The document discusses recent advances in controlled ovarian stimulation (COS) protocols for infertility treatment. It describes how recombinant gonadotropins are purer and safer than urinary gonadotropins, while having similar clinical efficacy. COS protocols now utilize GnRH antagonists to simplify treatment and decrease the risk of ovarian hyperstimulation syndrome compared to agonists. Overall, novel COS protocols incorporate recombinant gonadotropins and GnRH antagonists to provide patient-friendly stimulation with good outcomes.

1. “Understanding Infertility” – New Delhi, India – August 2011
Recent Advances in
Stimulation Protocols
Esteves, 1

2. 1. Present an evolution perspective of
gonadotropins
2. GnRH analogues for controlled ovarian
stimulation (COS)
3. Protocols for COS: taking advantage of
new products
Esteves, 2

3. Esteves, 3

4. Patients Doctors Industry

6. 600,000 120,000
Worldwide urine quantity (1000 litres)
Number of donors
600 120
1965 1975 1985 1990 1995 2000
Donors from different regions around the world
Esteves, 6

7. Up to 65% of couples dropout
from IVF without achieving
pregnancy before they
complete 3 cycles1-5
Reasons1,5,6
Psychological burden 49%-26%
Prognosis 40%-23% Oocyte retrieval 52%
Embryo transfer 29%
Cost of treatment 23%-0%
Injections 29%
Relationship/divorce 15%-9%
Physical pain 20%
Physical burden 7-6% Blood tests 14%
1. Olivius K t al, Fertil Steril 2004;81:258; 2. Land JA et al, Fertil Steril 1997; 68:278; 3. Schroder AK, et al, RBM Online 2004; 5:600; 4.
Osmanangaoglu K et al, Hum Reprod 2002; 17:2655; 5. Rajkhowa M et al, Hum Reprod 2006; 21:358; 6. Brandes M et al, Hum Reprod 2009;
24:3127; 7. Hammarberg K et al, Hum Reprod 2001; 16:374.

8. Typical Cycle (long protocol):
Daily SC GnRH-a: x21
FSH/LH: x10-15
hCG: x1
Progesterone: x14
Blood tests: x4-7
Number of sticks: 36-57

9. Gonadotropins: better today
Milestones in the development of gonadotrophins
2001
1940 1962 Full recombinant 2008
First hCG 1993 2000 First
Purified u-hMG gonadotrophin
extracted from First highly purified First r-hLH r-hLH+r-FSH
(Pergonal®) and u- portfolio available
human urine FSH-only product launched combined
hCG (Profasi®)
launched (Luveris®) (Pergoveris®)
become available
(Metrodin HP®)
1949 1980s 1995 2001 2002
First hMG extracted First FSH-only First r-hFSH First r-hCG First filled-by-mass
from urine pools product launched launched launched product launched
(Metrodin®) (GONAL-f®) (Ovidrel®/Ovitrelle) (GONAL-f® FbM)
Milestones in the development of r-hFSH
1980 1983 1985 1988 1992
-subunit -subunit -FSH gene cloned and Human FSH expressed First pregnancy
sequenced sequenced expressed in fibroblasts in CHO cells with r-hFSH
1. Bassett et al. Reprod Biomed Online 2005;10:169–177 CHO, Chinese hamster ovary
2. Lunenfeld. Hum Reprod Update 2004;10:453–467

10. Culture
media Bioreactor Harvest
Cell attachment and Concentration of
proliferation supernatant
r-hFSH production and Chromatographic
secretion purification
steps
Collection of cell
culture supernatant Ultrasterile filtration
medium containing
Characterization
r-hFSH
and full QC of
In-process QC bulk r-hFSH
Esteves, 10

11. Gonadotropins: better today
Quality
Safety
Key Clinical
issues Efficacy
Patient
conven-
ience
Esteves, 11

12. From urinary to recombinant
Choragon
Urinary -hCG from different manufacturers
Ferring PregnylOrganon
-
hMG HP
30% of impurities
per vial with
39 different
proteins identified
(varied from
batch to batch)
Protein
FSH
impurities
Laboratoire De Spectometrie de MBO – October/2009
van de Weijer et al. Reprod Biomed Online 2003;7:547–557
Esteves, 12 Kuwabara Y et al, J Reprod Med 2009; 54:459–466

