Riassunto esame Pharmacology in drug discovery, Prof. Gaetani Silvana, libro consigliato Pharmacology in Drug Discovery - Understanding drug response, Terry P. Kinakin

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allosteric site.)

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NAM negative allosteric modulators, they are allosteric antagonists, as they

antagonize agonist activation of a receptor, reducing its

affinity and/or the efficacy.

(e.g.: palmitate is the negative allosteric modulator for fatty

acid synthase)

Note that the activity of an allosteric modulator needs the

activity of an orthosteric ligand.

Other molecules act as:

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PEM they shift up the response to a drug.

(e.g.: Barbiturics bind GABA receptor on another allosteric site, closer to the

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channel, so that it does not interfere with the orthosteric site but affects the shift of

the channel. Indeed, barbiturics are more toxic than benzodiazepines because they

favour the shifting of the receptor to the activated state. They are NOT activating

the receptor, but interacting with the transduction activity, meaning that they could

even act without GABA; this makes their safety index value very low)

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NEM they shift down the response to a drug.

Often, the effect of a drug gradually diminishes when it is given continuously or

repeatedly; it’s the case of benzodiazepines administration: our brain shows

tolerance towards the drug.

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Tolerance when the same dose in no more able to show the same original

response. It is a gradual process.

In the case of benzodiazepines, tolerance is acquired gradually, but there are also

compounds that induce tachyphylaxis (e.g.: capsaicin, a molecule contained by chili

pepper, acts on TRPV1 receptor, stimulating the excitation of neurons, that is sensed

as heat. It happens that, the next time we eat spicy, we don’t sense the same intensity

of heat; this mechanism is related to receptors desensitisation, in particular, capsaicin

triggers receptor phosphorylation).

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Tachyphylaxis/Desensitisation when the same dosage in no more able to show

the same original response, happening in the course of few minutes.

Many different mechanisms can give rise to these phenomena. They include:

- change in receptors (e.g.: nicotinic receptors being desensitised at the

neuromuscular junction; G protein-coupled receptor being phosphorylated,

diminishing its ability to activate second messenger cascades.)

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- translocation of receptors Prolonged exposure to agonists often results in a

gradual decrease in the number of receptors expressed on the cell surface, that

are taken into the cell by endocytosis of patches of the membrane (e.g.:

internalisation of µ-receptor caused by overstimulation by opioids)

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- exhaustion of mediators can happen that the second messengers, as ATP,

have already been consumed.

- increased metabolic degradation of the drug

- physiological adaptation

- active extrusion of drug from cells (mainly relevant in cancer chemotherapy)

Note that also cross-tolerance can be developed, meaning that if we got used to a

drug that is metabolized by an enzyme that metabolizes also other compounds, we

could develop tolerance also to that drugs.

How is tolerance liked to addiction?

The relation relies on the consequently negative condition to the stimulation (the

drop): drug like benzodiazepines are lipophilic,

which allows them to cross the blood-

brain barrier, and elicit their effect

directly in the brain cells. In case of

repeated assumption, once drug

concentration decreases, it causes

what’s called rebound effect, that

enhances the symptoms the drug was contrasting. This implies that, to maintain the

calming effect, drugs like benzodiazepines must be taken continuously, and this

correlates with a withdrawal state.

PHARMACOKINETICS

We are entering into what is evaluated in Preclinical stages of drug development.

It describes whatever happens to the drugs when they enter a living organism. The

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latter is defined open system the steady state is not attained, so drugs

concentration depends on

the time the measurements

are taken.

In this system, the input is

the drug administration, and

the output is the drug elimination, so that,

only when these rates are equal, a steady

state can be reached. This happens because

the drug undergoes pharmacokinetics

processing, according to the ADME

parameters.

This implies that even in intravenous

administration the elimination rate does not

rely on different drug doses. This image represents a

pharmacokinetic model,

illustrating the journey of a

“new chemical entity”

(NCE) through the body

and its interactions with

various compartments and

processes. The main steps

include absorption,

distribution, metabolism,

and excretion (ADME).

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1. Absorption the process by which the drug enters the bloodstream

(corresponding to the central compartment) from the site of administration. There

are different routes of administration:

- Oral: ingestion through the

gastrointestinal (GI) tract.

Factors like gastric pH,

gastric emptying time, and

presence of food can affect

absorption.

- Intravenous (IV): direct

introduction into the

bloodstream, resulting in

100% bioavailability.

- Intramuscular (IM) and

Subcutaneous (SC):

injection into muscle or

fatty tissue, with absorption rates influenced by blood flow to the injection site.

- Transdermal: through the skin via patches; absorption depends on skin

permeability.

