for post graduate medical students

1. Pituitary Gonadotropin
Dr. Koustuv Chowdhury
MDPGT
Pituitary
Gonadotropin

2. Outline:
 Basic Physiology
 History
 Structure of Gonadotrophins
 Mechanism of action
 Regulator of Gonadotrophin production
 Physiological function of Gonadotrophin
 Use of Gonadotrophin
 Gonadotrophin Agonist and Antagonist
 Abnormality due to Gene Mutation
 Summary

3. Hypothalamic Neurons of Median Eminence
Dopamine influence
Secret GnRH or FSH/LHRH
Pulsatile secretion every 60 – 120 minute
GnRH transported to anterior Pituitary by Hypothalamic
– Hypophyseal Portal Venus System
Under influence of GnRH pulse
Anterior Pituitary secrets FSH & LH
LH a/k/a ICSH (Interstitial Cell Stimulating Hormone)
Basic Physiology

4. High E without P
stimulate
release of GnRH,
FSH, LH
• Moderate E inhibits
• Secretion of GnRH, FSH, LH
Hypothalamus
GnRH
Low E with
P promote
GnRH,, FSH,
LH secretion
FOLLICULAR
PHASE
Anterior pituitary
FSH +
LH
LUTEAL
PHASE
High E with P
inhibit GnRH,
FSH, LH
release
Primary
follicles
Secondary
follicles
Graafian
(mature) follicle
Ovulation
Corpus Iuteum
Corpus
albicans
6 14 281 Menstrual phase Proliferative phase (preovulatory)
Ovulation Secretory phase (postovulatory)
Menstrual
phase
ESTROGEN
1. PROGESTERONE + ESTROGEN
2. INHIBIN - secetion of GnRH and
FSH (-ive feedback)
3. RELAXIN – Relaxes and dilates
uterine cervix
Ovariancycle
Uteriancycle
FSH
ELH
P
+
-
)
+
-
-
+-
-
+
c
Correlation of Ovarian and Uterine Cycle With Hormonal Secretions ( Key : E – Estrogen ;P –
Progesterone)
PRINCIPLES OF PHARMACOLOGY
Fig 43.2
43

5. Gn-R
expression
• granulosa cells
• theca cells
• luteal cells
• interstitial cells
Extragonadal
Gn-R
 the uterus,
sperm,
seminal vesicles,
prostate,
skin,
breast,
adrenals,
thyroid,
neural retina,
neuroendocrine cells,
and (rat) brain.
physiologic role largely
unexplored

6. History
FSH originally first derived from animal (pregnant mare serum)
or human (postmortem Pituitary Glands) sources.
Abandoned because of safety concern.
Gonadotrophin first extracted from urine in 1940 (HCG).
In 1949 HMG was discovered.
Further purification produce FSH only product in 1980
Subsequent development of highly purified urinary FSH. (HP-
Hfsh)
In 1980 DNA Technology enabled the development of
recombinant human FSH (rec-Hfsh)
In 2000 recombinant human LH become available
In 2001 with the launching of recombinant human HCG
complete the recombinant portfolio.

7. Structure of Gonadotrophin
FSH, LH & hCG are Glycoprotein
They are composed of 2 non covalently linked
protein subunit – α & β
 α subunit contains 92 amino acids. It is identical in
FSH, LH & hCG.
 β sub unit confer unique receptor specificity and
differential biological and immunological properties.
Glycoproteins have two types of glycosylation
pattern. –
O - linked
N – Linked

8. Sialylation and Sulfonation
 Gonadotrophin further modified by addition of
Sialic Acid (Sialylation) or Sulfonic group
(Sulfonation) to the Carbohydrate moieties
 Increased number of Sialic Acid increases half –life.
 Increase number of Sulfonic group disappear faster
from the circulation due to their affinity to specific
receptor in the liver.
 Removal of the Carbohydrate moieties diminishes
gonadotropic activity.

9. FSH
 α subunit = 92 amino acids
 β subunit = 111 amino acids
 Both Sialylation and Sulfonation modulate the
half life of human gonadotrophin.
 Increased Sialic Acid = Hormone remains
longer in circulation.
 Production of different isoforms is controlled by
combination of steroidal feedback and GnRH
 Higher the estradiol level, lower the FSH
Sialylation
 The isoform profile is more acidic during early
follicular to mid follicullar phase and become
more basic before ovulation.

10. Characteristics of Native FSH
isoforms
Isoform Sialic acid
content
Biologic
activity
In vitro –
Half life
Predoimin
ance
Acid High Low Long Early/mid
– follicular
and Luteal
phase
Basic Low High Short Late
follicular/p
re
ovulatory
phase

11. LH
•α sub unit = 92 amino acids
•β sub unit = 121 amino acids
•β sub unit contains a single site of N-linked
Glycosylation and less Syalic Acid. So LH has a short
half life of only 20 to 30 minutes.
• Basic LH isoform are seen at mid cycle due to
decreased Sulfonation, and increased Sialylation.
•This change in isoform is physiologically important
for triggering ovulation.

