Quantification of drug-target interactions Flashcards
Define the Molecular mechanisms of action
The interactions between a drug and a target that leads to a specific outcome
What parameters must be considered When measureing MMOA?
The kinetics (Reversible versus irreversible binding) The Binding Site (Orthosteric vs allosteric) The functional effect (activation vs inhibition)
During reversible equilibrium binding of a drug to its target ligand or receptor, Kd and Ki are ____, Inhibition can be ___ or ____, and potential competition can come from ____, ____, or ____
equal
orthosteric
allosteric
Enzyme
Protein:protein interaction inhibitor
Receptor antagonist
Define competitive inhibition
Neither the ligand nor the drug can bind at the same time
Define uncompetitive inhibition
The inhibitor binds the enzyme/receptor only when the natural ligand is bound (potency is proportional to the physiological concentration of the ligand)
Define noncompetitive inhibition
The inhibitor reduces the activity of the enzyme/receptor and binds to an allosteric site, whether it has the ligand bound or not
Define agonism
Binding of compound activates the receptor or enzyme (can be full or partial)
Define allosteric modulation
Binding of compound will modify the activity of a receptor or enzyme, thereby shifting its substrate selectivity profile
How do you calculate the Kd for receptor:binding at equilibrium? What is the unit?
k_off/k_on
Molar
The lower the number, the stronger the interaction
What is receptor occupancy and how is it calculated during Receptor:ligand binding at equilibrium?
The fraction of receptors bound over all receptors
Y = [R:L]/[Rtotal]
Where [Rtotal] = [Rfree] + [R:L]
or
If the receptor concentration is much lower than the ligand concentration of Kd (often the case):
Y = [Ltotal]/([Ltotal] + Kd)
What property is seen when plotting Receptor occupancy over ligand concentration semilogarithmically?
All concentrations show the same curve, just shifted left/right
Define cooperativity
When the binding of a ligand influences binding at another site (oligomeric receptors display this behaviour often)
Can be positive or negative cooperativity
How is cooperativity modelled mathematically?
Y = [L]/([L] +Kd)
Raise the two [L]s and Kd by the Hill coefficient n
A Hill coefficient above 1 shows positive cooperativity
”” below 1 shows negative cooperativity (rare)
How do you calculate Receptor occupancy during reversible competitive inhibition?
Y = [RL]/[Rtotal] or Y = [L]/([L]+K_L(1 + [I]/K-I)) or [L]/([L] + K_app)
How do you calcualte receptor occupancy during irreversible competitive inhibition?
Y = [RL]/[Rtotal] or Y = [L]/([L] + K_L) or [R_unmodified]/[R_total]
When is K_app = [L]?
When Y = 0.5
What is the K_internal of a two-state model of receptor activation? what is the equation?
The rate at which a receptor switches from an inactive form to its active form
K_int = [R^A]/[R^I]
How d you calculate apparent K_d in a two-state model for an agonist?
K_appD = Kd(1+ 1/Kint)
How d you calculate apparent K_d in a two-state model for an inverse agonist?
K_appD = Kd(1+K_int)
What is a partial agonist and how do you calculate its apparent K_d?
An agonist that binds to both active an inactive forms of a two-state receptor (can be positive or negative depending of affinity)
K_appD = (Kint +1)/((K_int/K_A) + (1/K_I))
Where
K_A = [R^A][A]/[R^A:A]
K_I = [R^A][A]/[R^I:A]
At saturating concentrations, a full agonist will lead to _____ whereas an inverse agonist will lead to _____
A partial agonist will lead to ____, and a neutral antagonist leads to ______
Full receptor activation
Full inactivation
Partial activation or inactivation
Competitive inhibition (antagonizes the action of another agonist)
Why does Y not start at 0 in dose response curves?
Even without ligand there’s always an equilibrium between active/inactive forms
Where is K_app on a dose-response graph?
Halfway between minimal response and maximal response (the inflection point of the curve)
What is the enzymatic k_cat
The rate at which an enzyme catalyzes a substrate into its product
