Thyroid Hormones for Medical Diagnosis

1. 2. Thyroid Hormones
Thyroid gland is the endocrine gland located at the upper portion of the trachea just below larynx.
It provided the body with two fundamentally different hormones calcitonin, thyroxin and
liothyronine.
1. Calcitonin
It is a peptide hormone produced by specialized cells in the thyroid gland and released in
response to high level of blood calcium. Where, it decrease the amount of calcium entering the
blood from boon through inhibition the bone resorption.
Structure
It is a linear polypeptide of 32 amino acids with an disulfide ring between Cys1 and Cys7
amino terminal. The carboxyl terminal amino acid is prolineaminde. The calcitonin from hog
and salmon similar to that of human only in 9 amino acid. Despite these differences they exhibit
the same activity in human.
H2N-cys1
-gly...........................cys7
-met................gly10
....................pro32
CONH2

2. The Structure-activity relationships
1. All the 32 amino acids are essential for activity, shortening causes loss of activity.
2. Substitution of prolineaminde to proline or proline ester result in loss of activity.
3. Loss or opining of disulfide lead to loss of activity.
4. Acetylation of the terminal amino or even deamination resulting in increase activity from 20-
50% and increase duration.
5. Replacement of met.8 by val. and phe.22 by tyr.22 lead to increase activity from 4 to 5 time,
increase duration of action by 2 time.
2. Thyroxin and Liothyronine
I
HO
I O
I
I
COOH
NH2
T4: thyroxin
2-(4-(4-hydroxy-3,5-diiodophenoxy)-
3,5-diiodophenyl)-2-aminoacetic acid
L-(-)-3,3,5,5-Tetraiodothyronin
L-(-)-3,5-Diiodo-O-(4-hydroxy-3,5-diiodophenyl)tyrosine
I
HO
O
I
I
COOH
NH2
T3: liothyronine
2-(4-(4-hydroxy-3-iodophenoxy)-
3,5-diiodophenyl)-2-aminoacetic acid

3. 2. Synthesis
HO
NH2
COOH
L- Tyrosine
1. nitration
2. acetylation HO
NHCOCH3
COOH
O2N
NO2
HO
NHCOCH3
COOEt
O2N
NO2
O
NHCOCH3
COOEt
O2N
NO2
H3CO
1. Ph. SO2Cl
OH
H3CO
2.
Esterification
H2/Ni, HNO2
I2/KI
O
NHCOCH3
COOEt
I
I
H3CO
HI/P
HBr/HOAc Thyroxine (T4)

4. The Structure-activity relationships
1. The aromatic ring are designated as α and β
according to their position from carbonyl group.
2. The aromatic ring should be perpendicular to each other.
3. The L-(-) form is the active form and used in hormonal deficiency.
4- The D-(+) form is used to lower the blood cholesterol level.
5- the ether oxygen impart non-linear relationship of α and β rings.
6. Replacement of O by S retains the activity.
7. 4`- OH is essential for activity, replacement with Me group resulted in loss activity
8. 4`- OH replacement with H give in vivo active compound.!!
9. T3 is more active than T4.
10. T3 is readily absorbed from GIT and rapidly cleared from blood, it loosely bound to plasma
protein.
11. The reversed T3 (r-T3) is in active due to the free rotation of phenyl ring.
I
HO
I O
I
I
COOH
NH2


Analysis
1. Oxygen flask method.
2. By measure solution in 0.1 N NaOH spectrophotometry at 325 nm against standard.
-
O O O O-
Uses
1. Hypothyroidism
2. Decrease thyroid enlargement

5. Metabolism
I
HO
I O
I
I
COOH
NH2
Conjucation Deiodination
Oxidative deamination
Ether bond cleavage
Deiodinations
T4
T3
T2
r-T3 inactive
5- deiodination
5- deiodination
5- deiodination
5- deiodination
- ring - ring
3,3- diiodo
inactive

