Nathalie Deen
Biology, Radboud University
27-10-2019
Syllabus Pharmacochemistry
All images are derived from the lecture PowerPoints of the course NWI-MOL053
Lecture 1.1: introduction and receptor pharmacology, 2 September
History: an introduction
- Paracelsus: the dose makes the poison
- Diuretics in the foxglove plant lead to a decrease in swollen legs.
- First synthetic pharmaceutical drug was salicyclic acid. (to treat fever). > side effect>
stomach bleeding.
o Aspirin is made out of this substance without the side effects (by Felix Hoffman).
- Acetylation of morphine
o Morphine is the main component of opium (from the poppy plant)
o Very addictive drug
▪ Acetylation of morphine would make it less addictive> this did not work,
acetylation of morphine turns it into
heroin.
- Pregnant women who got treated with Softenon
on gave birth to children with short arms.
Pharmacological phases
- Pharmaceutical phase
o Concentrating to the active compound of the drug
- Pharmacokinetic phase
o The movement of drugs into the body, where they end up. What does the body
do with the drug. ADME
- Pharmacodynamic phase
o After excretion, the drug is available
for action. What does the drug in the
body> drug-receptor interaction
, Nathalie Deen
Biology, Radboud University
27-10-2019
Lecture 1.2: Receptor pharmacology, chapter 1
Drug interaction with receptor, quantifying these processes> try to optimize its activity
Drugs looking for receptors
- The usefulness of a drug is determined by the specificity and selectivity of the drug.
o Specificity: the number of different type of receptors the drug can interact with
o Selectivity: in an ideal world, the drug only has one therapeutic effect and no
other side effects. The selectivity is a property of a drug to exert a limited
amount of effect.
The discovery of High content, high throughput drugs
- Phenotypic screening> select a model system (mouse, flower etc) and library
substances> best compound identified
- Target-based screening> select the target protein, incubate the protein with all the
different compounds you have in the lab-library> then best compound identified
- Structure-based drug design (SBDD)> still in its infancy (not generally used) cause you
need the three dimensional structure of the target and where the binding pocket is
that could bind the protein
Discovery of first-in-class drugs
- First class drugs can be subdivided into target-based and system-based
- Target-based approaches> hypothesis based approaches that aim to manipulate a
biological system (you really know the target in this approach) , by pharmacologically
modulating a specific component or target (enzyme, receptor etc)
- Phenotypic screening> the testing of a large number of, in most cases randomly
selected, compounds in a systems-based assay.
- Chemocentric approach in which compounds with known pharmacology will serve as
the starting point. > Finding the lead compound (?)
- Systems- based approach> hypothesis agnostic assay or approach is based on a
phenotypic change in vitro or in vivo.
Types of targets of drug action
Receptor
Agonist > compound that are able to bind to a receptor and can result in an effect.
Antagonist> compound has binding affinity, it binds to the binding pocket but it will not
result in an effect , it blocks the receptor from binding to an agonist.
- The endogenous mediators are blocked.
Ion channel
- Blockers and modulators
o Blockers, the permeation of the channel is blocked
o Modulators> increased or decreased opening probability
Enzyme
- Inhibitors can target enzymes> the normal reaction is inhibited
- False substrate> will result in an abnormal metabolite production
- Prodrugs> after enzymatic action, drug is produced.
Transporter
- Inhibitor > transport blocked
- False substrate> abnormal accumulation of compound (?)
Types of drug receptors> see also SSA1.3
, Nathalie Deen
Biology, Radboud University
27-10-2019
- Receptor operated channels > takes very short time before drug effect takes place.
o When ion passes, only hyperpolarisation of depolarisation takes place before
there is a cellular effect.
- G protein-coupled receptors> takes seconds
- Enzyme receptors > takes hours
- DNA- coupled receptors (has the longest signal transduction pathway before there is
a cellular effect)
o Example> estrogen receptor
o Drug is taken with carrier protein to nucleus where it lead to (inhibition of )
gene transcription, which will result in different protein synthesis.
- G-protein coupled receptors are a small family but many drugs act on these receptors
as they are involved with many diseases.
- 1800 of the 25000 genes are related with diseases (the diseasome). Drugome: 400
proteins that are produced by the diseaseome genes could be targeted by drugs.
o Proteome is 400 000 > the proteins that get formed out of all the genes.
Technologies used in target discovery
- Genomics (mutations in genes, genetic modification and post-translational
modification (modifications in RNA)) proteomics (researches post0translational
modification) and metabolomics
- If metabolites have changed , than you know there has also something changed in
the function of the protein(s).
- Small molecule (chemical) drugs and larger molecule biological drugs (antibodies)
Reasons for drug failure in clinical trials> low efficacy in humans
Drug-receptor interaction
- Tissue of an organ, incubate it with increasing concentration of the compound (drug)
that you want to test.
- Incubate it until there is an equilibrium of the drug that is bound and not bound (?) or
equilibrium of which how many receptors are bound and not bound (?)Lay tissue on
paper and count the radioactitivity (?)
, Nathalie Deen
Biology, Radboud University
27-10-2019
- Concentration at which 50 percent of the receptors are bound. The middle of the s-
shaped graph. The better the drug, the lower the needed concentration of the drug
to bind 50 percent of the receptors.
Determination of specific binding
- You have a total binding, non-specific binding and specific binding.
- Radioactive-labelled ligand
- First add protecting ligand: All the non-specific sites are now occupied, then add the
radioactive- labelled ligand, they can then only bind the specific binding spots. Then
with that labelled binding amount, you can find the 50% bound receptor amount.
