their resistance to
cleavage by endopeptidases & reduced
susceptibility to enzymatic degradation
• Half life is of few hrs.

17. • It is possible to readily synthesise GnRH
agonists by using solid peptide
synthesis,also possible to introduce amino
acid substitution to prolong the half life .
• Multiple amino acid substitution permits
synthesis of antagonists.which bind to
GnRH receptors and provide competitive
block for naturally synthesised GnRH.

18. GnRH Analogs: Historical Perspective
• Prior to use of GnRH analogs, up to 20% of treatment
cycles were cancelled due to spontaneous LH surges that
interferes with follicular development1
• GnRH agonists were the first GnRH analogs available for
suppression of LH secretion1
– Clinical pregnancy rate was significantly improved
– Decreased the likelihood of cycle cancellation
1. Hughes et al. Fertil Steril. 1992;58:888.
2. Devroey et al. Hum Reprod. 2009;24:764.

19. AGONISTS
Seven approved analogues are;
• Buserelin
• Leuprorelin
• Goserelin
• Histrelin
• Deslorelin
• Nafarelin
• Triptorelin

20. GnRH AGONISTS
Agonist Structure Route Dose, mg Potency
GnRH =1
Leuprorelin D-Leu6, Subcutaneous 0.5-1.0 15-100
Pro9-NHET intramuscular, 3.75-7.5*
depot
Buserelin D-Ser(Bu)6, Subcutaneous 0.2 100
Pro9-NHET intranasal 0.9-1.2
Histrelin D-His(BZ)6, Subcutaneous 0.1 100
Pro9-NHET
Nafarelin D-[Na(2)6] Intranasal 0.4-0.8 200
Triptorelin D-Trp6 Intramuscular, 2-4* 100
Polymer
Goserelin D-Ser(tBu)6 Subcutaneous, 3.6* 230
aza-Gly10 implant

21. Preparations Available
• Zoladex depot
• Decapeptyl depot monthly/3 monthly
• Lupride depot & daily
• Many other preperations daily dose
(blossom/luprofact/lucrin/gonapeptil)
• Nasarel (multiple doses)
• Buserlin/Zerlin/suprafact sub cut daily dose

22. MODE OF ACTION
• GnRH agonists intially have similar short term
effects as GnRH.
• They bind to the receptors which undergo
dimerization & initiate a cascade of events
culminate in synthesis & secretion of FSH & LH
• FLARE UP 4 times FSH and 10 times LH release

23. UP REGULATION
• An increase in number of receptors
amplifies the synthesis and release of
gonadotrophins.
• This occurs during pulsatile endogenous
GnRH release or its superactive agonists.
• Thus upregulation of receptors facilitates
ready synthesis and release of large
amount of gonadotropins when needed.
• FLARE UP EFFECT

24. • Over the time b/o over abundance of agonists with
longer half life, the dimer form of receptors is favoured
and the receptors go into the cell and cannot come back
so
• Cannot respond to subsequent pulses of GnRH
• Thus the gonadotrops become desensitised and this is
called down regulation of GnRH receptors

25. DESENSITIZATION
• When the GnRH receptors exposed to
GnRH agonists for a prolonged period,
the cells lose their ability to respond to
the stimulus with their original
sensitivity.
• Process is rapid & reversible.
• Process operates at both the receptor
level & by post receptor modification
WHAT IS
DOW
N
ON
Down regulation

26. • GnRH is released in a pulsatile
fashion but a continuous supply of
GnRH suppresses gonadotrophin
secretion by desensitization of
gonadotrophs.
• This property is utilised in number
of therapeutic options.

27. • Slowly and steadly FSH and LH
conc falls,and so does estradiol
and progesterone
levels,indicating inhibition of
ovarian steroidogenesis and
follicular growth.
• This property is utilised in
number of therapeutic options.

28. CLINICAL APPLICATIONS
• Diagnostic use
• Gonadal stimulation
• ART
• Precocious puberty
• Endometriosis
• Fibroids
• Hormone dependent tumours
• Hirsutism
• DUB
• Endometrial ablation
• PMS
• Prostatic malignancy
• Trigger ovulation

29. Diagnostic use: GAST TEST
Similar to clomiphene challenge test used
to evaluate hypo-thalamic pituitary
ovarian axis.

30. Gonadal stimulation
• Pulsatile administrationof GnRH in
physiologic doses mimics endogenous
release & stimulates the ovaries.
• Sub –cut 5-20 mcg every 90 mins or IV
20-25mcg every 90mins. Especially for
hypothalamic amenorrhoea and PCOD

31. Precocious Puberty
• Suppression of pituitary-gonadal
function is the aim
• Long term agonists are safe and
effective
• Can be given for yrs till normal age of
puberty reaches.
• More striking aspect is regression of
secondary sex characters

32. Endometriosis
• Ability to produce amenorrhoea and
anovulation is the main basis
• Mild to moderate stages medically
managed.
• Severe managed with surgery &/or in
combination with medical.
• Depot preparations are widely used.