6. Hyperthyroidism (thyrotoxicosis)
Causes
Graves disease (diffuse toxic goiter), toxic adenoma, goiter,
thyroiodites, ………….etc
Treatment
1. Removal of part or all of the thyroid gland by surgery or by destruction with radioactive I131.
2. Inhibition of thyroid hormone synthesis, through using of
a. alkylthiouracil b. Imidazoles
3. Blocked of hormone release by iodides which are used prior to surgery or in fatal thyrotoxic
crises
Antithyroid drugs
Hyperthyroidism is usually treated surgically, but the patient must be prepared by administration
of antithyroid drugs. Thiourea and related compounds posses antithyroid action but they are too
toxic. Thionamides are potent inhibitors of thyroid peroxidase enzymes responsible for
iodination of the tyrosine residue. These drugs have no effect on iodide trap or on thyroid
hormone release. The most clinically thionamides are thiourylenes (thiouraciles and imidazoles)
which are five or six membered heterocyclic derivatives of thiourea.

7. N
N
N
N
R
O
S
H
R SH
OH
H
Thioketo Thioenols
R Compound Activity
H
CH3
C3H7
Thiouracil
6- methythiouracil
6- Propylthiouracil
1
10
i. Alkyl thiouraciles
Synthesis
R O
O
OEt
R OH
O
OEt NH2
H2N S
N
N
R
O
S
H
H
Alkyl acetoacetate Thiourea
NaOEt

8. Analysis
1. Gravimetrically: by digest with HBr which converts sulfur to H2SO4. then, sulfuric acid
precipitated with BaCl, the formed BaSO4 washed, dried and weigh.
2. According to USP 2000
by worm with small amount of 0.1 N NaOH till complete to form its sodium salt. Add 0.1 N
AgNO3 and worm the mixture. The precipitated AgNO3 is then filtrated and the filtrate is
titrated with 0.1 N NaOH using bromothymol blue as indicator.
N
N
R
O
SH
H
AgNO3 2 OH-
N
N
R
O
SAg
Ag
2 HNO3
N
N
R
S
CH3
if R = H N
N
HS
CH3
R = H Methimazole
2- Mercapto-1- methylimidazole
R = COOEt Carbimazole
Ethyl 3- methyl-2-thioxo-4-imidazoline-1-carboxylate
ii. Imidazole derivatives

9. Synthesis
H3C N C S H2N H2C CH
OEt
OEt H3C
NH
C
H
N
CH2
CH
S
OEt
EtO
HN
N
S
CH3
ClCOOEt
-2 OEt
aminoacetal
methyl isothiocyanate
N
N
EtOOC
S
CH3
Carbimazole Methimazole
Analysis
1. Methimazole: by silver nitrate method
2. Carbimazole: by spectrophotometrically
Metabolism
Carbimazole is metabolized by ester
hydrolyses followed by decarboxylation
to give methimazole, so carbimazole can
be considered as prodrug for methimazole.
N
N
EtOOC
S
CH3
Carbimazole
N
N
HOOC
S
CH3
HN
N
S
CH3
Methimazole
Estrase - CO2

10. 3. Pituitary and hypothalamic hormones
The ventromedial part of hypothalamus connect with
the CSF and contain capillaries of the portal vein.
Various parts of the hypothalamus produce number
of peptide that inter the portal vein and reach the
pituitary gland situated immediately bellow the
hypothalamus. The pituitary is not part of CNS.
The hypothalamus peptides are hormones but
can be considered as neurotransmitters. These
hormones regulate the synthesis of the other
peptide hormones produced by the pituitary
and are classified to:
1. Releasing hormones
2. Releasing factors
3. Inhibitory factors
The hypothalamic neurohormones are regulated by
higher brain centers through cholenergic, dopamenergic
and GABAergic intervention. Their synthesis is adjusted
by feedback mechanisms from the target organs. These
peptide hormones were isolated and identified and their structure were elucidated. All peptide
hormones posses a very high binding affinity to their receptors.