- Inhalation: through the respiratory tract, offering rapid entry into the

bloodstream. →

2. Plasma Protein Binding once in the central compartment, the NCE may bind

to plasma proteins, such as albumin and lipoproteins, which can affect its

bioavailability, as only the unbound fraction is pharmacologically active. Many

factors influence the binding:

- Lipophilic drugs tend to bind more to plasma proteins, as they are poorly

soluble in water and are drawn to the hydrophobic regions of proteins.

- At high drug concentrations, binding sites may become saturated, causing

nonlinear binding. In this case, an increase in drug dose disproportionately

increases the free drug concentration, resulting in toxicity.

- Drugs that bind to the same protein site can displace each other, leading to

higher free concentrations of one or both drugs.

Note that a graph displaying exponential and linear rates of absorption is implying

that both simple

diffusion and transport

mechanisms are

occurring at the cellular

level. Thus, the overall

transfer of molecules

through cells is the sum

of diffusion and the

equilibrium between

influx and efflux.

Initial negative absorption implies the high activation of efflux systems, meaning

that low concentrations may be dominated by efflux processes, but then these can

be overcome through bulk diffusion of higher concentrations.

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3. Plasma Esterase Destruction it’s the enzymatic breakdown of ester-

containing drugs by plasma esterases, which can reduce drugs concentration.

Some drugs are designed as prodrugs, with ester linkages that are cleaved by

esterases to release the active form.

(e.g.: Dopamine is used to treat Alzheimer disease, but it is administered it its

prodrug form L-Dopa, which bypasses the poor permeability of the active drug,

that can cause lower bioavailability at the target site).

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4. Hepatic Metabolism it’s the biochemical modification of drugs in the liver,

primarily through enzyme-mediated processes, transforming them into

metabolites. It involves the Cytochrome P450 family (e.g., CYP3A4, CYP2D6).

Some factors can influence Hepatic Metabolism:

- Genetic Polymorphisms: variations in metabolic enzymes can lead to

differences in drug metabolism rates.

- Certain drugs or foods can induce or inhibit hepatic enzymes, affecting

metabolism (e.g., grapefruit juice inhibits CYP3A4, enhancing the

concentration of drugs in the blood).

- Impaired hepatic function (e.g., liver cirrhosis) can reduce metabolic capacity.

Note that because hepatic degradation can be a major obstacle to attaining a stable

steady-state, molecules are tested in vitro in hepatic enzyme preparations at an early

stage of drug development to identify possible problematic chemical scaffolds.

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5. Renal Excretion it’s the elimination of drugs and their metabolites from the

body via the kidneys, through urine excretion.

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6. Tissue Distribution it’s the dispersion of substances throughout the fluids and

tissues of the body. Some factors influence the distribution:

1. Highly perfused organs (e.g., liver, kidneys, brain) receive drugs more

rapidly.

2. Tight junctions (e.g., blood-brain barrier) restrict drug entry into certain

tissues.

3. Plasma Protein Binding: only unbound drugs can diffuse into tissues.

4. Lipophilic drugs can easily cross cell membranes and accumulate in fatty

tissues.

In this Distribution Model, 2 compartments are classified:

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5. Central Compartment blood and highly perfused organs.

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6. Peripheral Compartments less perfused tissues where drugs may

distribute more slowly.

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Bioavailability it is the amount of drug that is available for physiological action

in the central compartment after absorption. It is particularly measured in case of

oral administration, one of the most common routes of drug administration: once

drugs are absorbed by the gastro-intestinal tract, they deal with the first pass

metabolism (or pre-systemic), in which they are initially metabolized by liver

enzymes before they reach systemic circulation. This significantly reduces their

bioavailability. This is why some drugs, administered as sublingual, could be

completely metabolized if assumed orally.

The bioavailability of a drug is denoted as F and is calculated as the plasma drug

concentration versus time curves in a group of subjects following oral intravenous

administration of the same drug

(meaning 100% of availability,

because this way directly leads to the

portal system). The areas under the

plasma concentration time curves

(AUC) are used to estimate F as

AUC(oral)/AUC(intravenous).

Note that t=0 is not collected because it would correspond to 100% in IV and it is

estimated to be 0% in case of oral administration.

Two drugs having the same AUC but different curves shape are NOT equivalent: if a

drug is completely absorbed in 30 min, it will reach a much higher peak plasma

concentration, and have a more dramatic effect, than if it were absorbed over several

hours; so it might be toxic.

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Bioequivalence when 2 drugs have comparable bioavailability. This happens

when they show:

- Similar Rate of Absorption: often measured as Cmax, is the maximum plasma

concentration.

- Similar AUCs.

- Similar time to reach the peak concentration, known as tmax.

For most drugs, each of these parameters must lie between 80% and 125%. The

equivalent (or “generic”) drug is cheaper because it is not covered by