Part I: Pharmacodynamic Principles Flashcards
Define the term ‘drug‘
Chemical with a selective therapeutic effect
List the main receptor superfamilies from fastest (ms) to slowest (day)
Ligand gated ion channels, G Protein Coupled Receptors, catalytic and nuclear receptors
Most common type of catalytic receptor
Receptor tyrosine kinases
Catalytic receptor meaning
Have a binding site and enzyme activity associated directly within the receptor
Define substrate
Molecule acted upon by an enzyme
Define inhibitor
Substance blocking enzyme activity
Define agonist
It reproduces the effects of the endogenous messenger. (Activator of a receptors response)
Define antagonist
It blocks the effects of the endogenous messenger (Blocker of receptor activation)
Define receptor
Proteins which respond to to an endogenous messenger by initiating a signal
Affinity def
Ability of the drug to bind to its target
Efficacy def
Ability of drug to activate receptor by a conformational change
Example of a nuclear receptor
Steroid hormones
Why are drugs selective but not specific?
Higher affinity for certain receptors, but at high enough concentration they will interact with additional binding sites - side effecs
Examples of an agonist + antagonist for the sympathetic nervous system
Adrenaline -> increase heart rate. <beta blocker like bisoprolol -> decrease rate, blood pressure
What is Law of Mass action in D+R<->DR
Where KD = Koff/Kon =[D][R]/[DR]
Equation for receptor occupancy (α)
α = [D] / ( [D] + KD )
KD def in terms of receptor occupancy
Concentration of drug needed to occupy 50% of receptors (measure of its affinity)
What is dissociation rate constant. WRITE MROE WHEN WATCHED LECTURE (ADVANCED ONE)
Koff = rate that drug unbinds from receptor
Steps of radioligand binding studies
1) Make radiolabelled drug (eg swap hydrogen with titrium). 2) Make preparation containing drug target (membranes, cells, brain). 3) Mix drug + receptors in assay buffer. 4) Equilibrium reached to measure KD. 5) Separate bound from free drug (using filter) and count radioactivity.
How to separate specific binding from total binding in radioligand binding
Use excess non-radioactive drug. Competes for target binding site - displaces them. Radioactive left = non-specific. Total - non-specific = Specific binding
What is saturation analysis in radioligands and how to find Bmax and KD
Vary concentration of radioligand to get a graph of bound ligands (count) against conc of radioligand. Bmax is the amount of bound ligands where it plateaus. KD is the radio conc at 1/2 of the bound ligands of Bmax.
What is competition analysis in radioligands. How to find IC50
Same conc of radiolig but change unlabelled ligand conc. As it increases, bound radioligands decrease. Graph of count against unlabelled conc. The middle between the 2 plateaus is the IC50.
Find dissociation constant (Ki) using IC50 from competing ligand and the name of the equation.
Cheng-Prusoff correction:
Ki = IC50 / (1 + [radio]/KD(radio))
Use of bioassay
To quantify drug action as a functional response
Different types of bioessays and some disadvantages
Callular, Tissue, Organ, System. Cellular may not behave same in actual organ. For the rest: response composed of many effects, drug could be working on other receptors too
Describe agonist conc-response curve. What is Rmax and EC50?
Muscle respons compared to reference (in %) against conc of agonist. Rmax is max response where it plateaus. EC50 is Effective Conc of agonist at 50% response.
What does EC50 tell you about the agonist
Its potency
Sigmoid equation to find R from con-response curve
R = Rmax * [D] / ([D] + EC50)
Difference between full and partial agonist
Full has the same Rmax, partial has less compared to the other agonist therefore it has lower efficacy (ability to activate receptors)
What is equipotent vs a drug that is more potent than another
Equipotent has the same EC50 value, more potent has a LOWER EC50
What is receptor reserve
Spare receptors bc not all needed due to signal amplification as 1 active receptor can cause a cascade of responses
What factors do EC50 (50% response) and Rmax depend on
EC50: Affinity, efficacy, amplification. Rmax: Efficacy, amplification
3 types of antagonists and examples
Receptor. Physiological antag: 2 drugs that counteract by separate molecular mechanism eg noradrenaline and acetycholine on heart. Antag of the stimulating messenger: Antibody therapy that inactivate the protein growth factors which stops them from stimulating receptors.