12. HCG
 HCG β sub unit contains two sites of N-linked
Glycosylation (LH has single site).
 Higher no. of Glycosylation and Sialic Acid residues
than LH, HCG exhibits a longer terminal half life of
24 hours of intravenous injection in comparison
with 30 minutes for LH.

13. Structural characteristic of
FSH, LH, HCG molecules
Alpha
Subunit
Beta
Subunit
N-Linked
glycisilatio
nsites
(alpha)
N-
Linked
glycisila
tionsites
(beta)
O-
Linked
glycisila
tionsites
(beta)
Carboxyl
terminal
segment
Terminal
half life
FSH 92AA 111AA 2 2 - Absent 3-4h
LH 92AA 121AA 2 1 - Absent 20-30h
HCG 92AA 145AA 2 2 4 Present 24h

14. Mechanism of Action
 The LH and HCG are mediated by LH receptor
 FSH are mediated by FSH receptor
 Both are GPCRs
 FSH & LH receptor activate adenylyl cyclase/ cAMP
activate PKC & Calcium signalling pathway.

15. Function of FSH
In female
1. Estrogen Production,
to some Progesteron
2. Follicular
Development
3. Gametogenesis
4. Regulates Menstrual
Cycle
In male
1. Spermatogenesis

16. Function of LH
In female
1. Corpus Luteum
formation
2. Ovulation induction
3. Estrogen and
Progesteron
production
4. Regulates Menstrual
Cycle
In male
1. Androgen and
Testosteron Synthesis
by interstitial cell of
leydig.

17. Lower pulse frequency favours – FSH Secretion
Higher pulse frequency favours – LH Secretion
Positive Regulator
 Activin
Negative Regulator
 Inhibin
 High blood level of
Estrogen, Progesteron &
Testosteron
GnRH is the main regulator of FSH & LH Secretion
REGULATOR

18. KISSPEPTIN

19. Altered Physiology of HPG
Axis
Altered Physiology Pathological State
Premature activation of HP System Precocious puberty
Derangement of HP System Hypogonadotropic hypogonadism
Delayed puberty
Excess production of FSH & LH PCOD
Overian enlargement
Overian rupture
Decrease production of FSH & LH In female – Sterility and Amenorrhea
In male – Oligospermia and

20. Gonadotrophin preparation
Gonadorelin
• Synthetic human GnRH
• Short plasma half life ( 4-6 minutes)
• Administered IV or SC
• Pulsatile IV administration every 1 – 4 hour
stimulates FSH/LH secretion
• Continuous therapy desensitises.
Use
• Diagnostic
To determine whether hypogonadism due to defect
in anterior pituitary release of LH or hypothalamic
release of LHRH
If 100μg SC or IV rise in LH level the
defect in hypothalamic level.
• Treatment of infertility
a) Male
b) Female

21. hMG
•First extracted from urine of post menopausal
woman in 1949.
•It contains varying amount of FSH, LH & hCG (only
5% of pure forms)
•First hMG preparation was registered in Italy in
1950
•In 1999 purified hMG were introduced
•Both conventional and highly purified hMG are
commercially available in FSH:LH 1:1.

22. Urinary FSH
•In 1980, pure urinary FSH preparation were
produced by removing LH with polyclonal antibodies.
•Urofollitropin or purified urinary FSH still contain high
amount of urinary protein.
•Use of highly specified monoclonal antibody extract
FSH and produce highly purified FSH (HP-FSH)

23. Recombinant FSH Preparation
 Gene coding for human FSH α subunit and β subunit
incorporated into nuclear DNA of a host cell via plasmid
vector.
 Chinese hamster ovary cell line used to produce it.
 1995, first rec-hFSH (Folitrophin-α) licensed for clinical use.
 In 1996, Folitrophin-β licensed
 In Folitrophin-α --------------- 2 separate vectors for each
subunit used
 Folitrophin-β-------------- single vector is used.

24. Differences between hMG and
FSH formulatios
Purity (FSH
content)
Mean
Specific FSH
activity
U/mg
protein
LH activity
(IU/vial)
Injected
Protein
hMG <5% -100 75 -750
HP – hMG <70% 2000-2500 75 -33
Folitropin β >99% 7000-10000 0 -81
Folitropin α >99% 13649 0 -71

25. Recombinant LH Preparation
Purity
(LH
content
)
FSH
Activity(I
U/vial)
LH
activity(
IU/vial)
Hcg
content
(IU/vial)
Specific
Activity
(LH/mg
protein)
Lutropin - α >99% 0 75* _ 9000
Follitropin
α+lutrofin α 2:1
ratio
>99% 150 75 _ 9000
HP-hMG Unknow
n*
75 75* -8
Lutropin – α is highly pure and high biological activity
It is the only recombinant form of human LH developed for
Ovarian Stimulation.

