Hormone Biosynthesis Flashcards
Rate limiting steps in:
Steroidogenesis?
Prostaglandin production?
Melatonin synthesis?
FSH/LH/TSH synthesis?
Estrogen synthesis?
Steroid biosynthesis – StAR enzyme transport of cholesterol form the outer to the inner mitochondrial membrane
Prostaglandin formation – release of free arachidonic acid
Melatonin synthesis – increase in cAMP leading to N-acetyltransferase activity
FSH/LH/TSH synthesis – availability of β subunits
Estrogen synthesis – aromatase activity
Responsible for cholesterol transport across mitochondrial membrane
STAR
Congenital lipoid adrenal hyperplasia (mutation, location, inheritance, functional change)
Loss of function mutation in STAR, chromosome 8, AR, limited intracellular transport of cholesterol 000? intracellular lipid accumulation 00> cellular destruction
Locations of steroid synthesis
Mitochondrial membrane, cytoplasm, endoplasmic reticulum
Locations of steroid receptor
Nucleus, cytoplasm
Mechanism of steroid hormone transport across cell membrane
Simple diffusion
Characteristics of steroid hormones
Small, non-polar, lipophilic
G-protein receptors
cAMP, calcium messenger, protein kinase/MAP kinase
cAMP second messenger hormones
Which G subunit?
FSH, LH, HCG, ACTH, TSH, CRH
Gs-alpha
Calcium second messenger hormones
Which G subunit?
GnRH, TRH, GHRH, kisspeptin, oxytocin, vasopressin, AGII
Gq-alpha
Protein kinase/MAP kinase hormone
Oxytocin (here also, through PKC)
cAMP receptor function
o Hormone binds cell membrane receptor
o Adenylate cyclase activated
o Gα-GTP subunit binds catalytic unit forming active enzyme converting ATP to cAMP
o Forms cAMP-receptor protein complex which activates protein kinase A (PKA)
o Inactive form: tetramer, 2 regulatory subunits and 2 catalytic subunits
o Bound: Catalytic units released, regulatory units form dimer
o Catalytic units phosphorylate serine and threonine residues of cellular proteins (enzymes and mitochondrial, microsomal, and chromatin proteins) (energy-producing)
o Physiologic effect
o Enzyme activity terminated by hydrolysis of GTP to GDP returning the enzyme to its inactive state
Calcium messenger receptor function
o Phospholipase C (PLC) catalyzes hydrolysis of polyphosphatidylinositols (IPI2) into two intracellular messengers: IP3 (inositol triphosphate) and DAG (diacylglycerol)
o IP3 binds with a receptor in the smooth ER and mitochondria and opens the Ca2+ channel
o DAG activates protein kinase C
o Calmodulin binding Ca2+ causes a conformational change
o Modifies calcium transport, enzyme activity, calcium regulation of cyclic nucleotide and glycogen metabolism and secretion and cell motility
Single transmembrane domain receptor types
Tyrosine kinase, cytokine, serine/threonine kinase
Tyrosine kinase receptor hormones
Insulin, IGF, EGF, PDGF, FGF
Cytokine receptor hormones
GH, PRL, hPL, leptin
Serine/threonine kinase ligands?
& what do ligands act by?
