4. Principles Of Drug (& Hormone) Action Flashcards
What is pharmacology?
The study of how drugs affect biological systems.
Describe the branches of pharmacology.
- Pure pharmacology -> Identification and study of chemically sensitive sites and their mechanisms
- Applied pharmacology
- Chemical physiology -> How do these sites play a role in normal function?
- Therapeutics -> Can these sites be exploited in treatment?
- Toxicology -> Do these sites mediate toxic effects of chemicals?
Define a drug.
- Any chemical agent which affects any biological process.
- Generally organic chemicals of small molecular weight (<500g/mol)
Do drugs have to be synthetic?
No, the term also encompasses bodily chemicals.
What are some examples of biopharmaceuticals?
- Therapeutic proteins from genetic engineering (e.g. hormones, blood clotting factors, enzymes)
- Nucleic acids
- Vaccines
- Monoclonal antibodies
- Cell-based therapies
What are the different methods of drug classification?
Based on:
- Chemical structure
- Main effect (e.g. analgesics)
- Therapeutic use (e.g. antidepressants)
- Mechanism of action
What are pharmacodynamics and pharmacokinetics?
- Pharmacodynamics -> What the drug does to the body (e.g. drug receptors, efficacy, responses, toxicity, etc.)
- Pharmacokinetics -> What the body does to the drug
What mnemonic is used to remember the branches of pharmacokinetics?
ADME:
- Absorption
- Distribution
- Metabolism
- Excretion
How can drugs cause a toxic effect on a molecular and cellular level?
On a molecular level, chemicals can:
- Interact with proteins, lipids and DNA
On a cellular level, chemicals can:
- Interfere with receptor-ligan binding
- Interfere with membrane function
- Interfere with cellular energy production
- Bind to biomolecules
- Perturb homeostasis (e.g. calcium levels)
Define a receptor.
A protein or macromolecule in or on a cell that is a recognition site for endogenous ligands or drugs. When a drug binds it initiates or blocks a response to an endogenous chemical.
Define endogenous.
Originating from within an organism.
What are main subtypes of drugs that bind to a receptor and what do they do?
- Agonists -> May ctivate more than one type of receptor (e.g. ACh activates nicotinic and muscarinic receptors)
- Antagonists -> Selectively block one type of receptor subtype (e.g. curare)
What are some criteria for a receptor?
- Recognition -> Receptor protein must exist in a conformational state that allows for recognition
- Saturability
- Reversibility
- Stereoselectivity -> Receptor must recognise only one of the optical isomers
- Agonist specificity -> Structurally related drugs should bind well
- Tissue specificity -> Binding should occur in the correct tissues
- Transduction -> The binding of the agonist must be transduced into some sort of functional response
Give some examples of endogenous agents that act on receptors.
- Hormones
- Neurotransmitters
- Growth factors
- Vaso-active factors
Why are drugs so potent?
Their action is amplified by, for example, second messenger systems.
Define affinity.
The tenacity by which a drug binds to its receptor.
State the primary assumption of receptor theory.
A single ligand is binding to a homogenous population of receptors.
Describe the concept of receptor kinetics.
[L] + [R] -> [LR]
- kon is the rate of the forwards reaction
- koff is the rate of the backwards reaction
- At equilibrium, the rate of formation equals the rate of dissociation, so: [L][R]kon = [LR]koff
- KD is the equilibrium dissociation constant: KD = koff / kon = [L][R]/[LR]
- KD is the [L] which produces 50% receptor occupancy
What is KD in receptor kinetics? What are the units?
- It is an inverse measure of receptor affinity for a particular ligand
- It is the concentration of the ligand that produces 50% receptor occupancy
- Units: M/L
What does a high KD value indicate?
A low receptor affinity for a certain drug.
Describe the idea behind drug binding studies.
- Measure the affinity not efficacy
- No need for intact cells or tissues
- Need a radio-ligand
- Saturation binding curve may be plotted, where the drug concentration producing 50% binding is the KD value
How do agonists work?
Bind to receptor causing a change of conformation, leading to a cellular response.
Do agonists and antagonists have both an affinity and efficacy?
- Agonist -> Affinity and efficacy
- Antagonist -> Affinity but no efficacy
How do antagonists work?