33. Though surgical removal is the most effective method
But the main goal of pre-operative agonists is;
*Reduce blood loss during subsequent surgery
*reduce the tumor size, lesser operating time
*lesser complications
*where surgery not possible
WITH 3 MTHS TT 40-60% REDUCTION IN SIZE
FIBROIDS

34. Agonists and
ART
• Down regulation of HPO axis -
prevention of LH surge
• Suppression of endogenous FSH / LH,
elevated basal LH levels (PCOS) and
active endometriosis
• Recruitment of a uniform cohort of
follicles
• Programming of cycles
• Avoiding weekends

35. AGONISTS AND ART
• augmentation of follicular
activity
• achieve higher number of
oocytes
• higher number of embryoes
• increased pregnancy rates

36. MODE OF DELIVERY
ORALLY ANALOGUES ARE RAPIDLY DESTROYED
MAYBE GIVEN:
• PARENTRALLY
• NASAL SPRAY
• VAGINAL PESSARIES
• DEPOT PREPARATIONS ARE ALSO AVAILABLE
• BIOLOGICAL EFFICACY OF NASAL SPRAY IS
LOWER THAN SUB CUTANEOUS

37. Assess Downregulation
RULE OF FIVES TO ASSESS DOWN REGULATION
LESS THAN 5 FOLLICLES
LESS THAN 5 MM SIZE OF FOLLICLES
LESS THAN .5 MM ENDOM THICKNESS(actually .3)
LESS THAN 50 pgm ESTRADIOL(actually 30)
LESS THAN .5 ngm PROGESTERONE
TVS

38. GnRh Agonists
1. Long Protocol
2. Short flare up
3. Ultra short
4. Ultra long

39. agonist
agonist
agonist
2 3 4
2 3 4 5 6 7 8 9 10 11 12
21 (luteal) 2 3 4 5 6 7 8 9 10 11
hCG
hCG
hCG
150 /225 FSH / hMG daily
150 rec FSH / 225 hMG daily
Conventional Protocols
150/225 FSH / hMG daily
Flare up of a dying corpus luteum
Elevated LH levels in the early
follicular phase
Follicular dominance - one follicle
races ahead of the others
- no uniform cohort
Premature LH surge
Better suited for poor responders
(advantage of the flare-up effect)
Reduced consumption of gonadotropins
hence reduced cost of therapy
Premature LH surge
Increased consumption of gonadotropins
result of complete suppression
of endogenous FSH and LH
Increased cost of treatment
Inadvertent administration of GnRHa
during early pregnancy
Reversibility takes weeks

40. Disadvantages of agonists
• Increased consumption of
gonadotropin as a result
of complete suppression
of endogenous FSH and
LH
• Increased cost of
treatment
• Inadvertent
administration of GnRHa
during early pregnancy

41. Side effects
•Hypo estrogenic state;
•Hot flushes
•Head aches
•Vaginal dryness
•Mood instability
•Insomnia
•Decreased libido
•Dizziness
•Altered lipid profile
•Osteoporosis

42. estrogen deprivation
• GnRH agonists administration in luteal phase ,a
stage of hypoestrogenism is induced ,during this
period women may suffer from common
symptoms of hypoestrogenism;
• Hot flashes
• Insomnia
• Short term memory lapses
• Head aches

43. THERE AROSE A NEED TO
LOOK FOR ANOTHER
DRUG TO PREVENT LH
SURGE

44. • The introduction of GnRH antagonists in
assisted reproductive technologies (ART) at
the beginning of this decade was met with
mixed reactions. A series of theoretical and
evidence-based advantages supported their
use
Tarlatzis et al., 2006

45. GnRH Agonist vs GnRH Antagonist:
Mechanism of Action
Flare-up
GnRH receptor
desensitization
Long GnRH Agonist
Protocol
Pituitary
GnRH
Gonadotropin
suppression
FSH
LH
GnRH
receptor
Hypothalamus
GnRH
agonist
GnRH
FSH
LH
GnRH Antagonist
Protocol
Pituitary
GnRH
Direct
gonadotropin
suppression
GnRH
receptor
Hypothalamus
GnRH
antagonist
FSH
LH
GnRH
agonist

46. Thus the need for antagonists is a
real one

47. ANTAGONISTS
• There is no dimer formation
• It outcompetes the GnRH for the GnRH
receptors and blocks the ability of GnRH to
initiate dimer formation
• As a result monopolization of GnRH
antagonist, there is no secretion of FSH and
LH.
• As long as sufficient antagonist is present
suppression is sustained

48. GnRH ANTAGONISTS
• Competitively bind to pit.GnRH receptors &
do not release gonadotrophins.
• Possible to modulate hormone suppression by
dose .
• Do not cause initial flare up.
• Within hours secretion of gonadotropins
decreases.
• Do not cause pituitary exhaustion.
• Can respond to adequate stimulus immediately.

49. • Introduction of GnRH antagonists
provides prevention of premature LH
surge but with distinct advantages
– More timely suppression of LH
– No symptoms associated with GnRH
agonist flare
and downregulation
1. Hughes et al. Fertil Steril. 1992;58:888.
2. Devroey et al. Hum Reprod. 2009;24:764.