11. A. Hypothalamic hormones
Thyrotropin releasing hormone (TRH) stimulates the secretion of TSH.
1. Corticotropin releasing factor (CRF, CRH), stimulates ACTH secretions.
3. Growth hormone releasing factor (GHRF) which stimulates GH secretions.
4. Gonadotropin releasing hormones (GnRH, LHRH) which stimulates the secretion of LH
and FSH.
5. Follitropin releasing factors (FRF).
6. Prolactin releasing factors (PRF).
7. Melanoliberin or melanocyte stimulant hormone releasing factor (MRF).
8. Leutropin releasing inhibitor factor (LHRIF).
9. Prolactin releasing inhibitor factors (PRF).
10. Somatostatin (SS), (Somatotropin R inhibitor).
11. Melanostatin, melanocyte stimulant hormone inhibitor factor (MIF).
NB: Some of these hormones are already marketed and show activity in certain disease, e.g.,
cancer, fertility and diagnostic.

12. i. Thyroliberin (Thyrotropic RH) TRH
N
O N
H O
O
N
CONH2
H
N NH Histidyl
Pyroglutamyl
Prolineamide
HOOC COOH
NH2
H2O
N
O COOH
H
Glutamic acid Pyroglutamic
It is tripeptide; consist of L-pyroglutamyl-L-histidyl-L-prolineaide (pyroglu-his-pro-NH2)
H2N
COOH
N NH
Histidine
H N
CONH2
Proline amide
N
O COOH
H
Pyroglutamic

13. Function Hypothalamus
TRH Pituitary
Thyrotropin Thyroid
T3 and T4
SAR
1. Methylhistidyl derivatives is 8 times more active.
2. N-π methylation of imidazole give inactive compound.
3. Pro-pyroglutamyl derivatives is 10 time more active.
N
O COOH
H
N
O N
H O
O
N
CONH2
H
N NH

14. ii. Gonadoliberin (Gonadotropin RH, GnRH)
It known as leuteinizing hormone-follicle stimulating hormone releasing hormone (LH, FSH, RH).
It controls the release of both LH and FSH from pituitary gland. It used clinically for treatment
of hypogonadotrophic infertility in male or female. It is a linear decapeptide.
Pyro-glu-his-trp-ser-tyr-gly-leu-arg-pro-gly-NH2
SAR
A. Agonist of higher activity and longer duration
1. Omission of gly10 and protection of pro9-COOH.
2. Replacement of gly6 by D-amino acid (ala, cyc, phe)
3. Combine both 1 and 2 (D-ser6-des-gly10) 140 more active.
B. Antagonist (used as contraceptives)
1. Omission of his2 (des-his2GnRH)
2. Replacement of his2 by D-amino acid
3. (D-pyro-glu1-D-ph2-D-tyr3,6) GnRH is a complete inhibitor of ovulation in rats.
Synthetic analogs
A. Agonist
1. Buserelin acetate: the gly6 replaced by D-ser6, it used foe treatment of prostate cancer,
advanced breast cancer and infertility.

15. 2. Goserelin acetate (Zoladex)
a. gly6 converted to D-trt-but-ser6 b. Des-gly10 and c. pro9-NH-NH-CO-NH2
It used for treatment of advanced prostate carcinoma and pre-menopausal breast cancer.
3. Leuprolide acetate
a. D-Leu6 b. N-Ethylprolineamide9 c. Glycineamide10
Used for prostate carcinoma.
B. Antagonists
They contain several D amino acids and are used as contraceptives.
e.g. Ganirelix, Antide, Citrorelix,
iii. Somatostatin (SS)
It isolated from hypothalamus, stomach and pancreas. It affect the release of growth hormone,
insulin, glucagon, gastrin, pepsin, secretin and vasoactive intestinal peptides.
Structure
It composed from tetradecapeptide with 3-14 disulfide bond. It have very short duration of action
(t1/2 2-4 min). It is inactivated by enzymatic cleavage of trp8-lys9.
ala-gly-cys-lys-asn-phe-phe-trp-lys-thr-phe-thr-ser-cys