- Prevent agonist-mediated response by preventing a drug from binding and eliciting its normal response.
- They may be competitive or non-competitive.
What factors can be meaured in antagonist action?
- Affinity
- Potency
- Selectivity
What is efficacy?
The ‘strength’ of an agonist-receptor interaction along with the transduction mechanism’s in eliciting a response.
Is the effect of a drug proportional to the fraction of receptors occupied?
Yes, that is the classic theory. Note, however, that in some cases not all receptors need to be bound in order to produce the maximal effect.
How can the effect of a drug depending on dose be represented graphically?
A dose-response curve can be plotted, with either the drug concentration or log of the drug concentration on the x-axis. The logarithmic graph may be more suitable due to the wide range in drug concentrations used.
Describe the shape of a non-logarithmic and semi-logarithmic dose-response curve.
- Non-logarithmic -> Usually a rectangular hyperbola
- Semi-logarithmic -> Usually sigmoidal
What is EC50?
- Effective concentration 50
- The drug concentration that produces response halfway between the baseline and the maximal response
What is the difference between KD and EC50?
- KD is a measure of the affinity of the receptor for the drug
- EC50 is a measure of the potency of the drug
The two values are not necessarily the same (LOOK UP WHY!)
What are the different degrees of agonist?
In order of efficacy: Full antagonist > Partial antagonist > Antagonist
What is a partial agonist?
An agonist that only has partial efficacy relative to a full agonist, so that they are unable to cause a maximal response (even when all receptors are bound).
Give some clinical uses of partial agonists.
Partial agonists can be used to “wean off” abused drugs, by replacing them with the partial agonist which produces a lesser response.
e.g. Methadone for heroine abuse treatment
Are agonists and antagonists structurally similar?
Yes, they may be similar structurally, so an antagonist could be synthesised by modifying an agonist so that affinity remains high while efficacy is reduced.
Define a drug target.
Any biological binding/recognition element (usually a protein) for a drug.
Give some examples of drug targets.
- Receptors
- Ion channels
- Carriers or transporters
- Enzymes
What are the different types of receptors?
Cell-surface receptors:
- Ligand-gated ion channels (ionotropic)
- G-protein coupled receptors (metabotropic)
- Enzyme-linked receptors
There are also nuclear receptors for hormones.
What is another name for ligand-gated ion channels?
Ionotropic
What is another name for GPCRs?
Metabotropic receptors
Why are there sometimes said to be 3 types of receptors and sometimes 4 types?
Because there are 3 types of cell-surface receptors, but there are also nuclear receptors which are sometimes also included as a 4th type.
Give some examples of ligand-gated ion channels (ionotropic receptors).
- nAChR
- GABA receptor
- Glutamate receptor
Give some examples of GPCRs.
- mAChR
- Alpha and beta adrenoreceptors
Give some examples of enzyme-linked receptors.
- Cytokine receptors
- Insulin receptors
- Atrial natriuretic factor (ANF) receptor
Give some examples of nuclear receptors.
- Receptors for steroid hormones
- Receptors for thyroid hormone
Describe the principles of how each of the receptor types works and how long the effects last.
What are receptor subtypes?
- Various different receptor kinds within each type of receptor.
- They have slightly different amino acid sequences, which may affect their sensitivity to different drugs
- e.g. Different nAChR types in brain and in muscle
Describe how different receptor subtypes may arise.
- Alternative splicing
- Post-transcriptional modifications, including mRNA splicing and mRNA editing
Describe the structure of ligand-gated ion channels (ionotropic receptors).
Describe the structure of nAChR.
What is the significance of the charge on ligand-gated ion channels?
They attract either anions or cations to the channel.
Are neurotransmitters acting on ion channels always excitatory?
No, inhibitory neurotransmitters (such as GABA) can cause membrane hyperpolarisation due to an inwards Cl- current.
Describe the structure of GPCRs (metabotropic receptors).
Describe simply the general functioning of GPCRs.
Why do GPCRs have a wide range of effects?
The G protein can interact with many different things.
What are the main types of G-protein types?
- Gs
- Gi
- Gq
- Other types -> Gt
Are GPCRs always activated by a neurotransmitter?
