Molecular and Cellular Mechanism of Action of Thyroid_Anas_Saifi.pptx

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1. Molecular and Cellular Mechanism of Action of Thyroid Hormone Anas Saifi M.Pharm (Pharmacology) 2 nd Sem. School of Pharmaceutical Education And Research Jamia hamdard 1

2. Presentation Outline Introduction Chemistry of thyroid hormones Biosynthesis Transport, metabolism and excretion Thyroid hormone receptors Regulation of Secretion Relation between T 4 and T 3 Mechanism of action Actions of thyroid hormones Clinical applications Anti-thyroid drugs Summary References 2

3. Introduction Thyroid gland secretes 3 hormones- Thyroxine (T 4 ), Triiodothyronine (T 3 ) Calcitonin. T 3 and T 4 are produced by thyroid follicles, having similar biological activity. Calcitonin is produced by interfollicular ‘C’ cells, Calicitonin is chemically and biologically entirely different, regulates calcium metabolism. Fig. 1: Location of thyroid gland 3

4. History The physiological role of thyroid gland was recognized after Graves and Basedow ( 1835, 1840 ) showed the clinical features of the 'Graves' disease and Gull correlated myxoedema with its atrophy (1874). Kendall (1915) obtained crystalline thyroxine and postulated its chemical formula which was confirmed in 1926 . Thyroxine was the first hormone to be synthesized in the laboratory. T 3 was discovered in 1952. 4

5. Chemistry of thyroid hormones Both T 3 and T 4 are iodine containing derivatives of thyronine , is a condensation product of two molecules of the amino acid tyrosine . Thyroxine (T 4 ) is 3, 5, 3', 5'-tetraiodothyronine and T 3 is 3, 5, 3'- triiodothyronine. a) b) c) d) Fig. 2: Chemical structure of a) Tyrosine, b) Thyronine , c) T4 & d) T3 5

6. Biosynthesis Thyroid hormones are synthesized and stored as amino acid residues of thyroglobulin (TG). TG is a complex glycoprotein made up of two apparently identical subunits, each of 330 KDa , containing 10% sugar . The major steps in the synthesis, storage, release, and interconversion are : Iodide uptake Oxidation and Iodination Coupling Storage and release Peripheral conversion of T4 to T3 6

7. Biosynthesis Iodide uptake Iodine is   obtained from food and water Thyroid cells have an active transport process (Na+: I¯ symporter or NIS) to concentrate the anion This trapping is stimulated by TSH to exceed a gradient of more than 100 fold I 2  content of thyroid gland somehow regulates the uptake mechanism. Oxidation and Iodination Iodide trapped by follicular cells is carried across the apical membrane by the transporter ‘ pendrin ’. The trapped iodide is oxidized by the membrane bound thyroid peroxidase enzyme to iodinium (I+) ions or hypoiodous acid (HOI) or enzyme linked hypoiodate (EOI) with the help of H2O2. These forms of iodine combine with tyrosil residues of thyroglobulin and form monoiodotyrosine (MIT) and diiodotyrosine (DIT). 7

8. Biosynthesis Fig. 4: Biosynthetic Pathway of Thyroid Hormones NIS-Na-iodide symporter; HOI- Hypoiodous acid; EOI-Enzyme linked hypoiodate MIT- Monoiodotyrosine ; DIT-Diiodotyrosine; T 3 -Triiodothyronine; T 4 -Thyroxine 8

9. Biosynthesis Coupling Pairs of iodinated tyrosil residues couple together to form T3 and T4.  Normally much more T4 than T3 is formed. Coupling is an oxidative reaction and is catalysed by the same thyroid peroxidase. Oxidation of iodide and coupling are both stimulated by TSH . Fig. 3: Coupling of MIT & DIT to form T3 9

10. Biosynthesis Storage and release Tg containing iodinated tyrosil and thyronil residues is transported to the interior of the follicles. These Tg remains stored as thyroid colloid till it is taken back into the cells by endocytosis and broken down by lysosomal proteases. The T 4  and T 3  so released is secreted into circulation while MIT and DIT residues are de-iodinated and the iodide released is reutilized. Peripheral conversion of T4 to T3 Peripheral tissues convert T 4  to T 3 . The conversion is carried out by iodothyronine deiodinase (D1, D2 & D3) Almost equal amounts of 3, 5, 3´ triiodothyronine (normal T 3  : active) and 3, 3´, 5´ triiodothyronine (reverse T 3 : inactive) are produced in the periphery. 10