16. Structural modifications
1. gly2 to ala2, asp5 to ala5, trp8 to D-trp8, this modification lead to enhancement of potency and
increase duration of action.
2. phe6,7,11, trp8 to tyr6,7,8,11, give more active compound.
3. Opining the disulfide bridge to 2SH dos not cause loss of activity because it convert to
sulfide bridge by metabolism.
4. Omission of ala1-gly2 tail and replacement of ser13 with D-ser increase specificity to
insulin release.
5. Cys14 to D-cys14 increase the glucagon activity.
Somatostatin analogs
Sandostatin
It is ac octapeptide synthetic analog of somatostatin (D-trp4) has been prepared to resist the
cleavage by endopeptidase, it characterized by very high potency and long duration.
Clinical use
1. Treatment of acromegaly through inhibition of GH secretion.
2. Diabetes through suppression of glucagon release.
3. Bleeding ulcer through inhibition of gastric release.
D-phe-cys-phe-D-trp-lys-thr-cys-thr-OH

17. B. i. Anterior pituitary hormones
1. Corticotropin (ACTH)
This hormone stimulates the adrenal cortex to convert cholesterol to steroidal hormones.
It posses 39 amino acid residues arranged in a single chain in a highly flexible random coil
form in solution.
The SAR
1. Sequential removal of the amino acids from carboxyl terminal from 1 to 20 amino acids
does not cause loss of activity.
2. The 5 amino terminal are essential for activity.
3. The core from 6 to 13 are essential for activity.
4. Amidation of terminal COOH, D-ser or D-ala at NH2 terminal decrease enzymatic
degradation so increase the duration of action.
Synthetic analogs
1. Tetracosactin acetate (hexa acetate)
It is an synthetic polypeptide with sequences identical to the 1st 24 amino acids of ACTH,
contain 76% peptide, 13% acetic acid and from 5-16% water.
It used to test the adrenal cortex.
2. Codactide
It composed of octadecapeptide with (D-ser1, ser17,18) corticotropin. It resists degradation by
aminopeptidase.

18. B. ii. Posterior pituitary hormones
They are synthesized by hypothalamus and transported to the posterior lobe of pituitary gland
where they are stored in the protein bound form. These hormones are released by the pituitary
gland in response to reflex CNS action.
a. Dilatation of the birth canal stimulates oxytocin release, which induce labor.
b. Increase of the osmotic pressure of blood stimulates vasopressin (Anti-diuretic hormone,
ADH) release. Vasopressin is used for treatment of diabetes insipidus in emergencies. Both
oxytocin and vasopressin are nonapeptides with 1-6 disulfide bridge. Vasopressin differs from
oxytocin in having a phenyl alanine residue at position 3 and argenine residue at position 8
instead of the corresponding isoleucine and leucine.
NH2
cys-tyr- lle
S
S
cys-asn-gln
pro-leu-gly-NH2
Oxytocin
NH2
cys-tyr- phe
S
S
cys-asn-gln
pro-arg-gly-NH2
Vasopressin (ADH)

19. Preparation
Pitressin: It is sterile water soluble preparation of vasopressin.
Vasopressin tannate: Water insoluble preparation with long duration of action for IM injection.
felypressin: (2L-phe-8L-lys vasopressin) it posses low antidiuretic activity with higher
vasoconstrictor activity.
Lypressin: (8-Lys vasopressin) it is more stable more rapidly absorbed with the same potency
of vasopressin.
Desmopressin: 1-deamino-8-D-argenine ADH of longer duration, devoid of side effects and
can be used IM and IV.