Activin, inhibin
- Activin acts via Type I and Type II serine/threonine specific receptor kinases
- Inhibin acts by blocking the Type II serine/threonine specific receptor kinase
Tyrosine kinase receptor function
o 3 domains: extracellular domain for ligand binding, single transmembrane domain, cytoplasmic domain
o Receptor has 2 alpha and 2 beta subunits (each w/ 3 domains as above) linked by disulfide bridge
o Ligand specificity determined by unique AA sequence that determines 3D conformation
o Ligand binding –> conformational change of cytoplasmic domain –> autophosphorylation
Cytokine receptor second messenger
JAK-STAT
Serine/threonine kinase receptor second messenger
SMAD4 –> FOXH1
A/B Regulatory Domain
Amino acid terminal
Most variable in superfamily (i.e. only 18% homology between ERα and ERβ)
In ER-α contains TAF1 which can stimulate transcription in absence of hormone binding
C DNA Binding Domain
Most homologous
Hormone binding induces conformational change in the 3 helices allowing binding to HRE (hormone responsive elements) of target genes
Contains 2 zinc fingers: determine specificity for binding to enhancer site in gene promoter
D HInge REgion
Contains nuclear localization signal
E Hormone Binding Domain
Harbors TAF2 which requires hormone binding for full activity
Functions:
o Pocket for hormone binding
o Sites for cofactor binding
o Responsible for dimerization
o Harbors TAF-2
o Binding site for HSP (when no hormone bound)
F Carboxy Terminal
no notes
TAF1 location/function
(A/B, regulatory domain) can stimulate transcription in the absence of hormone when fused to DNA
TAF2 location/function
(E, hormone binding domain) must have hormone binding for full activity
TAF3 location/function
(B-upstream segment [BUS]) autonomously activates transcription OR synergizes w/ other TAFs
TAFs on ER’s
ERα has TAF1 and TAF2
ERβ only has TAF2
TAFs on PR’s
PRα has TAF1 and TAF2
PRβ has TAF1, TAF2, and TAF3
Steroid hormones with nuclear receptors
Estrogen, thyroid (alpha-chrom 17, beta-chrom3), retinol, vit D
maybe progesterone & androgens?
Steroid hormones with cytoplasmic receptors
(still called nuclear receptors because the action is done there)
Mineralocorticoids, glucocorticoids
ER receptor expression during menstrual cycle
Peak late proliferative (self induced by E2), declines in early secretory (increased P4), increased in mid- and late-secretory (decreased P4)
ER alpha dominant tissues
Uterus/cervix/vagina, ovarian STROMA, breast, bone, hypothalamus, pituitary, kidney / adrenal (E2 and EE most sensitive)
ER beta dominant tissues
Granulosa cells, brain, colon, bladder, prostate
ER alpha and beta tissues
ovary, breast (alpha in dev and fxn, beta as natural suppressor of alpha activity)
2 SERMS and MOA
Raloxifene, tamoxifen; competitive inhibitor of estrogen binding to ER
Tamoxifen SEs
VTE, vaginal bleeding, endometrial hyperplasia/cancer, hot flashes, cataracts
Raloxifene SEs
Hot flashes, vaginal dryness, VTE (less than tamoxifen), decreases LDL, increase HDL, no change in TG
Raloxifene location of estrogenic and antiestrogenic effect
Estrogenic effect on bone
Antiestrogenic effects on breast and uterus
Tamoxifen location of estrogenic and antiestrogenic effect
Estrogenic: liver (decrease AT3, total chol, LDL, increase binding globulins, stimulates P4-R synthesis), bone, vaginal mucosa, endometrium
Antiestrogenic: cystostatic at breast, cytotoxic with breast ca
PR-alpha action and TAFs
Negative action (inhibits activity of PR-B) TAF1 on reg domain, TAF2 on hormone binding domain
PR-beta action and TAFs
Positive reg of progesterone response genes
TAF1 on reg domain, TAF2 on hormone binding domain, TAF3 in B-upstream segment (BUS) at 5’ terminal
PR receptor expression
Induced by E2, inhibited by P4
Peak late-proliferative phase, nearly undetectable by midpoint of secretory phase
First gen progestins
Estranes derived form testosterone
Pregnanes derived from 17-OHP
Second gen progestins
Gonanes derived from testosterone
Third gen progestins
Gonane (levonorgestrel) derivatives
Fourth gen progestins
Non-ethylated estranes
Estranes
Norethindrone (MIcronor) Niorethindrone acetate (Loestrin) Ethynodiol diacetate (Kelnor)
Pregnanes
MPA, Megestrol acetate (Megace)
Gonanes
Levonorgestrel (Seasonale)
Norgestrel (Ovral)
Gonane derivatives (3rd gen)
Gestodene
Desogestrel (Kariva)
Norgestimate (OrthoCyclen)
Etenogestrel (NuvaRing, Implanon)
Non-ethylated estranes
