Nuclear Receptors Flashcards
1
Q
What are examples of ligands?
A
- Hormones
- Growth factors
- Neurotransmitter sugars
- Lipids
- Ions
2
Q
How do nuclear receptors differ from cell surface receptors?
A
- Cell surface; act as sensors for extracellular molecules
- Nuclear receptors; act as sensors for intracellular levels of small molecules/metabolities, potentially inducing a signal transduction cascade binding to DNA and activating gene transcription
3
Q
What signals do nuclear receptors respond to?
A
(small) Lipid-soluble molecules (entering into the cell, into the nucleus binding in situ):
- hormones
- vitamins
- retinoids
4
Q
What are the outcomes of nuclear receptor activation?
A
- Activating gene transcription
- Repressing gene transcription
5
Q
What forms do nuclear receptors exist in?
A
- Homodimers; 2 molecules of the same NR protein
- Heterodimers; 2 molecules of differing nuclear receptor protein
- Monomers
6
Q
What are orphan receptors?
A
Nuclear receptors that are thought not to have ligands (e.g. interaction w/another protein) or of which their ligands have not yet been discovered.
7
Q
What does ligand binding do to a NR?
A
Induces a conformational change of the nuclear receptor protein e.g. in folding/shape, altering its ability to induce gene transcription switching it from an inactive form to an active form or vice versa.
8
Q
What is a xenobiotic compound?
A
Compounds/foreign chemical substance not normally produced by the organism; e.g. PXR nuclear receptor can detect the xenobiotic and respond accordingly, drugs pollutants etc.
9
Q
What do natural NR ligands all have in common?
A
- Lipophilic
- Lipid solubility enables them to cross cell membranes with ease by passive diffusion
10
Q
What are the different classes of natural NR ligands?
A
- Sex steroids (cholesterol derivatives)
- Reinoids; Vitamin A
- Prostaglandins/fatty acids
- Vitamin D
- Thyroid hormone
11
Q
Why do ligands have different affinities for their respective NRs?
A
- Steroids, vitamins etc. have high affinity for their NR as there is a lower circulating concentration
- Than metabolites etc. such as fatty acids where there is lots of availible
12
Q
What effects can ligands have on their NRs?
A
- Agonists
- Partial agonist
- Antagonist
13
Q
What processes do NRs regulate in human physiology?
A
- Cell growth, proliferation, apoptosis (death), homeostasis
- Tissue differentiation
- Development
- Metabolism
- Endocrine systems
- Body clock/circadian rhythms
- Reproduction
- CNS function
- Cardiovascular system
- Respiratory system
- Renal function
- GI function
- Immune system
- Stem cell renewal
14
Q
When are NRs important clinical targets?
A
- Inflammatory disease
- Solid tumours
- Leukaemias and lymphomas
- Developmental abnormalities
- Metabolic diseases
- Growth disorders
- Diabetes
- Kidney disease
- Obesity
- Liver disease
- Endocrine disorders
- Auto-immune disease
- Retinopathies
- Thyroid disorders
- Behavioral disorders
- Alzheimer’s disease
- Parkinson’s disease
- Stroke
- CVD
- Hypogonadism
- Polycystic ovary disease
15
Q
What are the top NR drug targets?
A
- Estrogen receptor
- Glucocorticoid receptor
- Progesterone receptor
- PPARs
16
Q
What is the NR domain structure?
A
- 2 main functional domains
- DNA Binding Domain (DBD)
- Ligand Binding Domain (LBD)
17
Q
What enables the DBD of a NR to specifically recognise sequences?
A
- 2 Zinc Fingers
- 2-3 alpha helices arranged to read dsDNA
- Helices interact w/bases and read sequences, allowing receptor to recognise specific sequences
18
Q
What is the structure of the LBD and what does it contain?
A
- Makes up most of the C-terminus
- 10-12 alpha helices; NRs have conserved folding but differed enough in sequence to give specificity
- Contains activation helix
- Many contain a hydrophobic cavity known as the ligand binding pocket
- Ligand binding changes conformation of LBD
19
Q
What are the functions of the DBD NR Domain?
