Drug-Receptor Binding Kinetics

Classical Binding

• Ligands bind to receptors, shifting the equilibrium toward association, but the bound complex can also dissociate.
• Association rate involves the ligand's saturation concentration, reaching a steady state where drug binding and unbinding are equal.
• Equilibrium reverses via dilution or washout, enabling measurement of the first-order rate decay (k off).
• Fitting these equations yields both K on and k off parameters, indicating the binding kinetics.

Drug Residency Time Calculation

• Drug residency time = 1 / k off.
• K off is measured in reciprocal time, but the equation converts residency time to minutes, seconds, or hours.
• Represents the average time a drug is bound to its target.
• Longer residency can increase efficacy, as the drug stays on the receptor longer, increasing its affinity.
• This increases duration of action at the site of action.
• KD (equilibrium constant) = k off / K on.

Experimental Determination

Fluorescent Labeling:

• Attach fluorescent label to the ligand and receptor.
• When the ligand binds, proximity causes fluorescence.
• On rate is measured as ligands bind by monitoring increased fluorescence.
• Data fitted to exponential association curve determines K on rate.

Saturation and Washout:

• Saturate receptor with ligand to reach equilibrium (plateau).
• Clear excess free drug (washout) or dilute to shift equilibrium to dissociation.
• As bound complex dissociates, monitor fluorescence decrease to calculate K off.
• K off rate calculated by monitoring first-order decay rate, fitting data to exponential decay curve.

Surface Plasmon Resonance (SPR):

• An optical measurement that measures changes in refractive index.
• Target immobilized on a thin gold-plated sensor.
• Refractive index changes measured as ligand (analyte) flows across surface.
• Ligand binding increases refractive index, measuring binding association rate.
• Washing off ligand measures off rates (refractive index decreases).
• Association and dissociation curves fitted to quantify K on and K off rates.

Enzyme Example

• Enzymatic activity decreases with an inhibitor.
• The normal activity is without an inhibitor.
• Adding an inhibitor decreases the enzymatic activity
• Removing the inhibitor allows the enzymatic activity to return.
• Inhibitors with longer drug residency times affect enzymatic activity longer.
• For example, Dasatinib, which has a long residency time, results in the lowest enzyme activity, indicating it stays bound to the enzyme longer.
• Longer occupancy = longer residency time.
• Drugs differ based on off rate, taking one over the K off yields residency time in minutes.

Clinical Relevance

• Clinical drugs have varying residency times.
• Antiviral drugs have long residency times for a long duration of action.
• Verapamil (calcium blocker): short residency time due to rapid calcium channel dynamics.
• Morphine (opioid): moderate residency time.
• Desloratadine (antihistamine): longer residency time.
• Residency time correlates with drug type being developed.
• The k off value is important in differentiating drugs in the pipeline.
• Tailoring drug residency time allows prioritization of drugs in development.
• Drug residency time shows important clinical effects for drug duration of action and pre-clinically for optimizing drug types.

Target Occupancy

• Drug concentration and target occupancy are related.
• Drug residency time affects occupancy and duration of action.
• Long resonance drug (e.g., half-life of 72 hours) has high occupancy.
• Rapid off rate drug (e.g., 0.7 seconds) has receptor occupancy that dissapates quickly.
• Intermediate residency times affect drug occupancy accordingly.
• Binding kinetics are tied to receptor occupancy and duration of action.
• Drug residency time ties together many aspects of drug activity.

Case Studies

Short-Acting Drugs:

• Drugs like sleep aids, emergency medications, and anesthetics require relatively quick on and off receptor activity.
• Rapid turnover prevents hangover effects or ensures rapid onset.
• Rapid kinetics competes with drug elimination.

PKPD Modeling:

• In vivo, endogenous ligand concentration changes over time.
• Added drugs maintain a steady state concentration.

Sleep Drug (Ambien) Example:

• 40 nanomolar concentration
• ~2 hour elimination rate
• ~23 nanomolar affinity
• 5 minute residency time (fast)
• ~30% occupancy for efficacy
• Rapid kinetics with rapid onset and ~6 hour duration above 30% efficacy.
• Suitable for sleep drug due to efficacy without long-lasting hangover.

Longer-Acting CNS Drug Example:

• 40 nanomolar concentration
• Same elimination rate as previous drug
• Same affinity as previous drug
• 6 hour residency time (longer)
• Changing the off rate also changes the on rate.
• Efficacy lasts almost 10 hours above 30%.
• Not suitable for sleep drug due to slow onset and long duration of action.
• Changing the residency time affects the onset, occupancy, and duration of action.

Optimal Long Duration Drugs:

• Longer duration drugs include antivirals, anti-addiction drugs, neuropsychiatric drugs, and endocrine drugs.
• Less frequent dosing, fewer adverse effects, and lower costs are advantages of long duration drugs.
• High concentration drug with long residency time remains protected on the receptor and is less affected by endogenous ligands or elimination rates.
• This leads to high occupancy for an extended period.
• For Parkinson's medications, this allows for less frequent dosing (e.g., once a day or even weekly).
• Lowering the dose affects receptor occupancy.

Tuberculosis Drug Example

• Modifying a tuberculosis drug at different positions (1-5) can affect its residency time and in vivo activity.
• Evaluating the off rate and correlating residency time with percent survival in vivo demonstrates the impact of modifications.
• Longer residency times directly correlate with higher patient survival rates in animal studies.
• Failing to measure kinetics over a relevant timeframe may lead to missing crucial information in structure-activity relationship (SAR) studies.
• Analyzing binding parameters and kinetics allows for differentiation and optimization of drugs, leading to improved survival rates.

Influence on Drug Effectiveness and Duration of Action

• Determines how long a drug remains active in the body, which influences its therapeutic effect.
• Longer residency times may be more effective for sustained therapeutic action.
• Shorter residency times may require more frequent dosing.
• Understanding pharmacokinetics and drug elimination is important.
• Drug elimination may affect drugs with shorter residency times more significantly.
• Optimizing dosing regimes can be achieved by understanding drug residency time.
• Adverse drug effects can be avoided by controlling drug accumulation through shorter drug residency times.

Implications for Drug Design

• Traditional binding assays may not capture the full picture for compounds with long residency times.
• Assess the binding kinetics and measure the off rate over relevant timeframes.
• Directly measuring the off rate and considering the half-life allows for differentiation and optimization of drugs pre-clinically, improving in vivo efficacy.
• This approach reduces costs and lowers attrition rates.
• Drug residency time is essential for understanding affinity, in vivo efficacy, and duration of action.