13. Impurities
cannot be
associated
with a better or
worse outcome
but certainly
are not needed
for COH
u-hMG HP
Molecular (5 batches)
r-hFSH
weight (follitropin
markers alfa)
Esteves, 13 Merck Serono data on file

14. Australia 1996:
Recognizes higher UK 2003: Metrodin HP
standards of purity and withdrawn
safety of recombinants; unacceptable risks given
Encourages their use over that there are alternatives
urinary, human derived.
Esteves, 14

15. Purity Mean specific Injected
(FSH FSH activity protein
content) (IU/mg protein) per 75 IU
(mcg)
hMG < 5% ~100 ~750*
hMG-HP < 70% 2000–2500 ~33*
r-hFSH
Follitropin beta – 7000–10,000 8.1*
Follitropin alfa > 99% 13,645 6.1
Esteves, 15 1. Bassett et al. Reprod Biomed Online 2005;10:169–177

16. Novel analitycal
Conventional method
Bioassay
Physiochemical
technique
High
in vivo variability Minimal batch-to-
(rat) batch variability
(1.6%)1,2
Urinary gonadotropins
Follitropin beta Follitropin alfa
Esteves, 16
1. Bassett et al. Reprod Biomed Online 2005;10:169–177; 2. Driebergen et al. Curr Med Res Opin 2003;19:41–46

17. Concept of Dose Precision
Clinical implications
Batch variability Batch variability
+20%, -25% 2%
IU
Risk of OHSS
270
16.5 mcg
225
(225 IU)
170
Poor response
Bioassay Filled by Mass
Urinary and Follitropin beta Folitropin alfa

18. Patient Compliance
sc rec FSH (pen device)
sc rec LH
sc hCG (prefilled syringe)

19. Evidence-based truth:
Scientific truth:
recFSH is more
potent recFSH is purer
↑ 3.1 oocytes
(Bosch, 2008) Non urine-
extracted product
↑ 1.8 oocytes
(MERIT, 2006) Recombinant
technology
↑ 2.8 oocytes
(Hompes, 2007)
Esteves, 19

20. GnRH analogues in ART
LH surge prevention by GnRH agonists
pyro (Glu) – His – Trp – Ser – Tyr – Gly – Leu – Arg – Pro – Gly – NH2
Activation of the Regulation of Regulation of receptor
GnRH receptor receptor affinity biological activity

21. GnRH analogues in ART
LH surge prevention by GnRH antagonists
1 2 3
pyro (Glu) – His – Trp – Ser – Tyr – Gly – Leu – Arg – Pro – Gly – NH2
Activation of the
Antagonistic Regulation of Regulation of receptor
GnRH receptor
effect receptor affinity biological activity

22. LH surge prevention
GnRH antagonists
Agonist
Follicular
LH, FSH
30
E2 , P 4
Luteal
20
2-4 weeks
10 Synchronized follicles
0
6
• Half-life ~20h (Cetrorelix)
Antagonist 5 • Suppress LH by 80% of
4 baseline levels
LH (IU/L)
3 Antagonist
2
1
0
-6 0 6 12 18 24 30 36 42 48
Hours

23. LH surge prevention
GnRH antagonists vs agonists
Prevent OHSS
Can be integrated by GnRH-a
No flare in spontaneous
Single or multiple effect with No hormonal and OI cycles Antagonist
dose GnRH administration
possible cyst withdrawal
antagonist protocol formation Gonadotropin administration
Less gona-
Can exclude dotropins
early
pregnancy
Flare up Pituitary
effect suppression
Gonadotropin administration
Long GnRH
agonist protocol Longer Agonist administration
treatment
Pre-treatment cycle Treatment cycle