A
- Binds specific DNA sequence; normally 6 base pairs long
- Normally AGGTCA
- Or steroid receptors bind AGAACA
- Dimer nature; recognise 2 copies of the sequence
- Dimerisation (on the DNA)
20
Q
What are the functions of the LBD NR Domain?
A
- Ligand binding
- DImerisation
- Co-factor binding (needed for transcriptional activity)
- Transcriptional activation or repression
21
Q
Upon ligand-binding, how does the NR become repressed/activated?
A
- NRs recruit respective co-factors; for an antagonist ligand/no ligand, co-repressor proteins are recruited which in turn represses gene transcription
- With an agonist ligand, co-activator proteins are recruited which allow for transcription of the desired gene (facilitating RNA polymerase etc)
22
Q
How do recruited co-factors interact with NRs?
A
- Via conserved helical motifs, promoting NR/cofactor complexes
- Ligand binding to NR = conformational change which accommodates binding to helical motifs of cofactors
- Strong protein-protein interaction occurs
- Co-activators: LXXLL motif (3 leucines + 2 other AAs)
- Co-repressors: LXXXI/LXXXL (1 leucine 1 isoleucine + 3 AAs or 2 leucines and 3 AAs)
- They form amphipathic alpha helices with respective hydrophobic and -philic ends, leucines pointing in the same direction
23
Q
How do corepressors exert their suppressive effect?
A
- DNA is packaged as chromatins; nucleosomes of 8 histone proteins w/200 bases of DNA wrapped around
- Corepressors contain histone deacetylases
- Enzyme removes acetyl groups from histones, which keep nucleosomes tightly packed and generates a compact chromatin structure
- This blocks the recruitment of polymerases; transcription does not occur
24
Q
How do coactivators exert their agonist activity?
A
- DNA is packaged as chromatins; nucleosomes of 8 histone proteins w/200 bases of DNA wrapped around
- Coactivators contain histone acetylases
- These add acetyl groups to the histones; this occurs on the lysine residue which is positive - DNA is negative (normally enhancing the interaction), thus acetylating removes the positive charge relaxing interaction
- This generates an open chromatin structure, making it easier to invade thus there is recruitment of polymerases and transcription of the gene can occur
25
What is different about steroid receptor dimerisation at the LBD?
- Most NR LBDs function as dimers
- Steroid receptors function as homodimers, binding inverted repeats (palindromes; reading the same forwards and backwards)
26
How does LBD heterodimer formation contribute to greater complexity of NR function?
- Ligand-binding NRs (but not steroids) form heterodimers with RXR (retinoid X receptor; which binds 9-cis retinoic acid)
- They bind direct repeats of AGGTCA (unlike palindromic nature of steroids)
- But spacing between repeats 'n' differs giving rise to unique functionalities etc
27
How does the activation helix influence cofactor binding?
- Activation helix normally is chillin' and protrudes away when inactivated
- Ligand binding at ligand binding pocket of LBD brings about conformational change due to molecular interactions with AAs etc
- Activation helix moves (a lot)
- This opens up a surface on the NR for the LXXLL motif of a coactivator (e.g.) to dock and bind
- Intimate hydrophobic interaction established with leucine and ligand binding domain
28
What is the activation helix also known as?
- Helix 12
| - AF2`
29
What are the different gene mutations that can result in human disease?
- Mutation
- Deletion
- Amplification
30
What diseases can arise from PPAR Mutations?
- Type II Diabetes
- Insulin resistance
- Obesity
- Dyslipidemia
31
What diseases can arise from thyroid hormone receptor mutations?
- Thyroid disorders
| - Graves disease
32
What diseases can arise from mineralocorticoid receptor mutations?
- Severe familial hypertension
33
What diseases can arise from Retinoic Acid Receptor mutation?
