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Describe Drug Binding (briefly)

Drugs exert effects by binding to a molecular target which is normally a protein.

There are some exceptions e.g. Some anti microbial and anti tumour drugs bind DNA.

Specificity indicates how effective the drug will be.

Most drugs bind reversibly with receptors, with binding governed by association and disassociation rates.


What are Current Drug Targets?

47% Enzymes

30% GPCRs - very druggable, ~800 GPCRs in man but currently ~30 targets

7% Ion channels

4% Transporters

4% Nuclear hormone receptors

4% Other receptors

2% Miscellaneous

1% Integrins

1% DNA


Explain the relevance of the concentration of drug molecules

The concentration of drug molecules around receptors is critical in determining drug action and therefore regulate drug activity

It is important to consider drug concentrations in molarity as drugs as equivalent molar concentrations have the same concentration of drug molecules but drugs of equivalent concentrations by weight may not.

Molarity = (grams per litre) / (molecular weight)

NOTE: binding obeys the law of mass action (related to concentrations of reactants and products)


Explain Affinity and Intrinsic Efficacy

Drug action (receptor activity) is dictated by: AFFINITY AND INTRINSIC EFFICACY

Agonist: a ligand that causes a response; has both affinity and intrinsic efficacy.

Antagonist: has affinity only, no intrinsic efficacy

Affinity: a measure of how well these 2 molecules interact; high affinity - ligand binds tightly to the receptor. Affinity GOVERNS BINDING

Intrinsic Efficacy: a measure of the ability of a compound to activate the receptor. Intrinsic Efficacy GOVERNS ACTIVATION

To generate a response, activation of the receptor is required.


How do we measure Drug-Receptor Interactions by binding?

Often by binding of a radioactively-labelled ligand (radio ligand) to cells or membranes prepared from cells.

A cell can have multiple receptors, e.g. 10,000-100,000per cell

How much binding occurs is governed by concentration of radio ligand.


Discuss the Quantification of Drug-Receptor Interaction (Binding)

B(max) is the maximum binding capacity (all receptors are occupied)

K(d) = the concentration of ligand required to occupy 50% of the available receptors = dissociation constant

K(d) is a measure of affinity - the reciprocal of affinity; the lower the K(d) the higher the affinity (a drug has high affinity if only a little amount of it is required to occupy 50% of receptors)


What is K(A)?

Affinity can be measured by means other than radioligand binding e,g, pharmacologically, often known as K(A) instead of K(D)


Describe the scale of [Drug]

[Drug] is usually on a logarithmic scale! not linear.

Logarithm = exponent by which a fixed (base) value has to be raised to give a particular number.

The curve is normally sigmoidal.


Discuss the relationship between [drug] and response

Response could be e.g. A change in a signalling pathway or a change in cell or tissue behaviour (contraction)

EC(50) = effective concentration of drug giving 50% of the maximal response.

This is a measure of agonist potency (how good the drug is at giving a biological response)


What is Potency?

EC(50) is a measure of Potency.

Potency depends on BOTH affinity and intrinsic efficacy PLUS cell/tissue-specific components (thus these factors combined reflect efficacy).

The same potency could occur with different combinations of affinity and efficacy

Potency dictates how much of a drug you need to take in order for it work.

The number of receptors available also governs potency.


What is the difference between concentration and dose?

Concentrations the known concentration of drug at site of action e,g, in cells and tissues

Dose: concentration at site of action unknown - e.g. Dose to a patient in mg or mg/kg

The two terms are often used interchangeably


How is Efficacy measured?

In relative terms, with no absolute scale.

Agonists with different E(max) values have different efficacy however agonists with the same E(max) values may not have identical efficacy.

The two drugs may differ in affinity, meaning that the relationship between occupancy and response will be different for the two agonists.

One may be more able to convert binding into function.


Discuss Affinity and Efficacy in a clinical setting

In Asthma, the treatment goal is to activate the B2-adrenoceptors to relax the airways, however there are B-adrenoceptors elsewhere in the body e.g. B1 in the heart increase the force and rate of contraction.

Salbutamol is a B2-adrenoagonist that has a K(d) of 20 micro moles for B1 and 1micromoles for B2. As the K(d) for B2 is lower, it has a higher affinity. As well as this, B2-selective efficacy and route of administration (oral spray) limits B1 -activation and side effects.

This highlights the need for selective activation of B2-adrenoceptors - there is a need for drugs with enhanced selectivity, affinity and/or efficacy.

Salmeterol is a longer acting B2-adrenoagonist but it has no selective efficacy. It prevents B1-activation and side effects purely through differences in affinity.

Salmeterol has a K(d) of 1900nm for B1 and 0.55nm for B2 therefore B2's lower K(d) value gives it 3,455 times greater affinity than B1


What are Spare Receptors?

In some cases

Often seen when receptors are catalytically active e,,g tyrosine kinases or GPCRs.

They are not needed even though ligands can still bind because once a receptor is activated, it can activate many signalling molecules which can each activate multiple effectors (amplification)

So spare receptors exist because of amplification in the signal transduction pathway and response is limited by a post-receptor event.

In the lungs, only 10% of occupancy of Muscarinic receptors is needed for maximal contraction (full biological response)


Why have Spare receptors?

They regulate sensitivity of the tissue to the ligand - increase sensitivity which allows responses at low concentrations of agonist; increasing receptor numbers eg from 10,000 to 20,000 lowers concentration of drug required to produce the full biological action.

Changing receptor number changes agonist potency and can affect the maximal response (position of curve is dictated by number of receptors).