Competitive vs non-competitive antagonist def
Comp: Antag binds at orthosteric (same) site as agonist. Non-comp: Antag binds at an allosteric (different) site
Reversible vs irreversible antag
Irreversible is covalent bond, other is not
What is use-dependence
In non-comp antagonists, agonist must open channel before antagonist can block
What is surmountable antagonism and what does it do to conc-response curve
For reversible antags: If agonist conc is sufficiently high, it can reoccupy enough receptors to reach original Rmax before antagonists
When does non-surmountable antagonism happen + conc-response curve effect
If antag is irreversible OR if antag is non-competitive as it binds at allosteric (different) site, Rmax is decreased
How is concentration ratio (CR) measured for antagonists
Agonist EC50 in presence of antag/Original EC50 of agonist
What is the Gaddum equation
KD (or Kb for Gaddum) = [Antagonist] / (CR - 1) to show affinity of antagonist
What is Schild plot
KD is worked out using many different [antagonist]. Then plotted on log(CR-1) against log(antag) for a straight line. x-intercept is - logKD (pA2)
What is pA2
-log(KD) found from Schild plot (x-intercept)
What type of antagonist must it be for Gaddum and Schild to work?
Competitive reversible and surmountable
2 types of enzyme drug action
- Communication via cellular messengers. 2. Chemotherapy
Neurotransmission pathway
- Synthesis of NT in presynaptic 2. Storage 3. Release from presynaptic neuron in response to an Action Potential. Then calcium entry. Then presynaptic inhibitory receptors which inhibits further NT release. 4. Postsynaptic receptors lead to signalling. 5. Metabolism (removing NT from synapse) by enzyme. 6. Uptake of post NT back into presynap unchanged
3 ways drugs csn be used
As substrates, inhibitors, false substrates
Explain drugs as substrates
Boosts conversion to a desired product. Eg A–enzyme–>B so add A which produced desired B.
Explain drugs as inhibitors
Molecule that competes with original molecule of rate limiting step -> prevents synthesis
Explain drugs as false subtrates
Are substrates for target enzyme BUT produces slightly different product - reduced biological activity
How do NSAIDs work
Cyclooxygenase (COX) synthesises prostaglandins (inflammatory lipid mediators) but NSAIDS inhibits synthesis
Explain chemotherapy as enzyme drug action
Agents that are selectively toxic to the target cells compared to normal human cells using biochemical differences. Several of these targets are enzymes
Examples of chemotherapy
Treatment of infectious disease (fungal, bacterial, viruses).
Cancer chemotherapy - targeting rapidly dividing tumour cells
What is a suicide substrate
Antibiotics that target final enzymes in BACTERIAL cell wall synthesis (cell walls don’t exist in human cells)
What are 2 cancer chemotherapies where enzymes are involved in mitosis
Inhibitors of purine nucleotide synthesis (methotrexate) and inhibitors of DNA/RNA synthesis (doxorubicin)
What is on a Michaelis-Menten graph
Enzyme acticity (% to control) vs [substrate]. It was Vmax and half is Km
Michaelis menten equation
V = Vmax * [S] / ([S] + Km)
Definition of Michaelis-Menten constant, Km
The concentration of substrate at which 50% of the maximal enzyme velocity (V) is obtained
What is the steady state approximation and why is it useful
That [ES] (enzyme + substrate) does not change. Can derive an expression for Km based on it
What is kcat
Catalytic rate constant. Between ES comples -> E + P
Km equation
(koff + kcat)/kon
KD equation and what does it represent
koff/kon. The affinity of the substrate for the enzyme - 50% enzyme occupancy
What does it mean if Km > KD
The concentration of substrate required for 50% velocity is greater than for 50% enzyme occupancy
How does competitive reversible inhibitor change Michaelis-Menten plot
Same Vmax, higher Km. Shows it is surmountable inhibition.
If binding equilibrium is reached fast aka koff»_space; kcat then what happens to KD and Km
They are almost equal to each other
Equation to work out dissociation constant Ki (inhibitor affinity) from Km
Km(obs) = Km(control) * (1 + [inhibitor]/Ki)
What is on a competition curve for comp reversible inhibitors
Enzyme activity % against Log[inhibitor] at constant [substrate]. Can compare different inhibitors. At 50% of acitivity -> IC50 of inhibitor!
Work out Ki using Cheng-Prusoff eq:
Ki = IC50 / (1 + [substrate]/Km )
How does irreversible inhibitor change Michaelis-Menten plot
Decreased Vmax (not surmountable) but no change in Km
How does non-competitive reversible inhibitor change Michaelis-Menten plot
Vmax decreased and Km unchanged
What does a non-competitive inhibitor bind to
Both enzyme and ES complex at an allosteric site
How to calculate Ki from non-comp reversible inhibs
Vmax(obs) = Vmax(control) / (1 + [inhib]/Ki )