- Gene fusion PML-RAR in acute promyelocytic leukaemia (100% remission achieved by treatment with All Trans Retinoic Acid
34
What diseases can arise from photoreceptor specific nuclear receptor mutations?
- Retinopathies
| - Retinitis pigmentosa
35
What diseases can arise from estrogen receptor mutation?
- Metastatic breast cancer
36
What diseases can arise from androgen receptor mutation?
- Metastatic prostate cancer
37
What are endocrine cancers?
- Tumours arising in endocrine tissues (hormone producing)
- Tumours of adrenals, pancreas, pituitary, thyroid, parathyroid glands
- Poor outlook
38
What are steroid hormone-related cancers?
- Tumours of breast, prostate, ovary, endometrium, testis, osteosarcoma
- Growth of tumour stimulated by sex (steroid) hormones
- Treatable w/hormone therapies
39
What are the potential origins of breast cancer?
- Ductal
- Lobular
- Mucinous
- Basal-type
- Medullary
40
What is the difference between benign and malignant breast cancer?
- Benign; localised - DCIS (Ductal Carcinoma In Situ; contained within duct
- Malignant; invasive - metastatic, spreading outside local origin
41
How is breast cancer classified?
- Tumour morphology
- Biomarker staining (staining w/antibodies to determine if receptors present and genetic factors): ER (estrogen receptor), PR (progesterone receptor), EGFR2 (HER2), BRCA1, BRCA2
42
What gene transcription signatures does breast cancer possess?
- Luminal A
- Luminal B
- Basal
- HER2-enriched
43
When would breast cancer be treated with hormone therapies?
- If they are ER+; estrogen receptor expressing
44
When is herceptin used for breast cancer therapy?
- If the patient is ER-, but HER2 (human epidermal growth factor receptor 2) enriched
- No point using hormone therapy as ER-
45
What is the outlook for a triple negative breast tumour?
- Don't express ER
- Don't express HER2
- No approved targeted therapy; poor outcome
46
What is available for breast cancer treatments?
- Surgery
- Radiotherapy (often following surgery)
- Chemotherapy (pre and post surgery, shrinking tumour to cause minimal damage etc)
- Hormone therapy
- Targeted therapy
47
What surgery is available for breast cancer treatment?
- Lumpectomy,
- Mastectomy - whole breast,
- Lymph node biopsies
48
What chemotherapy regimens are available for breast cancer treatment?
- 5-FU,
- epirubicin,
- docetaxel,
- cyclophosphamide,
- methotrexate
49
What hormone therapies are available for breast cancer treatment and what do they entail?
- Antiestrogen (Tamoxifen); binds estrogen receptor as antagonist ligand
- Aromatase inhibitors (Arimidex; Aromasin; Femara); block estrogen synthesis, so that it's not available to stimulate cancer cell growth
- Ovarian ablation (Zoladex); GnRH agonists (gonadotropin releasing hormone agonist, blocking the function of the ovaries - where the estrogen is produced)
50
What targeted therapy is available for breast cancer treatment?
- HER2+; herceptin (trastuzumab); monoclonal antibody
51
How do tamoxifen and raloxifen exert their therapeutic activity?
- Growth of many breast tumours is estrogen (estradiol)-dependent
- Tamoxifen/raloxifen (SERMs - Selective Estrogen Modulators) are antagonists that block estrogen-dependent growth, mimicing estradiol (estrogen) and binding reversibly to ER LBD
- Bulky nature pushes activation helix out of position; occupying position where cofactors normally bind
- Co-activators are thus unable to form a complex with ER
52
What are the other tissue-specific effects for the hormone therapies tamoxifen/raloxifen?
Tamoxifen/Raloxifen:
- are ER antagonists in breast tissues
- are partial ER agonists in bone (prevent osteoporosis - menopausal women lose bone density due to lack of estrogen productio)
Tamoxifen:
- ER partial agonist in endometrium ( inner mucous membrane of the mammalian uterus); increased risk of endometrial cancer over time
Raloxifen:
- increased risk of blood clots
53
How do aromatase inhibitors exert their therapeutic effect?