50. ANTAGONISTS
• Multiple amino acid
substituition permits
synthesis of
antagonists.
• Which bind to GnRH
receptors and provide
competitive block for
naturally synthesised
GnRH.

51. Antagonists
• Cetrolix
• Iturelix
• Azaline B
• Ganirelix
• Abarelix
• Antarelix

52. Antagonists Results in Rapid Reduction in LH
0
2
4
6
8
6 7 8 9 10 11 12
Days
Serum
FSH/LH
(IU/L)
0
3
6
9
12
Orgalutran
(ng/mL)
FSH LH Orgalutran
-32%
-74%
Serum concentrations of Orgalutran, FSH, and LH following the
final (day 7) Orgalutran 0.25 mg injection
Out and Mannaerts. Hum Fertil. 2002;5:G5.
Oberye et al. Fertil Steril. 1999;72:1006.

53. Controlled Ovarian Hyperstimulation Regimens for
Assisted Reproductive Technology
GnRH Antagonist
Protocols
GnRH Agonist
Protocols
225 IU per day
(150 IU Europe)
Individualized Dosing of FSH/HMG
250 µg per day antagonist
Individualized Dosing of FSH/HMG
GnRHa 1.0 mg per day
up to 21 days
0.5 mg per day of GnRHa
225 IU per day
(150 IU Europe)
Day 6
of FSH/HMG
Day
of hCG
Day 1
of FSH/HMG
Day 6
of FSH/HMG
Day
of hCG
7 – 8 days
after estimated ovulation
Down regulation
Day 2 or 3
of menses
Day 1
FSH/HMG
OCP

54. KEY DIFFERENCES BETWEEN
ANTAGONISTS AND AGONIST
• No initial flare effect
• No estrogen deprivation
symptoms
• Shorter tt protocol
• Reduced gonadotropin use
• Flexibility
• Rapid reversibility
• Recovery phase is much
shorter(2-4 days)
• Patients more inclined to drop
out of agonist regime.

55. NO INITIAL FLARE UP
• Initial response of agonists is stimulatory.
• For achieving a clinically suppressed
state –14-21 days.
• Antagonists in contrast have immediate
action.

56. • GnRH antagonists are not started until
stimulation is well under way and
estradiol levels are high
• So no estrogen deprivation symptoms

57. SHORTER TREATMENT
• Duration of antagonists cycle is much
shorter
• Difference is 4 days vs 22 days (north
american study group trial)
• Albano et al. difference is 6 days vs 27
days

58. REDUCED GONADOTROPIN USE
• North American ganirelix study group
trial showed that duration of
recombinant FSH was slightly shorter (8
days)as compared to leuporide group(10
days)
• Total dose was also lower( 1800 i.u vs
2025 i.u)

59. Antagonist what more…
• Holds promise for the PCOS
• Reduction in the incidence of OHSS
• Agonist can be used as a trigger instead of
hCG to drastically reduce risk of OHSS
– More physiological
– Lower half life (60 mins vs 32 – 34 hrs)
– Lower incidence of OHSS

60. FLEXIBILITY
• ANTAGONISTS SHOW
RAPID SUPPRESSION OF
ENDOGENOUS LH/FSH
DURING ALL PHASES OF
MENSTRUAL CYCLE

61. Fixed Start Day Protocol
Day 6 of stimulation
1 2 7 10 12
0.25 mg
Cetrorelix
HCG OPU/IUI
Rec FSH

62. Flexible Start Day Protocol
1 7 10
0.25 mg
Cetrorelix
HCG OPU/IUI
1 6 7 8 9 1011 13
USG > 14 mm
E2 > 200-300 pg/ml
Rec FSH/hMG

63. Timing of antagonist addition
• Controversial
• Introduction too soon counterproductive and
leads to shut off potential endogenous FSH,
interfering with early follicular recruitment.
• Too late will not prevent premature LH surge

64. RAPID REVERSIBILITY
• Because GnRH receptors remain
functional although blocked ,if sufficient
GnRH or GnRH agonists are given to out
compete the antagonists, secretion of
FSH and LH results

65. GnRH antagonists in normal
responders
• Start FSH or HMG on d-2 and as soon
the follicle reaches 12-14 mm add
antagonist till hCG day.

66. POTENTIAL OTHER USES
• Suppressing LH surge in cases where pt
forgets to take agonists by chance.
• Triggering ovulation in antagonist cycle by
agonist in hyperstimulated cases
• Synchronising ovum recepient cycles

67. Major points to consider in
normal responders
• Expect a high estradiol level on D-6
• Do not cutback on dose of recombinant
FSH
• OC pretreatment can be included for cycle
timing,but donot expect better response
• Use flexible start of antagonist ,not
preferably after D-8

68. IN POOR
RESPONDERS
• Who is a poor responder ?
• Acc. To reproductive medicine associates of
NY
• Exhibits 1 or 2 follicle response
• Peak estradiol levels not >500 pg at hCG
administration
• Requiring excessive amount or duration of
gonadotropins(>450 I.u)

69. Soft Ovarian Stimulation:Poor responders
Craft et al
– CC/Gn with antagonist in poor responders & PCOS
USG
E2
USG
E2
USG
E2
HCG
5000
10,000
ET
D2
ET
D3
ET
D5
Blastocyst
OPU
IUI
35-37
hr
1 2 3 4 5 6 7 8 9 101112
Progesterone
IM
Oral
Vaginal
CC
GnRH
antagonist
FSH/HMG

70. Advantages of antagonist cycles
• Same success rate as agonists
• Lower cancellation rate
• Reduction in duration of GnRH agonists
• Lower risk of OHSS
• Avoidance of estrogen deprivation
symptoms
• Lower dropout rates.