No, they may also be activated by protease cleavage of the extracellular N-terminal tail.
What is the clinical relevance of GPCRs?
About 50% of modern therapeutic drugs act on GPCRs.
Describe the mechanism of G-protein activation.
- αβγ is a trimer formed from the covalently bound β/γ, but not α
- When the ligand is not bound, the αβγ trimer is not connected to the receptor and GDP is bound to the α unit
- When the ligand binds, it triggers a conformational change in the receptor, which increases the affinity for the αβγ trimer -> αβγ trimer binds to the receptor
- α’s affinity for GTP increases -> So the GDP bound to α is replaced by GTP
- The α (bound to the GTP) and β/γ units travel to their targets
- Hydrolysis of GTP terminates signalling
Note: An individual agonist-receptor complex can activate more than one G-protein, so this process can happen with one G-protein binding after another.
Does an individual agonist-GPCR complex activate just one G-protein?
No, one G-protein can bind, then another, then another…
Describe the amplification mechanism in GPCRs.
- Individual agonist-GPCR complex can activate more than one G-protein
- An active G-protein can interact with the effector enzyme for a duration that is sufficient to generate multiple molecules of product
- If the product is a second messenger, further amplification occurs
Define a second messenger.
A substance whose release within a cell is promoted by an agonist and which brings about a response by the cell.
Name some of the targets for G-proteins.
- Adenylate cyclase -> Leading to cAMP production
- Phospholipase C -> Initiating IP3 and DAG signalling pathways
- Ion channels -> Modulated especially via interaction with β/γ subunits
- Rho kinase
- MAP kinase
What do Gi, Gs and Gq proteins stand for?
- Gi -> Inhibitory
- Gs -> Stimulatory
- Gq -> No name really
What mechanisms do Gi/Go, Gs and Gq proteins modulate?
- Gi/Go -> Inhibiting adenylate cyclase activity (increasing cAMP production)
- Gs -> Stimulating adenylate cyclase activity (decreasing cAMP production)
- Gq -> Inositol phosphate pathway
Which G-proteins function by an equivalent mechanism?
Gi and Gs proteins both modulate adenylate cyclase (an enzyme in the plasma membrane) activity, but Gi inhibits it, while Gs stimulates it.
Describe the Gi and Gs protein mechanism of action.
- After the G protein is activated (see flashcard on this!), the α subunit bound to GTP moves to adenylate cyclase
- With αs subunits, this results in the stimulation of adenylate cyclase, so that cAMP production is increased
- With αi subunits, this results in the inhibition of adenylate cyclase, so that cAMP production is reduced
- cAMP acts as a second messenger to activate various processes, including activation of cAMP-dependent protein kinases
What are Go-proteins?
Gi proteins are often called Gi/o proteins. (Check this!)
Give an example of when adenylate cyclase may be inhibited due to receptor binding.
M2 and M4 inhibitory muscarinic receptors are GPCRs that are coupled to Gi proteins, so that adenylate cyclase activity is inhibited.
What is the full name for cAMP?
Cyclic 3’,5’-AMP
Describe the synthesis and breakdown of cAMP.
Synthesis:
- Synthesised from ATP
- Involves removal of PPi
- Catalysed by adenylate cyclase
Breakdown:
- Hydrolysed by cAMP phosphodiesterase
- Produces 5’-AMP
What terminates the effects of cAMP?
cAMP phosphodiesterase hydrolyses the cAMP to 5’-AMP.
What does cAMP do?
It activates cAMP-dependent protein kinase A (PKA). This protein kinase can then phosphorylate specific enzymes.
Describe the entire cAMP pathway when it is activated by a GPCR.
- Ligand binding to GPCR causes activation of G-protein
- G-protein activates (if Gs) or inhibits (if Gi) adenylate cyclase
- Adenylate cyclase catalyses the formation of cAMP from ATP
- cAMP activates protein kinase A (PKA)
- PKA phosphorylates specific enzymes, modifying their activity
How is cAMP degraded and how can this be inhibited?
- By cAMP phosphodiesterase
- This is inhibited by methylxanthines
Describe the Gq protein mechanism of action.