Drosperinone: progestin analogue of spironolactone; high affinity for the MC receptor and anti-androgenic activity
Relative progestin potency
Levonorgestrel
Norgestimate
Desogestrel
NE
NEA
Drosperinone
MPA & Megace
*Most potent on primed endometrium = levonorgestrel
Component that determines efficacy of OCPs
Progestin dose (inhibits LH surge, thickens mucus)
Emergency contraception options (4)
Levonorgestrel: 0.75mg given twice 12h apart or in combined single dose (1.5mg)
Mifepristone: 600mg single dose, same efficacy/SE as levonorgestrel
Ullipristal: 30mg single dose, slightly more effective and similar SE to levonorgestrel
ParaGard: up to 5-10d after unprotected intercourse, failure rate as low as 0.1%
Aromatase inhibitors SEs
- Lower risk VTE, uterine ca, hot flashes and bleeding than tamoxifen
- Higher risk fracture and joint pain/stiffness, vaginal dryness
Androgen receptor forms
A (shorter) and B (full length)
Androgen receptor chromosome location
Xq11-12 (only steroid receptor on X chromosome)
Reason for Androgen and Progesterone receptor cross reactivity
Similar steroid binding
Androgen receptor mutation diseases (2)
AIS (similar presentation to 17-B HSD (type 3) deficiency
Kennedy’s disease (X-linked spinobulbar muscular atrophy)
Flutamide
Nonsteroidal androgen receptor antagonist, can cause severe hepatotoxicity; teratogenic
Cyproterone acetate (CPA)
Derivative of? competes with what? similar profile to what drug?
1st gen progestin
17-α-OHP derivative with potent progestational activity that inhibits gonadotropin secretion, competes with DHT for binding to AR and reduces serum LH and ovarian androgen concentrations, shares similar pharmacological profile to RU486 (mifepristone)
Spironolactone
MOA?
Competive or non-competitive? and with what hormone?
Aldosterone and androgen receptor antagonist structurally similar to progestins, competes with DHT for binding to AR and inhibits enzymes involved in androgen biosynthesis, can increase potassium
Finasteride
MOA?
why only limited action in hirsutism treatment?
Inhibits type 2 5α-reductase, enzyme converting T to DHT, only partial inhibitory effect when used for hirsutism because enhanced 5alpha reductase activity involves both type 1 (skin) and type 2 (reproductive tissues) enzymes
Danazol
isoxazol derivative of 17α-ethinyl testosterone
o Mechanism of action:
• Weak-moderate binding to androgen receptor (also weak binding to PR and ER=least)
• Inhibits the midcycle urinary LH surge and induces a chronic anovulatory state
• Inhibits a number of steroidogenic enzymes (by inhibiting ovarian steroidogenesis)
• Increases free testosterone levels (by displacing from SHBG)
• Decreases serum estrogen levels
o Side effects: weight gain, acne, hirsutism, fluid retention, fatigue, reduced breast size, oily skin, atrophic vaginitis, hot flashes, muscle cramps, emotional lability, can irreversible deepen the voice, unfavorable lipid profile changes (total cholesterol and LDL increased, HDL lowered)
o Associated with virilization of female fetus in utero
o First drug approved for treatment of endometriosis in the United States
DHEA-S source
100% adrenal – [3-20 mg/day]
DHEA source
50% adrenal, 20% ovary, 30% peripheral conversion of DHEA-S – [6-8 mg/day]
Androstenedione source
50% ovary, 50% adrenal – [1.5-6 mg/day]
DHT source
100% peripheral conversion
Androgen potency of androgens
DHT (higher affinity/slower dissociation from AR)>Testosterone>Androstenedione>DHEA-S
Testosterone source
50% from peripheral conversion of androstenedione, 25% adrenal, 25% ovary
Most abundant circulating androgen
Testosterone (DHT is formed intracellularly)
Measuring Peripheral Androgen Activity
3α-androstanediol glucuronide is peripheral metabolite of DHT
Marker of target tissue cellular activity; 3AG correlates w/ 5α reductase activity in skin
BUT, 3AG also reflects hepatic conjugation activity, and impact of precursors (androstenedione and T) derived from adrenal gland – not just from peripheral sources
3AG is not SOLELY a measure of cutaneous androgen metabolism
Insulin/IGF-1 Receptors
Grossly elevated insulin levels stimulate ovarian androgen production in theca cells via insulin, IGF-1 and hybrid receptors
Adrenal zonas
Zona glomerulosa: produces mineralocorticoids
Zona fasciculata: produces glucocorticoids
Zona reticularis: produces sex steroids
Half life hCG
24 hours
Half life FSH
2-4 hours
Half life LH
20 minutes
Half life GnRH
2-3 minutes
SHBG
How many binding sites? What is necessary before binding of ligand?