71. Issues to be settled

72. Q.1
• In a fixed dose regime of antagonist,is there
a need to increase the dose of FSH or LH
supplementation while starting the
antagonist on D-5 ?

73. Best Practices for GnRH Antagonist Protocols: Evidence
Does Not Support Supplementation of LH Activity
29.5
31.7
0
5
10
15
20
25
30
35
40
35
32.1
0
5
10
15
20
25
30
35
40
Bosch et al1 Meta-analysis2
recFSH recFSHrecFSH
+ rLH
hMG
Ongoingpregnancyrate
(%)
P=0.6
1
P=NS
1. Bosch et al. Hum Reprod. 2008;23:2346.
2. Baruffi et al. Reprod Biomed Online. 2007;14:14.
Ongoingpregnancyrate
(%)

74. Q. 2
Fixed or flexible protocol?

75. Ganorelix / cetrorelix day 9
OR
Gano / cetro day 6 - 12
2 3 4 5 6 7 8 9 10 11 12
Single shot

76. Q.3
• PCOS with delayed cycles ,when to start
antagonists ?

77. Q.4
• What is the time gap between the last
antagonist injection and hCG?

78. •Prevent premature LH surge
•Protect oocyte from deleterious effect of high LH
•Compared to agonist :-
• shorter cycle of treatment,
• more conception,
•fewer miscarriages,
• reduced amount of gonadotropins needed
• monofollicular growth,
• reduced risk of OHSS & multiple pregnancy
GnRH antagonist in IUI cycles

79. The ideal Indian protocol
100 mg CC /L day
225 hMG / 225 IU rec FSH
0.25 mg antagonist/day
2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17
7500units hCG

80. “Think it over...”
Understanding of the pituitary down
regulation enables you to take maximum
advantage of the interplay exo and
endogenous hormonal milieu of the body.
BUT
In future antagonists might play a major
role in ART cycles.

82. Antagonists Phase 3 Trials:
Duration of FSH Stimulation
9
8
9
10 10
11
0
2
4
6
8
10
12
EU
trial
NA
trial
EU/ME
trial
DaysofFSH
stimulation
Triptorelin
Buserelin
Leuprolide
Orgalutran
Out and Mannaerts. Hum Fertil. 2002;5:G5.

83. Antagonists Phase 3 Trials:
Amount of recFSH Required
1,500
1,800
1,350
1,800
2,025
1,800
0
500
1,000
1,500
2,000
2,500
Triptorelin
Buserelin
Leuprolide
IUofFSHrequired
EU
trial
NA
trial
EU/ME
trial
Orgalutran
Out and Mannaerts. Hum Fertil. 2002;5:G5.

84. Increased consumption and days of
stimulation

85. Antagonists Phase 3 Trials: Number of Oocytes
and Good Quality Embryos
EU
trial
NA
trial
EU/ME
trial
Triptorelin
Buserelin
Leuprolide
8.7
11.7
7.9
9.7
14.1
9.6
0
2
4
6
8
10
12
14
16
3.3
4.3
2.7
3.5
4.8
2.9
0
1
2
3
4
5
6
7
8
9
10
EU
trial
NA
trial
EU/ME
trial
EU = European; NA = North American; ME = Middle East.
Out and Mannaerts. Hum Fertil. 2002;5:G5.
Orgalutran
Number
Number
Good
Quality
Embyros
Oocytes

86. Antagonists Phase 3 Trials: Ongoing
Pregnancy Rate per Started Cycle
20.3
30.8 31
25.7
36.4
33.9
0
5
10
15
20
25
30
35
40 Orgalutran
Triptorelin
Buserelin
Leuprolide
EU
trial
NA
trial
EU/ME
trial
Ongoingpregnancyrate(%)
Out and Mannaerts. Hum Fertil. 2002;5:G5.