- Estrogen synthesis dependent on CYP19A aromatase
- CYP19A converts testosterone to estradiol, and estrone to estradiol
- Inhibiting this enzyme inhibits the synthesis of estrogen; tumour is starved
54
What is the normal function of the prostate?
- Sits just under bladder
- Exocrine gland
- Secretes alkaline fluid; part of ejaculate/sperm
- Protectant for sperm
55
What are the characteristics of prostate cancer?
- May be symptomless; it's a slow-growing cancer
- Urinary problems; pressing on bladder etc
- Pelvic pain (in more advanced carcinoma)
- Stage I - IV (localised - invasive)
56
How is prostate cancer diagnosed?
- Digital rectal exam
- PSA test
- MRI or CT scan
- Biopsy; ultrasound-guided
57
What does the Gleason Score entail and how does it help diagnose prostate cancer?
- Helps clinician decide how advanced tumour is
- 10-12 samples scored
- Most common grade + highest grade:
1 = small, uniform glands (well differentiated)
2 = more stroma between glands
(moderately differentiated)
3 = Distinctly infiltrative margins
(poorly differentiated/anaplastic)
4 = Irregular masses of neoplastic glands
5 = Only occasional gland formation
- 1 + 2 = not cancerous
- 3 + 4 = intermediate grade
- 5 + 5 = most aggressive
58
What does the PSA test entail for prostate cancer diagnosis?
Blood test for prostate specific antigen (Pca biomarker):
| - 20ng/ml; high risk
59
What are androgens?
Male sex hormones:
- testosterone
- 5'-dihydrotesterone (DHT)
60
How are androgens linked to prostate tumours?
- Androgens mostly produced by testes (90%), but small amount (10%) produced by adrenal glands
- Required for normal prostate growth & function BUT also stimulates prostate cancer cell growth
1. ) Hypothalamus secreted GnRH/LHRH (gonadotropin releasing hormone)
2. ) GnRH stimulates pituitary to secreted Luteinizing Hormone (LH)
3. ) LH controls testosterone production in testes
4. ) Testosterone converted to more active DHT in prostate
5. ) DHT binds and activates androgen receptors which regulate gene expression
61
What are the treatment for prostate cancer?
- Active surveillance; watchful waiting (intervention postponed until/unless necessary)
- Radical prostatectomy; if tumour is localised
- Radiotherapy/chemotherapy (docetaxel - taxotere)
- Seed brachytherapy; radioactive seed implants in the prostate
- Cryotherapy; kill tumour cells by localised freezing
- HIFU; High Intensity Focussed Ultrasound, killing tumour cells with heat
- Antihormone therapy
- Androgen deprivation therapy
62
What are the potential side effect of antihormone/androgen deprivation therapy?
- Erectile dysfunction, loss of libido
- Weight gain, muscle loss, bone thinning, breast swelling, hair loss
- Memory, mood, concentration effects
63
What androgen deprivation therapies are available for prostate cancer?
- GnRH Agonists
- GnRH antagonists (medical castration); pituitary
- CYP17A inhibition; block androgen synthesis
- Antiandrogens; AR antagonists (at receptor level)
- Bilateral orchidectomy (surgical castration)
64
How can castration resistant prostate cancer develop?
Resistance to hormone therapies emerges after 1-2 years Androgen Deprivation Therapy treatment
65
What differs in castration resistant prostate cancer to emerging prostate cancer?
- Advanced metastatic disease
- Incurable; no effective treatment
- 60% of tumours have mutations/amplifications in AR gene
- LBD mutations common
- Mutant AR activated by antagonists such as flutamide, bicalutamide; disease worsens with previous ADT treatment driving cancer growth
- Mutant AR no longer requires hormone
66
What are the future therapies for castration resistant prostate cancer?
- Epigenetic targets
- HAT inhibitors
- Bromodomain inhibitors
- Demethylase inhibitors
- Protein-protein interactions