Number of receptors therefore has an effect on potency.


Is the number of receptors fixed?


Tends to increase with low activity (up-regulation)

Tends to decrease with high activity (down-regulation).

For drugs this can contribute to tolerance/tachyphylaxis (loss of responsiveness to drugs).

Receptor numbers change due to physiological, pathological or drug induced changes.


What is a Partial Agonist?

A drug that cannot produce a maximal effect, even with full receptor occupancy.

The EC(50) is a partial agonist is equal to its K(d).

The potency of a drug is dependent on both its affinity and efficacy and therefore partial agonists can be more or less potent than full agonists.

A partial agonist may not always be a partial agonist, depending on the tissue and the biological response.

Increasing receptor number can change a partial agonist into a full agonist. The partial agonist still has low intrinsic efficacy at the each receptor BUT there are sufficient receptors to contribute to a full response.


Discuss the clinical use of Partial Agonists

Opioids are used for pain relief and also for recreational use (e.g. Heroin - euphoria).

However they can cause respiratory depression which can lead to death.

They act primarily through the miu-opioid receptor (GPCR).

In a heroin (diamorphine) addict: full agonist at miu-opioid receptor --> full response.

If an addict injects a stolen narcotic instead of herring which turns out be buprenorphine, he immediately becomes very ill because buprenorphine antagonises the effect of herein - efficacy at receptor insufficient so addict suffers withdrawal symptoms.


Compare the action of buprenorphine to morphine

Morphine is a full agonist of the receptor, used for pain relief

Buprenorphine is a partial agonist has a higher affinity but lower efficacy than morphine.

It binds more tightly and occupies the receptors and limits response.

Heroin can't access receptors

This is how a partial agonist can act as an antagonist of a full agonist and are sometimes referred to as a mixed agonist/antagonist.

Partial agonists can be used in the treatment of opioid addiction e.g. Buprenorphine enables gradual withdrawal and prevents use of other illicit opioids.


Why might buprenorphine be advantageous to morphine in some clinical settings?

E.g. Pain control - adequate pain control, less respiratory depression

Because buprenorphine had a higher affinity but lower efficacy than morphine


How is efficacy used clinically?

Full agonists with identical intrinsic activites and the same affinity may have different efficacies - B is greater - much better at turning receptors on (more to the left on the graph).

Efficacy is often used clinically to describe how good a drug is at producing a response.

Different to the strict pharmacological meaning


What is an antagonist and what are the three different types?

Antagonist block the effects of agonists I.e. Prevent receptor activation by agonists.

1. Reversible competitive antagonism (commonest and most important in therapeutics)

2. Irreversible competitive antagonism

3. Non-competitive antagonism


Explain about Reversible Competitive Antagonism

Relies on a dynamic equilibrium (created by constant association and disassociation) between ligands and receptors

Antagonist and agonist are competing at the same binding site

Adding more antagonist will eventually outcompete the agonist - less agonists will be bound at any one time.

Depends on affinity of these 2 compounds

Greater [antagonist] = greater inhibition.

IC(50) (Inhibitor Constant) is an index of antagonist potency - concentration of antagonist giving 50% inhibition.

K(d) describes antagonist affinity - concentration of antagonist needed to give 50% occupancy.


How can reversible competitive antagonists be overcome?

Can be overcome by high concentrations of agonists, thus causing a parallel shift to the right of the agonist concentration-response curve.


Describe a clinical example of reverse competitive antagonism

Naloxone is a high affinity, competitive antagonist at miu-opioid receptors

It may be useful clinically because can be used for reversal of opioid-mediated respiratory depression (high affinity -binds more tightly, making it more difficult for heroin to access receptors - it will compete effectively with opioids such as heroin for receptors)


Explain about Irreversible Competitive Antagonism

Occurs when the antagonist dissociates slowly or not at all (agonist is nerve able to access the receptor - can't overcome this inhibition) - covalent bonding is involved.

With increased [antagonist] or increased time, more receptors are blocked by antagonist - NON-SURMOUNTABLE.

Irreversible competitive antagonists cause a parallel shift to the right of the agonist concentration-response curve and at higher concentrations suppress the maximal response.

The maximal response is lowered even when you add more agonist.


Give a therapeutic example of irreversible competitive antagonism

Phenoxybenzamine is a non selective irreversible alpha1-adrenoceptor blocker used in hypertension episodes in pheochromocytoma.

Pheochromocytoma is a tumour adrenal Chromaffin cells which pH secrete excessive adrenaline/noradrenaline leading to vasoconstriction.

Irreversible antagonism might be advantageous here because it prevents adrenaline accessing receptors.


Explain about Non-competitive antagonism

Orthosteric site (where a natural ligand/s bind on a receptor)

Allosteric site are where other molecules e.g. Synthetic ligands bind.

Allosteric and non-competitive antagonism provide binding sites for agonists (potential novel drug targets) and molecules that enhance or reduce effects of agonists (no competition for binding site but affect Orthosteric ligand affinity and/of efficacy by causing a conformational change --> non-competitive antagonism).

Effect on pharmacology is similar to irreversible competitive antagonism. Additional experiments are needed to distinguish the differences.


Give an example of Non-Competitive Antagonism

NMDA (glutamate) receptor- ligand gated ion channel (ionotropic receptor)

glutamate binding site is Orthosteric binding site and ketamine binding site is allosteric binding site.


What is the effect of frequent repeated drug application on the tissue response?

Tissue becomes less responsive (desensitised) to a drug (desensitisation/tachyphylaxis)