87. Pregnancy outcome slightly better
with agonist

88. MORE EXPENSIVE MORE
EFFECTIVE

89. • It is not uncommon to see that pts who are
normally ovulating when kept on long luteal
protocol,then given stimulation, fail to
stimulate and become refractory.
• These pts have been oversuppressed

90. Antagonist vs long protocol poor
responders

91. Modifying protocols for poor
responders
• Minidose agonist protocol
• Microdose flareup protocol
• Antagonist protocol

92. GnRH Antagonists vs Agonists
poor responders or PCOS
• Two studies on poor responders comparing
the protocols demonstrated that both had
similar pregnancy rates
• Four RCTs related to PCOS also
demonstrated similar results.
Greisinger G et al 2006

93. GnRH Antagonist Strategy Is Associated With
a Lower Dropout Rate vs Long GnRH Agonist Strategy
SET = single embryo transfer; DET = double embryo transfer.
Adapted from Verberg et al. Hum Reprod. 2008;23:2050.
Likelihoodofcontinuingtherapy(%)
Cycle number
95.9%
93.7%
78.6%
88.3%
75.9%
P=0.034
GnRH antagonist plus SET
GnRH agonist plus DET
Continuation of therapy following each cycle
50
60
70
80
90
100
0 1 2 3

94. GnRH Antagonists For Treatment Of PCOS
Why should the antagonists be used rather than the agonists?
Three reasons
• Significantly less gonadotropin is required
• Better compliance due to the number of injections are approximately 4 to 5 with
the GnRH antagonist compared with anywhere between 21 to 25 with the GnRH
agonist.
• Reduced risk of developing OHSS.
• GnRH agonist can be used to trigger ovulation
Fertility and Sterility Vol.80, Suppl.1, Jul 2003

95. Ongoing pregnancy rate per couple with one cycle of FSH/IUI with and
without GnRH antagonist treatment.
Hum. Reprod. Update 2009;15:265-277
© The Author 2009. Published by Oxford University Press on behalf of the European Society of
Human Reproduction and Embryology. All rights reserved. For Permissions, please email:
journals.permissions@oxfordjournals.org

96. GnRH Antagonists Are Associated With More Favorable
Outcomes vs GnRH Agonists Among Women at High Risk
for OHSS
56.3
43.7
76.3
27.6
32.2
67.8
96.6
11.5
0
10
20
30
40
50
60
70
80
90
100
Canceled
cycles
Oocyte
retrievals
Embryo
transfer
OHSS
GnRH agonist
GnRH antagonist
P<0.001
P<0.001
P=0.003
P=0.006
Percent
Ragni et al. Hum Reprod. 2005;20:2421.
Investigator-driven, prospective observational study of women (N=87) at
high risk for OHSS, who were treated with a GnRH antagonist protocol
following a previous cycle with a GnRH agonist protocol

97. OHSS

98. Antagonists vs Long GnRH Agonist:
Effects on the Endometrium
• No relevant alteration in endometrial thickness or
pattern was observed with standard or high dose of Orgalutran
• Endometrial dating, estrogen and progesterone receptor expression,
and endometrial surface structure were unaffected with Orgalutran
treatment
• Buserelin was associated with indications of an arrest
in endometrial development
• Expression of “window of implantation” genes with Orgalutran
treatment more closely paralleled the pattern observed during a natural
cycle compared with buserelin
Simon et al. Hum Reprod. 2005;20:3318.

99. Antagonists vs Long GnRH Agonist:
Effect on Expression of Implantation Genes
Number of Genes With Differential Regulation Between the Natural Cycle (Day LH +7)
and the Treatment Regimens (Day HCG +7) and Within the Window of Implantation*
Regimen No. of Genes With Expression Increased; Decreased†
Window of Implantation Genes
All Genes
Typically Up-
regulated‡ (n=894)
Typically Down-
regulated‡ (n=504)
Orgalutran 0.25 mg/d ↑ 22; ↓ 69 ↑ 0; ↓ 46 ↑4; ↓ 0
Orgalutran 2 mg/d ↑ 88; ↓ 24 ↑ 0; ↓ 15 ↑7; ↓ 1
Buserelin long protocol ↑ 22; ↓ 100 ↑ 3; ↓ 76 ↑4; ↓ 2
*Differential regulation was defined as a ≥100% increase or ≥50% decrease in expression.
†Compared with values on day LH +7 of natural cycle.
‡Genes whose expression is typically upregulated or typically downregulated during the window of implantation (day
2–7 after the LH surge), according to Horcajadas et al 2005.
Adapted from Simon et al. Hum Reprod. 2005;20:3318; Horcajadas et al. Mol Hum Reprod. 2005;11:195.
Expression of “window of implantation” genes more closely
paralleled the pattern observed during a natural cycle with
Orgalutran than with buserelin

100. Pooled Antagonists Clinical Studies:
Development of Pregnancies and Birth Outcomes
No. (%)
Orgalutran GnRH Agonist
Ongoing pregnancies
(≥16 wk)
340 134
Singleton 258 (75.9) 91 (67.9)
Multiple 82 (24.1) 43 (32.1)
Total fetuses
(≥16 wk)
432 184
Total liveborn infants 419 (97.0) 179 (97.3)
Fetal deaths None None
Boerrigter et al. Hum Reprod. 2002;17:2027.
Data on birth outcomes pooled from 5 clinical studies in women
with ongoing pregnancy (N=474)

101. Pooled Antagonists Clinical Studies:
Neonatal Outcomes
Orgalutran GnRH Agonist
n=419 n=179
Mean gestational age, weeks 38.0 37.4
Term birth, no. (%) 306 (73.0) 107 (59.8)
Preterm birth, no. (%)
(≥33 weeks and <37 weeks)
87 (20.8) 47 (26.3)
Very preterm birth, no. (%)
(<33 weeks)
27 (6.2) 25 (14.0)
Mean birth weight, g 2,834 2,716
n=424 n=181
Congenital malformations (%) 7.5 5.5
Major malformations (%) 4.5 3.3
Data on neonatal outcomes pooled from 5 clinical studies in
women with ongoing pregnancy
Boerrigter et al. Hum Reprod. 2002;17:2027.