11. Transport, Metabolism and Excretion Thyroid hormones are avidly bound to plasma proteins— Thyroxine binding globulin (TBG) Thyroxine binding prealbumin (transthyretin) Albumin Only the free hormone is available for action, metabolism and excretion. Metabolic inactivation of T4 and T3 occurs by deiodination and glucuronide/sulfate conjugation. Liver is the primary site and the conjugates are excreted in bile. A significant fraction is deconjugated in intestines and finally excreted in urine . Plasma t½ : T4 - 6–7 days and T3 -  1–2 days. The half lives are shortened in hyperthyroidism and prolonged in hypothyroidism . 11

12. Thyroid Hormone Receptors Thyroid Hormone Receptors(TR) are nuclear receptors. Have two DNA binding domains, one ligand bindng domain and one transactivation domain. Act as transcription factors, affecting the regulation of transcription and translation. Non-genomic effects lead to second messenger activation and cellular response. Isoforms Common Location of Expression TR- α1 cardiac and skeletal muscles, brown fat and bone TR- α 2 skeletal muscle, brain and kidney TR- α 3 skeletal muscle, brain and kidney TR- β 1 brain, liver and kidney TR- β 2 retina, hypothalamus, anterior pituitary and cochlea TR- β 3 & TR- β 4   THRA   THRB 12

13. Regulation of Secretion Hypothalamic regulation Pituitary regulation Feedback regulation of thyroid secretion Blood iodine level Fig. 5: Regulation of thyroid function SST- Somatostatin; TRH- Thyrotropine releasing hormone; TSH- Thyroid stimulating hormone 13

14. Regulation of Secretion Thyrotrophin -releasing hormone (TRH), released from the hypothalamus in response to various stimuli, releases thyroid-stimulating hormone (TSH; thyrotrophin ) from the anterior pituitary. TSH acts on receptors on the membrane of thyroid follicle cells through a mechanism that involves cAMP and phosphatidylinositol 3-kinase . The production of TSH is also regulated by a negative feedback effect of thyroid hormones on the anterior pituitary gland. The peptide somatostatin also reduces basal TSH release . The control of the secretion of TSH thus depends on a balance between the actions of T3/T4 and TRH on the pituitary. The other main factor influencing thyroid function is the plasma iodide concentration. 14

15. Relation between T4 and T3 T 3  is the active hormone, T 4  is a transport form, functions as a prohormone of T 3 . Thyroid secretes more T 4  than T 3 .   T 4  is the major circulating hormone because it is 15 times more tightly bound to plasma proteins. T 3  is 5 times more potent than T 4  and acts faster. T 3  is more avidly bound to the nuclear receptor than T 4  and the T 4 -receptor complex is unable to activate gene transcription. 15

16. Mechanism of action Both T3 and T4 produce actions by combining with a nuclear thyroid hormone receptor (TR), belongs to steroid and retinoid superfamily of intracellular receptors . TR is bound to the ‘thyroid hormone response element’ (TRE) in the enhancer region of the target genes along with corepressors which keeps gene transcription suppressed. When T3 binds to the ligand binding domain of TR → heterodimerizes with retinoid X receptor (RXR) → conformational change releasing the corepressor and binding the coactivator → gene transcription → production of specific mRNA and a specific pattern of protein synthesis → various metabolic and anatomic effects. Apart from the nuclear T3 receptor, it acts on cell membrane to enhance amino acid and glucose entry and on mitochondria to increase oxygen consumption. 16

17. Mechanism of action Fig. 6: Mechanism of action of thyroid hormone on nuclear thyroid hormone receptor (TR). TRE-Thyroid hormone response element; RXR-Retinoid X receptor; 5 DI-5 Deiodinase 17

18. Non-genomic Effects of Thyroid Hormone TRs can be found outside the nucleus where they can exert biological effects via rapid nongenomic mechanisms. TRs associate in a T3-dependent manner with the p85 subunit of phosphatidyl inositol 3-kinase (PI3K) → phosphorylation and activation of PKB/ Akt → broad effects on cellular metabolism. For example, it stimulates NO production by endothelial cells, which leads to vasodilation. T3 administration causes rapid vasodilation . Non-genomic actions of thyroid hormone via a plasma membrane receptor within integrin α V β3 → binds extracellular T4 in preference to T3, resulting in activation of MAP kinase. 18

19. Actions of thyroid hormones 19

20. Clinical Applications The most important use of thyroid hormone is for replacement therapy in deficiency states. Synthetic l-thyroxine is the preparation of Choice 20