Contains single binding site for androgens and estrogens (even though homodimer composed of two monomers)
Dimerization is believed necessary to form single steroid binding site
SHBG increased by…
Estrogen, pregnancy, hyperthyroidism, liver disease, anorexia
SHBG decreased by…
Androgens
corticosteroids/anabolic steroids
GH
insulin, IGF1, obesity,
menopause
HYPOthyroidism
Cushing’s disease
prolactin
SHBG affinity of hormones (% bound)
o Testosterone (70) o Estrogen (70) o DHT (30) o DHEA (8) o Androstenedione (8) o Progesterone (1)
Cholesterol Metabolism (exogenous & endogenous)
- Exogenous pathway
i. Lipids absorbed in intestine via microsomal triglyceride transfer protein (MTP) and form chylomicrons (triglycerides)
ii. Chylomicrons carried in bloodstream and broken down by lipoprotein lipase (LPL) into fatty acids and glycerol in peripheral tissues (also makes empty HDL)
iii. Chylo-remnants enter liver via apoE and low density lipoprotein receptor (LDLR) binding - Endogenous pathway
i. Newly synthesized triglycerides repackaged into VLDL
ii. VLDL enter bloodstream and release fatty acids and glycerol in peripheral tissues via lipoprotein lipase – becoming IDL + HDL then LDL
iii. IDLs absorbed by the liver and broken down into LDL via hepatic lipase
Main cholesterol carrier in blood: LDL, HDL or VLDL? (and facts of the other two)
- LDL main cholesterol carrier in blood – low protein:cholesterol ratio
- HDL carries cholesterol from tissues to liver for metabolism/excretion (reverse cholesterol transport) – high protein:cholesterol ratio
- VLDL are triglyceride rich particles
What happens if you have an LDL receptor mutation/defect? (and other s/sx)
CVD/atherosclerosis
Other symptoms:
i. Xanthelasma palpebrarum (yellowish patches around eyes)
ii. Arcus senilis corneae (cholesterol in iris)
iii. Tendon xanthoma (lumps in tendons)
Best predictor of heart disease in women?
Low HDL
How are steroids metabolized?
oxidized by cytochrome P450 oxidase enzymes
- P450 enzymes introduce oxygen into the steroid ring
- Byproducts of steroid breakdown then excreted as bile acids
What is special of conjugated estrogens?
What is the “conjugation”?
conjugated to make them hydrophilic
serve as steroid hormone precursors; cannot bind the estrogen receptor as is
- Conjugation groups
o Sulfate – most common
o Glucuronidase - Lengthens the ½-life of the compound
o Allows transport via circulation
o Increases solubility in bile
Most abundant conjugated estrogen?
Estrone sulfate
Which Progestin generation has:
low androgenic effects
increased risk of DVT/PE
little effect on lipids and glucose
3rd
Which Progestin generation has:
a diuretic effect
4th
Androgenic potency of progestins (highest to lowest)
i. Levonorgestrel
ii. Norgestrel
iii. Desogestrel
iv. Norgestimate
v. Norethindrone acetate
vi. Norethindrone
vii. Ethynodiol diacetate
viii. Drospirenone
ix. Medroxyprogesterone
easier: 2nd > 1st = 3rd > 4th
Each progestin generation’s effect on cholesterols
1st gen GOOD: Increase HDL, decrease LDL, no change/increase TG
2nd gen BAD: Decrease HDL, increase LDL, increase TG
3rd gen: No change/neutral
4th gen GOOD: Increase HDL, decrease LDL, no change/increase (same as 1st)
Inhibin
Production: increased by?