24. Day 1 Rec-hCG
Follicle (prefilled syringe)
of rFSH
13 mm or GnRH-a
rec-FSH (fbm) Individualized rec-FSH dose
112.5-450 UI (recLH supplementation)
Vaginal
menses progesterone
GnRH antagonist
Day 2 or 3 0.25 mg/day (flexible)
of menses
Day 1 Rec-hCG
Day 6 (prefilled syringe)
of rFSH
of rFSH
Cycle
recFSH (fbm) Individualized rec-FSH dose
day 21
112.5-450 UI (rec-LH supplementation) Vaginal
progesterone
GnRH Agonist
menses
Day 2-5 of menses
Esteves, 24

25. Group A (hMG; N=299)
Group B (HP-hMG; N=330)
Group C (r-hFSH; N=236)
Day
Day 1 Day 6 of hCG
Cycle
day 21 Gonadotropin rFSH/hMG
Individualized dose
112.5-450 UI Vaginal
progesterone
Agonist (nasal spray): Nafarelin acetate (400 mcg/day; fixed)
menses
Esteves, 25 Day 2-5 of menses

26. Outcome Measure HMG HP-hMG r-hFSH P-
n=299 N=330 n=236 value
Total gonadotropin dose (IU) 2,685 2,903 2,268 <0.01
Retrieved oocytes (N) 10.9 10.7 10.8 NS
MII oocytes (N) 8.9 8.9 8.7 NS
2PN fertilization rate (%) 72 72 71 NS
Implantation rate (%) 24 27 23 NS
Live birth rate per cycle (%) 24.4 32.4 30.1 NS
Moderate/severe OHSS(%) 2.3 1.8 1.3 NS
Esteves et al, Reprod Biol Endocrinol. 2009; 7:111

27. % Cycles with “Step-down”
during ovarian stimulation
53.4*
*P<0.01
18.7 20.3
HMG HP-HMG rec-hFSH (fbm)

28. Total Dose per Live Birth (IU)*
To achieve a
10.000 live birth,
52.2% 9,690
21-52% more
7.000 21.6% 7,739 HP-hMG and
6,324* hMG was
3.000
required
0 compared with
r-hFSH HP-hMG hMG
r-hFSH
* Mean total dose per cycle/Live birth rate (≤35 years)

29. Infertility Specialists Gonadotropin
Choice in South America?
r-hFSH
r-hFSH+hMG
hMG
1600
1400
1200
Gonadotrophin segment split
1000
(75 IU equivalent units)
800
52%
600
400
200
-
1998 1999 2000 2001 2002 2003 2004 2005
39%
Total r-hFSH Total u-FSH Total hMG/LH
9%
Esteves, 29 Data supplied by IMS and REDLARA 2007

30. GnRH antagonists vs agonists
Meta-analysis
Kolibianakis et al (2006)2
N studies 22
Included non peer-reviewed data No
Included IUI cycles No
N patients 3176
Odds ratio (probability of live birth) 0.86 (0.72-1.02; p=.08)
Duration of stimulation -1.54 days (OR: -2.42; -0.66; p=.0006)
Oocytes retrieved -1.19 (OR: -1.82; -0.56)
Risk of severe OHSS OR=0.61 (0.42; 0.89; p=.01)*
*For every 59 women treated with a GnRH agonist vs GnRH
antagonist, one additional case of severe OHSS will occur
Esteves, 30

31. GnRH agonist for triggering oocyte
maturation – Humaidan et al
Hum Reprod Update 2011
 GnRH-a triggering (0.2-1.5 mg): antagonist protocol
 Reduced if not eliminated risk for OHSS
 In specific high risk patients for OHSS and egg
donation programs should become the choice
 Challenge is to rescue luteal phase insufficiency
 Modified luteal support improved delivery rate
hCG bolus OPU day (1,500 UI) or 3x 500 UI boluses;
recLH; intense progesterone + estradiol; combined
Delivery rates: 18% risk difference favoring hCG
(before) X 6% risk (after)
Esteves, 31

32. Esteves, 32

33. Recent Advances in Stimulation
Protocols
 Recombinant gonadotropins purer and safer
compared to urinary gonadotropins
 Recombinant gonadotropins more patient-
friendly and have similar (or better) clinical
efficacy compared to urinary gonadotropins
 COS using GnRH antagonists simplify
treatment and decrease OHSS
 Novel COS protocols involve recombinant
products and GnRH antagonists
Esteves, 33