21. Clinical Applications Preparations: l-thyroxine sod. (synthetic levothyroxine sod.): ELTROXEN (25 µg, 50 µg, 100 µg) tabs by GlaxoSmithKline, THYRO NORM (12.5 µg, 25 µg, 50 µg, 62.5 µg, 75 µg, 100 µg, 125 µg, 137 µg, 150 µg) tabs by Abbott, THYROX (25 µg, 50 µg, 75 µg, 100 µg ) tabs by Macleods Pharmaceuticals Ltd. Pharmacokinetics and interactions: Oral bioavailability of l-thyroxine is 75% but severe hypothyroidism can reduce oral absorption. Should be administered in empty stomach to avoid interference by food. Sucralfate, iron, calcium and proton pump inhibitors also reduce l-thyroxine absorption. CYP3A4 inducers like rifampin, phenytoin and carbamazepine increase T 4 metabolism. 21

22. Anti-thyroid drugs A number of compounds are capable of interfering, directly or indirectly, with the synthesis, release, or action of thyroid hormones. Several are of great clinical value for the temporary or extended control of hyperthyroid states. The major inhibitors may be classified into four categories: Anti-thyroid drugs, which interfere directly with the synthesis of thyroid hormones; Ionic inhibitors, which block the iodide transport mechanism ; High concentrations of iodine, which decrease release of thyroid hormones from the gland and also may decrease hormone synthesis; Radioactive iodine, which damages the thyroid gland with ionizing radiation. 22

23. Anti-thyroid drugs CLASSIFICATION 23

24. Mechanism of action Anti-thyroid drugs inhibit the formation of thyroid hormones by interfering with the incorporation of iodine into tyrosyl residues of Tg . Inhibit the coupling of these residues to form iodothyronines. Inhibits the peroxidase enzyme, preventing oxidation of iodide or iodotyrosyl groups. When Graves' disease is treated with anti-thyroid drugs, the concentration of thyroid-stimulating immunoglobulins in the circulation decreases, act as immunosuppressants . 24

25. Site of action Propylthiouracil inhibits step 2 and 6, Thioamides & Carbimazole inhibit step 2, Ionic inhibitors block step 1, Excess iodide interfers with step 1, 2, 3 and 5. 25

26. Thioamides Thioamides bind to the thyroid peroxidase and prevent oxidation of iodide and iodotyrosyl residues, thereby, Inhibit iodination of tyrosine residues in thyroglobulin Inhibit coupling of iodotyrosine residues to form T 3 and T 4 . Propylthiouracil blocks hormone synthesis, partially inhibits the peripheral deiodination of T 4 to T 3 . Preparations and dose: Propylthiouracil : 50-150 mg TDS. Methimazole : 5-10 mg TDS. Carbimazole : 5-15 mg TDS, NEOMERCAZOLE, THYROZOLE, ANTITHYROX 5, 10, 20 mg tab . 26

27. Thioamides Pharmacokinetics: Antithyroid drugs are quickly absorbed orally, widely distributed in the body, enter milk and cross placenta metabolized in liver and excreted in urine primarily as metabolites. Carbimazole acts largely by getting converted to methimazole in the body and is longer acting than propylthiouracil . Adverse effects: Hypothyroidism and goiter - Due to over treatment High dose: causes excess TSH production- enlargement of thyroid gland (Goiter). Side effects: GI intolerance, Skin rashes, joint pain. Graying or loss of hair, loss of taste, fever & liver damage. 27

28. Thioamides Propylthiouracil and Carbimazole : Parameters Propylthiouracil Carbimazole Potency Less potent About 5 × more potent Plasma protein binding 80-90% Most of the drugs are free Crosses placenta Less transferred across placenta & in milk Large amounts cross to foetus and in milk Plasma t 1/2 1-2 hours 6-10 hours Dose Multiple daily doses are needed Single daily dose Active metabolite No Methimazole Peripheral conversion of T 4 to T 3 Inhibits Does not inhibit 28

29. Thioamides Therapeutic uses : As definitive therapy: In some patients, after 1-2 years of treatment remission may occur. After drug withdrawal if symptoms recur again, the treatment is restarted. This situation mostly occurs in patients having short history of Graves’ disease & small goiter. Preoperatively: Surgery (thyroidectomy) is advised to thyrotoxicosis patients, so preoperative treatment of carbimazole is given . Treatment along with I131: When there is prompt control of severe hyperthyroidism in older patient with heart disease, following sequence of therapy is employed; Starting treatment with antithyroid drug 1-2 weeks gap Radioiodine dosing Resume antithyroid drugs after 5-7 days and gradually withdrawal over 3 months. 29