Secretion:
what increases it?
what decreases it?
FSH stimulates inhibin production from granulosa cells → suppresses FSH
Secretion increased by IGF-1
Secretion decreased by GnRH
Inhibin:
Actions (2)
how?
what are low & high levels associated w/?
- Down-regulates FSH synthesis and inhibits FSH secretion
- Enhances LH stimulation of androgen synthesis in the theca to provide substrate for aromatization to E2 in the granulosa
Acts by blocking the Type II serine/threonine specific receptor kinase
Low = DOR
High = granulosa cell tumors
Epidermal growth factor
structural analog?
function?
TGF-alpha - can bind to EGF receptors
decreases FSH induction of LH receptors on granulosa cell (opposite the TGF-b superfamily)
TGF-b action on granulosa cells & theca cells
Granulosa: enhances FSH induction of LH receptors (positive)
Theca: negative action, inhibiting androgen production
Types of nuclear steroid receptors
Type I - reside in cytoplasm, then move to nucleus (mineralcorticoid, glucocorticoid receptors)
Type II - reside in nucleus (Estrogens (most), progestins, and androgens (most) receptors, thyroid hormone receptor, Vit D)
Most common HSPs in E region of steroid receptors that need to be dissociated before moving to nucleus
hsp 90 & 56
Zinc Fingers:
- # of AAs? most important AA?
- Essential for what?
- Where is functional specificity localized?
- 100 AA’s w/ 9 cysteines in fixed positions
- activation of transcription
- second zinc finger in an area called the d(distal) box
DNA sequences that are bound by steroid receptors called what?
Where is it in the gene? what is common about it?
steroid/hormone response element
Part of the promoter region of the gene
- most commonly a pair of inverted repeats separated by three nucleotides, which also indicates that the receptor binds as a dimer.
What aspect of binding is just as important as the dose?
give example (2)
Duration of exposure
- only small amounts of estrogen needed in circulation b/c long half-life of estrogen hormone-receptor complex. (Potency differences among the various estrogens related to length of time estrogen-receptor complex occupies the nucleus)
- Cortisol and progesterone must circulate in large concentrations because their receptor complexes have short half-lives
What do IP3 & DAG activate?
- IP3 – activates calcium ion channels
- DAG – activates protein kinase C
Mifepristone MOA
- Progesterone receptor antagonist (affinity 5x greater than natural hormone)
- Glucocorticoid receptor antagonist (but the circulating level of cortisol is so high, so doesn’t produce as much an effect)
Positive & Negative feedback of estrogen
Negative: Occurs during follicular phase when E2 levels are low
Positive: Occurs at high E2 concentrations near the end of the follicular phase – E2 becomes a positive inducer of the anterior pituitary - triggers the anterior pituitary to release more FSH/LH to produce more E2 – resulting in a substantial rise that triggers the LH surge
Progesterone receptor-A and B knock-out - what happens to:
Breast? Endometrium? Ovary?
- Receptor-A knock-out (PR-A opposes E2-induced proliferation, required for fertility/ovulation)
- Ovary - anovulation
- Endometrium – epithelial cell proliferation in response to E2/P4 (because ONLY PR-B expression present)
- Breast – no change in response of mammary gland to E2/P4
- Progesterone Receptor-B knock-out (PR-B enhances E2-induced proliferation, required for mammary gland function)
- Ovary – no change in response of ovary to E2/P4
- Endometrium – no change in response of endometrium to E2/P4
- Breast – Reduced mammary ductal morphogenesis and alveologenesis during pregnancy
Elagolix
MOA?
non-peptide oral, short-acting competitive GnRH receptor antagonist
Medroxyprogesterone Acetate risk in:
VTE / effect on lipids?
breast cancer?
CHD?
what is oral brand name?
all bad. the worst progestin when combined w/ estrogen
Provera
Micronized Progesterone risk in:
VTE / effect on lipids?
breast cancer?
CHD?
what is brand name?
metabolically neutral, doesn’t seem to have effect.
Prometrium