- After the G protein is activated (see flashcard on this!), the αq subunit bound to GTP moves to the PLC enzyme (phospholipase C)
- PLC in turn hydrolyses PIP2 to DAG (diacyl glycerol) and IP3 (inositol triphosphate)
- IP3 acts as a second messenger -> Binds to to IP3 receptors, triggering Ca2+ release from the ER
- DAG also acts as a second messenger -> Activates protein kinase C
Compare the protein kinases activated by Gi and Gs proteins with those activated by Gq proteins.
- Gi and Gs -> Protein kinase A (PKA)
- Gq -> Protein kinase C (PKC)
What is GTP?
Guanosine triphosphate -> A nucleoside triphosphate.
What does PIP2 stand for?
Phosphatidylinositol 4,5-bisphosphate (probably don’t need to know this though)
What does DAG stand for?
Diacylglycerol
What does IP3 stand for?
Inositol triphosphate
Where are IP3 receptors found?
On the endoplasmic reticulum
How is PIP2 resynthesised?
Name two important GPCR you need to know about, apart from muscarinic receptors.
- α1 adrenoreceptor -> Gq protein coupled
- β1 adrenoreceptor -> Gs protein coupled
Where are β1 adrenoreceptors found, how are they activated and what is the effect of this?
- Found on cardiac myocytes
- Activated by the binding of noradrenaline or adrenaline
- Gs protein triggers adenylate cyclase, which increases cAMP, which results in increased PKA activity
- PKA activates calcium channels in the cell membrane, so that intracellular Ca2+ concentration is increased -> This results in contraction
Where are α1 adrenoreceptors found, how are they activated and what is the effect of this?
- Found on vascular smooth muscle cells
- Activated by noradrenaline
- Gq protein activates PLC, which increases intracellular IP3
- IP3 leads to increased release of Ca2+ from the sarcoplasmic reticulum, which causes contraction
How do beta-blockers work?
- Block β1 receptors so that adrenaline can’t bind
- Therefore the cAMP pathway is not activated and contraction of the cardiac myocyte does not occur
- This reduces cardiac contraction
What are some other names for adrenaline and noradrenaline?
- Adrenaline = Epinephrine
- Noradrenaline = Norepinephrine
Describe the structure of enzyme-linked receptors.
Describe the principle on which enzyme-linked receptors work.
- Extracellular domain of receptor has receptor functions, while intracellular domain has enzymatic functions
- Binding of a ligand causes a conformational change in the enzymatic domain
- Frequently, this leads to control of gene transcription and therefore various cellular effects
What ligands bind to enzyme-linked receptors?
- Growth factors
- Cytokines
- Hormones
What cellular effects do enzyme-linked receptors lead to?
They affect gene transcription, so can alter:
- Cell division
- Inflammation
- Apoptosis
What are the main types of enzyme-linked receptors?
[EXTRA]
- Receptor tyrosine kinases
- Receptor serine/threonine kinases
- Cytokine receptors
Give an example of an enzyme-linked receptor signalling pathway.
Where are nuclear receptors found?
In the cytoplasm or the nucleus.
Describe the structure of nuclear receptors.
What types of molecules are nuclear receptors?
Transcription factors
Describe the principle on which nuclear receptors work.
- Have a DNA-binding domain and ligand-binding domain
- When the ligand binds, the receptor can act as a transcription factor, modulating DNA transcription
What ligands bind to nuclear receptors?
- Steroid hormones
- Lipids
What are the different types of nuclear receptor and how does each work?
Class I:
- Present in cytoplasm
- Bind steroid hormones
- Binding of ligand triggers homo-dimerisation and migration to nucleus
Class II:
- Present in nucleus
- Bind lipids
- Binding of ligand triggers dimerisation with retinoid receptor
What are the main types of drugs that act on ion channels?
- Blockers -> Occlude pathway directly
- Modulators or “gating modifiers” (can be inhibitors or activators)
What are the main types of drugs that act on ion transporters?
- Inhibitors
What are the main types of drugs that act on enzymes?
- Inhibitors -> Prevent normal reaction
- False substrate -> Drug is processed by gives abnormal product that interferes with a cellular pathway
- Prodrug -> Require processing to be turned into an active form