26. GNRH Agonists and Antagonists

27. Mechanism of Action:
 GnRH agonists binds to specific G-protein coupled receptors
on the pituitary gonadotrophs.
 Prolonged activation of GnRH receptors by GnRH agonists
leads to desensitization and down regulation of GnRH
receptors, thereby suppressing gonadotrophin (FSH & LH)
secretion.

28. GNRH Agonist Use Dose
Leuprolide 1. Endometriosis
2. Uterine Fibroids
3. Prostate Cancer
4. Precautious puberty
3.75 mg/month im
3.75 mg/month im
7.5 mg/month im
0.05 mg/kg/day SC
Nafarelin 1. Endometriosis
2. Uterine Fibroids
3. Precautious puberty
0.2 mg BD(Nasal spray)
50-400 microgram BD
1.6 mg/day
Goserelin 1. Endometriosis
2. Uterine Fibroids
3. Precautious puberty
3.6 mg SC in plant every 28
days
Buserelin 1. Endometriosis
2. Uterine Fibroids
Histrelin 1. Endometriosis
2. Uterine Fibroids
Triptorelin Prostate CA 3.75 mg/month IMDepot
INDICATION

30. Side Effects of GnRH Agonists
A. Hypoestrogenism
1. Hot flush
2. Headache
3. Osteoporosis
4. Breast Atrophy
B. Local site pain
C. Bronchospasm and Anaphylaxis (Histerelen)
D. Sinusitis (Nafarelin)

31. GnRH
ANTAGONISTS

32. GnRH Antagonists:
 Ganirelix.
 Cetrorelix.
 Degarelix.
 Abarelix.
 Elagolix (under Phase III Clinical Trial).
 Relugolix (TAK-385  under clinical trial).
 KLH-2109 & ASP-1707 (under development).
 JV – 1 – 36 (under clinical trial)

33. Ganirelix
 Half life: 12.8-16.2 hours.
 Onset of Action: within 8 hours.
 Highly protein bound.
 Metabolized by the liver.
 Excreted through feces (75%) and urine (22%).
 Peak plasma concentration is attained within 1 hour.
 Contraindicated in Pregnancy, Lactation & Hypersensitivity
responses.
Adverse Effects:
Pelvic pain.
Ovarian Hyperstimulation Syndrome.
Abdominal pain.
Vaginal bleeding.
Injection site reactions.

34. Cetrorelix
 Half life: 20-63 hours.
 Onset of Action: within 12 hours.
 Highly protein bound.
 Excreted through feces and urine.
 Peak plasma concentration is attained within 1-1.5
hours.
 Contraindicated in Renal impairment, Pregnancy,
Lactation & Hypersensitivity responses.
Adverse Effects:.
Ovarian Hyperstimulation Syndrome.
Headache.
Nausea.
Elevated hepatic enzymes
Increase plasma
HDL levels.

35. Degarelix
 Onset of Action: within 3 days.
 Highly protein bound.
 Metabolized by the liver.
 Excreted through feces (70%-80%) and urine
(20%-30%).
 Contraindicated in Pregnancy.
 Adverse Effects:
 Hot flushes.
 Injection site pain & reactions.
 Weight gain.
 Elevated liver enzymes.

36. Use of GRH Antagonists
Indication Drug
Advance Prostate CA Abarelix (withdrawn in 2005)
Degarelix (SC)
Endometrios Elagolix
Uterine Fibroids Cetrolix (3mg SC once weekly for 2
months)
Adjunct during IVF

37. GNRH Antagonist Protocol
 Day – 3 of Menstrual period (USG for basic evaluation)
 Medication was initiated with rFSH at the day of USG
 More than 35 years 3 ampule (225 iu of Gonal F daily) X 5 days
 Less than 35 years 2 ampule (225 iu of Gonal F daily) X 5 days
 After 5 days TVS done to see the follicle
 rFSH dose depends on no. & size of follicle.
 Cetrorelix (0.25 mg/day) SC day in the morning (8AM – 12 noon)
from day 6 to the day of hCG administration
 TVSB done on 8th , 10th , 12th , day

38. Oocyte retrieval
 Gonal F and Cetrorelix administered continuously
untill 3 follicles reached more than 17 mm
 hCG (10,000 IU) then administered and serum
concentration of Estradiol, LH & Progesteron was
tested.
 Oocyte were retrived 34-48 hour after hCG
injection.
 Embryo transfer carried out 72 hours after Oocyte
retrieval. (Maximum 3 embryos are transferred in
each patient )

40. Summary