102. Antagonists Favorable Safety Profile
No. of Patients
Orgalutran
(n=1,217)
Buserelin
(n=236)
Leuprolide
(n=99)
Triptorelin
(n=111)
Subjects with AEs 280 (23.0) 74 (31.4) 41 (41.4) 24 (21.6)
Subjects with
SAEs 37 (3.0) 11 (4.7) 0 (0) 3 (2.7)
Subjects with
drug-related AEs
(including SAEs)*
36 (3.0) 9 (3.8) 6 (6.1) 2 (1.8)
*Judged by the investigator as definitely, probably, or possibly related to study drug.
Borm and Mannaerts. Hum Reprod. 2000;15:1490.
Fluker et al. Fertil Steril. 2001;75:38.
European and Middle East Orgalutran Study Group. Hum Reprod. 2001;16:644.
AEs With Orgalutran vs GnRH Agonists:
Pooled Data From Dose-Finding and Phase 3 Trials
No significant differences were seen in AEs between the 2 groups

103.  No OCP pretreatment
 Check patient cycle day 2
 FSH 100-225 IU
 Antagonist earlier than later
 LH not necessary
Suggested GnRH Antagonist Protocol
Cycle day 2
Transvaginal US +
(if desired) hormonal profile
This suggested protocol represents a “best estimate” given current data and
clinical experience. Further data are required before more concrete
recommendations can be made.
For regular IVF patients:
 5-9 antral follicles per
ovary
 Age <35 years
 No PCOS
 No history of poor
responses
 No endometriosis
Duration of treatment based
on clinical judgment in
consultation with patient
(usually 2 USs)
Cycle day 2/3
Start FSH 150-200 IU. Continue
Stimulation days 5-6
Start GnRH antagonist
administered daily. Continue
Monitoring according to clinic practice
 US (+ blood test if required)
 FSH dose adjustments may be considered
3 follicles 15-19 mm
Day of triggering
 Ensure interval between antagonist and hCG does not exceed 30 h
 hCG 5000-10,000 IU
Oocyte retrieval
36 h
YES
NO
US = ultrasonogram; OCP = oral contraceptive pill.
Devroey et al. Hum Reprod. 2009;24:764.

104. Antagonist as a Key Component of
Patient-Centered Therapy
• Good pregnancy rates
• Reduced risk of OHSS
• Reduction of stress associated with physical and
psychological treatment burden
– No side effects related to flare-up or downregulation
– Fewer injections
– Shorter treatment cycles
– Shorter duration of stimulation
Devroey et al. Hum Reprod. 2009;24:764.

105. ONE SIZE FITS ALL ?

106. Meta-analyses of GnRH Antagonists
vs GnRH Agonists: Pregnancy Outcomes
• The 2 studies had different results for
pregnancy outcomes
*Live birth rate included ongoing pregnancies (Al-Inany) or calculated rates (Kolibianakis).
OR = odds ratio.
1. Al-Inany et al. Cochrane Database Syst Rev. 2006;3:CD001750.
2. Kolibianakis et al. Hum Reprod Update. 2006;12:651.
Al-Inany1 Kolibianakis2
OR 0.82 0.86
95% CI 0.69-0.98 0.72-1.02
P value 0.03 0.085
Live Birth Rate* in Meta-analyses,
GnRH Antagonists vs GnRH Agonists

107. Meta-analyses Confirm That GnRH Antagonists Have a
Better Safety Profile vs GnRH Agonists
Al-Inany1 Kolibianakis2
Duration of analog
treatment
−20.90 days
(95% CI −22.20, −19.60)
−19.48 days
(95% CI −21.05, −17.91)
Duration of ovarian
stimulation
−1.54 days
(95% CI −2.42, −0.66; P=0.0006)
−1.13 days
(95% CI −1.83, −0.44)
Risk of
severe OHSS
OR 0.61
(95% CI –0.42, 0.89; P=0.01)
RR 0.46*
(95% CI 0.26, 0.82; P=0.01)
Interventions
to prevent OHSS
OR 0.44
(95% CI 0.21, 0.93; P=0.03)
*For every 59 women treated with a GnRH agonist vs GnRH antagonist, 1 additional case of severe OHSS will occur.
RR = risk ratio.
1. Al-Inany et al. Cochrane Database Syst Rev. 2006;3:CD001750.
2. Kolibianakis et al. Hum Reprod Update. 2006;12:651.