30. Ionic inhibitors They inhibit iodide trapping by NIS into thyroid because of similar hydrated ionic size—T4/T3 cannot be synthesized In large amounts, thiocyanates may inhibit the organification of iodine Thiocyanates : can cause liver, kidney, bone marrow and brain toxicity . Perchlorates: produce rashes, fever, aplastic anaemia , agranulocytosis It causes fatal aplastic anemia when given in excessive amounts (2-3 gm daily ). They are toxic and not clinically used now. 30

31. Iodine and Iodides Iodine acts as fastest acting thyroid inhibitor. Response to iodine and iodide is identical, iodine is reduced to iodide in intestine. Due to negative feedback mechanism iodide inhibits release of thyroid hormone within 1-2 days of starting of treatment. 10-14 days causes marked reduction in vascularity of gland & decrease in the size of gland. After which “ thyroid escape ” occurs . Preparations and dose: Lugol’s solution : orally, 5% iodine in 10% potassium iodide solution. Colloid iodine 10%: 5-10 drops /day Collosol : 8 mg iodine/5 ml liquid. 31

32. Iodine and Iodides Mechanism of action : Actual mechanism is unknown. It inhibits own transport into thyroid cell by acting on NIS. It attenuates TSH & cAMP & causes thyroid inhibition. Excess iodide rapidly interferes with iodination of tyrosil and tyronil residues of Tg , reduces T 4 /T 3 synthesis ( Wolff - Chaikoff effect ). Inhibition of hormone release causes thyroid constipation .   32

33. Iodine and Iodides Therapeutic uses : 1) Preoperative preparation: For thyroidectomy in Graves’ disease, iodine for 10 days before surgery, makes gland less vascular & easier to remove. 2) Thyroid storm: Lugol's iodine (6-10 drops) or iodine containing radio contrast media ( iopanoic acid) are used orally to stop further release and conversion of T4/T3 from the thyroid. 3) Prophylaxis or endemic goiter: It is used as “iodized salt ”. 4) Antiseptic : The tincture of iodine, povidone iodine is used as antiseptic. 33

34. Iodine and Iodides Adverse effects : Acute reaction : occurs in sensitive people. Symptoms like swelling of lips, eyelids, angio -edema of larynx, fever, joint pain, petechial hemorrhages, thrombocytopenia, lymphadenopathy . Chronic overdose ( iodism ) : Inflammation of mucous membranes, salivation, rhinorrhoea , sneezing, lacrimation, swelling of eyelids, burning sensation in mouth, headache, rashes, g.i. symptoms. Long-term use of high doses can cause hypothyroidism & goiter. If high dose given to pregnant women chances of hypothyroidism & goiter in foetus .  34

35. Radioactive Iodine Medically useful isotope is I 131 Half life : 8 days Chemical behaviour is similar to stable isotope I 127 Radioactive iodine administered as sodium salt of I 131 dissolve in water & taken orally.  Diagnostic: 25-100 μ curie is given Scanning is done at interval Use: Thyroid Carcinoma Therapeutic: Therapeutic dose is 3-6 m curie. High dose is required for toxic multinodular goiter. The response to radio active iodine starts after 2 weeks & peak is reached after 3 months, repeat dose is given after that. Uses 35

36. Radioactive Iodine Mechanism of action : I 131 emits both β particles and γ rays Destructive effect on thyroid cells Penetrates only 0.5-2 mm of tissue Pyknosis and necrosis followed by fibrosis Tracer studies Passes through tissues without damaging 36

37. β adrenergic blockers β blockers are used in - While response to carbimazole , propylthiouracil or I 131 is low β blockers given along with iodide for preoperative preparation before subtotal thyroidectomy Thyroid storm or thyrotoxic crisis Propranolol reduces peripheral conversion of T4 to T3 (1-2 mg i.v. or 40-80 mg oral every 6 hours) Diltiazem : 60-120 mg BD oral 37

38. References Brunton, L., Chabner, B. and Knollmann, B., 2011. Goodman & Gilman's the Pharmacological Basis of Therapeutics. 12th ed. New York: McGraw Hill Professional, pp.1249-1280. Tripathi, K., 2019. Essentials of Medical Pharmacology. 8th ed. New Delhi: Jaypee Brothers Medical Publishers, pp.267-278 Ritter, M. et al., 2020 Rang and Dale's Pharmacology. 9th ed. Edinburgh: Elsevier, pp. 450-454. Parry, T., Ledee, D., Willis, M. and Portman, M., 2017. Nuclear Receptors and the Adaptive Response of the Heart. Endocrinology of the Heart in Health and Disease, pp.249-284. Abdi, H., Amouzegar, A. and Azizi, F., 2019. Antithyroid Drugs. Iran J Pharm Res., pp.1-12. 38

39. Thank You 39