108. GnRH Antagonists vs GnRH Agonists: Patients
With Poor Response or PCOS
PCOS = polycystic ovary syndrome.
Griesinger et al. Reprod Biomed Online. 2006;13:628.
Poor Responders
PCOS
Clinical Pregnancy Rates

109. GnRH Antagonist and Long GnRH Agonist
Strategies Result in Comparable
Cumulative Pregnancy Rates
0
20
40
60
0 3 6 9 12
Adapted from Heijnen et al. Lancet. 2007;369:743.
GnRH agonist with DET
GnRH antagonist with SET
%ofpregnancies
leadingtotermlive
birth
Months since randomization
Singleton term
live birth
Proportion of pregnancies leading to cumulative
term live birth within 12 months after starting IVF

110. GnRH Antagonist and GnRH Agonist Strategies Result in
Shorter Treatment, Better Safety, and Lower Cost
Heijnen et al. Lancet. 2007;369:743.
GnRH
Antagonist
(n=444)
GnRH
Agonist
(n=325) P Value
Days of injections 8.5 25.3 <0.0001
Days of stimulation 8.3 11.5 <0.0001
Total dose of FSH (IU) 1,307 1,832 <0.0001
Incidence of OHSS (%) 1.4 3.7 0.04
Mean total costs €8,333 €10,745 0.006

112. Thanks

116. CLOMIPHENE CITRATE

117. EVALUATION

118. GONADOTROPHINS ROLE
• FSH IS THE KEY GONADOTROPHIN
• Plays crucial role in
• Recruitment
• Selection
• Dominance
during follicular phase
• Also stimulates granulosa cells LH receptor
expression
• Concept of FSH window

119. GnRH
• Plays a key role in functioning of
hypothalamic-pituitary axis .
• Is a decapeptide
• Regulates trophic function of anterior
pituitary gland.
• Regulates production & release of both LH
& FSH in the gonadotrophs of ant. Pituitary.

120. GnRH
• is produced & secreted by
a group of neurons located
in the arcuate nucleus of
medial basal hypothalamus
& preoptic area of ventral
hypothalamus.
• These molecules are
transported to pituitary by
intricate network of
capillaries.

121. • GnRH then binds to specific receptors in the
plasma membrane of the anterior pituitary
gonadotrops
• Where it causes synthesis ,storage and
release of luteinizing hormone and follicle
stimulating hormone.

122. PHYSIOLOGY OF GnRH ACTION
• Gonadotrophs of ant.pituitary release FSH and LH
rhythmically in response to rhythmic release of
GnRH.
• Pulsatile frequency varies from 71 mins in late
follicular phase to 216 mins in luteal phase .
• This secretion of GnRH is governed by GnRH pulse
generators which resides in mediobasal
hypothalamus.

123. • Puberty starts when GnRH pulse generator is released
from suppressive effects of gonadostats at around 8.5 –
12.5 yrs in girls and 9.5-13.5 yrs in boys.
•This pulse generator is operative during fetal life and
in early infancy but is suppressed by intrinsic inhibitory
control

124. • However under certain
circumstances estradiol
can have positive
influence,on GnRH
secretion like prior to
ovulation.
• GnRH secretions are
negatively modulated by
gonadal steroids.
• Prolonged strenuous
exercise and severe weight
loss can inhibit GnRH
secretion.

125. The number of receptors in the
membrane of gonadotrophs change
with altered physiological
conditions and are subjected to up
& down regulation.

126. UP REGULATION
• An increase in number of receptors
amplifies the synthesis and release of
gonadotrophins.
• This occurs during pulsatile endogenous
GnRH release or its superactive agonists.
• Thus upregulation of receptors facilitates
ready synthesis and release of large
amount of gonadotropins when needed.

127. GnRH AGONISTS
SYNTHETIC AGONISTS
• INCREASE STABILITY AND BINDING
AFFINITY TO ITS RECEPTORS.
• 100-200 TIMES HIGHER.
•GnRH has short half life of a few minutes & rapidly
degraded by endopeptidases.

128. •It is possible to readily synthesise GnRH agonists by
using solid peptide synthesis.
It is possible to introduce amino acid substitution to
prolong the half life .
Position 6 of amino acid sequence is important for
inactivation.
2 and 3 for gonadotropin release
1,6 and 10 for binding
•
1 2 3 4 5 6 7 8 9 10
GnRH shows two phases with half lives of 2-8 mins for
fast and 15-60 mins for slower phase

129. GnRH agonists
• First synthesised in 1972
• produced by altering the amino acids at position 6
and /or 10,resulted in compounds with higher
affinity for the GnRH receptors and a long half-
life due to their resistance to cleavage by
endopeptidases & reduced susceptibility to
enzymatic degradation
• Half life is of few hrs.

130. Agonists
and
ART
• Down regulation of HPO axis - prevention
of LH surge
• Suppression of endogenous FSH / LH,
elevated basal LH levels (PCOS) and active
endometriosis
• Recruitment of a uniform cohort of follicles
• Programming of cycles
• Avoiding weekends

131. GnRh Agonists
1. Long Protocol
2. Short flare up
3. Ultra short
4. Ultra long
(long follicular)
(long luteal)
(ultra long)

132. 2 3 4
2 3 4 5 6 7 8 9 10 11 12
21 (luteal) 2 3 4 5 6 7 8 9 10 11
hCG
hCG
hCG
150/225 FSH / hMG daily
150 /225 FSH / hMG daily
150 rec FSH / 225 hMG daily

133. • No difference between GnRH agonist or
antagonist in terms of cycle cancellation rate,
number of mature oocytes, or pregnancy rate.
• In analysis of GnRH agonist microdose flare
vs antagonist (4 trials) - significantly higher
rate of oocytes retreived in GnRH-a flare cycles
• no difference in pregnancy outcomes
• More trials needed
Franco, JG Jr et al, Reproductive Biomedicine Online, 2006

134. Stress Impacts IVF Success
• Several indicators of stress are significantly higher
in women undergoing stimulated IVF compared
with unstimulated IVF or undergoing gynecologic
surgery not related to infertility1
– Prolactin, cortisol, and state anxiety score all increased
during stimulated IVF treatment1
• Anxiety associated with IVF treatments may lead
to inadvertent noncompliance with recommended
gonadotropin dosing, a poor or excessive ovarian
response, and possibly a poor cycle outcome2
1. Harlow et al. Hum Reprod. 1996;11:274.
2. Noorhasan et al. Fertil Steril. 2008;90:2013.e1.

135. Stress Impacts IVF Success (cont’d)
• COS with less complicated treatment
regimens and fewer injections results in
significantly fewer (3% vs 23% with more
complicated regimens, P<0.001) patients
reporting that the stress associated with
treatment was considered “unacceptable”1
• The psychological burden of IVF treatments
was the primary reason cited among couples
who discontinued treatment before
2,3
1. Højgaard et al. Hum Reprod. 2001;16:1391; 2. Olivius et al. Fertil Steril. 2004;81:258;
3. Verberg et al. Hum Reprod. 2008;23:2050; 4. Smeenk et al. Hum Reprod. 2001;16:1420.

136. 4
4
5
5
6
7
16
18
0 5 10 15 20
Physical or Psychological Treatment Burden as
a Primary Reason for Dropout
Physical or psychological burden of treatment
Unknown
Relational problems/divorce
Others
Adoption
Poor embryo quality
Poor response/signs of ovarian aging
Ethical objections to ICSI treatment after failed
IVF treatment
Among 384 couples undergoing IVF treatment, 65 (17%) drop
Adapted from Verberg et al. Hum Reprod. 2008;23:2050.
Causes for Dropout
No. of patients

137. KEY
• Optimizing GnRH antagonists is clear
understanding of pharmacological
differences in agonists and antagonists
• More convenient
• Fewer injections
• Fewer side effects
• Easier to implement ,schedule and manage.

138. Structure of Orgalutran
NH2
NAcD2Nal D4CIPhe D3Pal DhArg(Et2) hArg(Et2) D-Ala
Orgalutran
NH2
pGlu His Trp Ser Tyr Gly Leu Arg Pro Gly
Endogenous GnRH
Ser Tyr ProLeu

139. 0
5
10
15
20
%ofpatientswithLH≥10IU/L
Daily dose of Orgalutran (mg)
The ganirelix dose-study finding group. Hum Reprod. 1998;13:3023.
Orgalutran Dose-Finding Study:
Suppression of LH Surges
Dose selected for phase
3 studies
0.0625
(n=31)
0.125
(n=65)
0.25
(n=69)
0.5
(n=69)
1
(n=65)
2
(n=30)

140. Orgalutran Phase 3 Trials:
Study Design
Inclusion Criteria
- Healthy women undergoing
COS for IVF or ICSI
- Women aged 18 to 39 years
- Regular cycles between
24 and 35 days
- BMI 18 to 29 kg/m2
Orgalutran
Puregon
Puregon
GnRH agonist
Downregulation
Cycle day
21-24
Day 2 or 3
of menses
hCG*
Day 6
of rFSH
*≥3 follicles ≥17 mm.
Adapted from Out and Mannaerts. Hum Fertil. 2002;5:G5.
Borm and Mannaerts. Hum Reprod. 2000;15:1490.
Fluker et al. Fertil Steril. 2001;75:38.
European and Middle East Orgalutran Study Group. Hum Reprod. 2001;16:644.
Luteal phase
support
Luteal phase
support

141. COH
• 3 choices
• FSH with antagonist
• Clomiphene with FSH/with antagonist
• Long luteal protocol

142. Advantages of GnRH Antagonists vs
GnRH Agonist Long Protocol
• No initial flare-up
• No estrogen deprivation symptoms
• Shorter treatment
• Reduced gonadotropin use
• Flexibility
• Administration only during period needed to
suppress endogenous LH surge
• Lower risk of OHSS
• Rapid reversibility of pharmacodynamic action

143. Meta-analyses of GnRH Antagonists
vs GnRH Agonists
• Two recent meta-analyses evaluated
randomized, controlled trials of GnRH
antagonists vs GnRH agonists in IVF1,2
• These meta-analyses included different studies,
used different measures of efficacy, and
reached different conclusions regarding
relative efficacy
– The Al-Inany study included non peer-reviewed
data and included studies on IUI
IUI = intrauterine insemination.
1. Al-Inany et al. Cochrane Database Syst Rev. 2006;3:CD001750.
2. Kolibianakis et al. Hum Reprod Update. 2006;12:651.