When DR/Rtotaal= 0.5, than 50% of binding
sites are occupied.
Concentration- effect relationship
-
- The Potency of a drug is the amount of drug that is required to achieve a defined
biological effect (the dose) (so bijv the amount that is needed to reach 50% of its max
effect. If 50% of the max effect of drug A is reached at a lower dose than 50% of the
max of drug B, that drug A is more potent. The max effect of B could yet be higher
than of A. then B has a higher efficacy, yet lower potency
- So see which drug reaches highest effect at lowest concentration. > that drug more
potent.
o The smaller the dose that is needed (to reach 50% of the effect for example),
the more potent the drug. It is possible for a drug to be active in a small dose
(potent) but have a low efficacy. A low number for potency means its very
potent.
- Effect is potency > binding : (affinity/ potency)= effect (?)
- If you use simplified forms of the molecules, than you need higher concentration of
the drug to get the same 50% potency (effect), that 50% of all receptors of the
testified tissue are bound. > chemical approach, you know how much receptors the
testified tissue has.
- Efficacy: maximum biological effect a compound (drug) can produce as a result of
receptor binding. : not always 100% (when a smaller form of the molecule is used >
lower efficacy)
- Affinity: a measure of how strongly a compound binds to a receptor.
o Difference affinity and efficacy: an antagonist can have high affinity for a
receptor but will have no efficacy
o A full agonist has 100% of the effect. A full antagonist has 0% efficacy (? Not
sure, not true) . a neutral antagonist, the baseline is at 30% response . with an
inverse agonist, there is a lower activity level than 30%. Antagonist will only lead
to 30% response and not lower (?)
, Nathalie Deen
Biology, Radboud University
27-10-2019
o Full agonist: has high affinity for active form of the receptor. It induces a
complete shift of the equilibrium to the active state. (the opposite of this one is I
guess the inverse agonist> shift to the inactive state).
o A partial agonist has affinity for the active and inactive form of the receptor.
o An agonist can induce a partial effect as it can for example bind both the active
and inactive conformation of the receptor.
- Summary: A partial agonist can stabilize (and has affinity for) the active and inactive
state of a receptor (lil bit more affinity for active state though) . An inverse agonist
has high affinity for the inactive state of the receptor and induces a complete shift of
the equilibrium to the inactive state. A competitive antagonist has equal affinity for
the active as well as the inactive state of the receptor and leaves the equilibrium
undisturbed (it will just block the binding site).
-
Competitive antagonism
- When there is a lot of agonist, it can replace the antagonist and it can still bind the
receptor (competition).
- An allosteric antagonist deforms the receptor. ( in competitive antagonism). The max
effect is always reached, only with a higher concentration of antagonist, a higher
concentration agonist is needed to eventually reach its max efficacy.
- Gaddum equation > this equation shows that if there is
more of B, than you need much higher concentration of A
(agonist ) to reach the same (AR/Rtotal) amount (to reach
the same effect).
o AR is agonist-receptor compound, R is Receptor total, B is
antagonist
- B/K= the dr (the dose ratio)
o The dose of the antagonist that is needed to shift the curve 10 agonists to the
right (10 times more agonist ) (?) . than the dose ratio is 10.
o When you have an concetration of Kb=B, aka 50% of receptors are bound, than
dr = factor 2.
o PA2> actual affinity of antagonist for the receptor.
o Schild equation: Log (dr-1)= log(B) -log (Kb)
o Shift to the right is 33, than dr = 33.
o Schild plot: Intersection of the x-as gives you the Pa2. With
that you can calculate log Kb : pA2= -logKb. So 10^-Pa2 = Kb
, Nathalie Deen
Biology, Radboud University
27-10-2019
Lecture 2.1: recaption of lecture 1. 9 September
Selfstudy assignment 1.3
Most drugs do not bind covalent to receptor cause its irreversible, than the effect lasts to
long and is difficult to control.
Receptor superfamilies:
o Membrane receptor with ion channel (fastest), nicotine receptor
o Membrane receptor that is coupled to an enzyme of ion channel via a G-protein.
Histamine receptor
o Membrane receptor with kinase activity (VEGF receptor)
o Intracellular receptor with ligand that acts on DNA by through
transcription/translation. (slowest signal transduction) (insulin receptor)
- Drugs that interact with only one type of receptor can exert a bread range of effects
in the body as the receptors are expressed in several tissues and organs, there they
are involved with different effect.
- Which molecule can cross a fatty acid membrane better has to do with its
hydrophobic character and not much with its size .
o Having more polar groups and less carbon skeleton means that you are more
polar/ hydrophilic.
o Voor adrenaline dus moeilijk om het membraan te crossen
o Ook met meer polaire groepen meer energy nodig om die los te maken van de
waterige omgeving.
o Valinomycin can let polar ion cross the membrane as it has polar structures
pointing inwards, so it can encapsulate the polar ion, and it has hydrophobic
alkyl chains on the outside.
- 7. Door een Cis binding ontstaat er een knak in the chain so the normal straight order
in the membrane is there less so the fluidity of the membrane increases.
Selfstudy assignment 1.4
Question 5.
receptor desensitization: after prolonged agonist exposure> less receptor response, the
receptor is then uncoupled from the signalling cascade, thus the biological effect is
attenuated (verzwakt) . when an antagonist is bound for a longer time period, receptor
sensitization occurs. The receptor will then synthesize more receptors so the cell will get
more sensitive for the agonists effect. (if the drug is an antagonist) then this could lead to
tolerance to drug, as there is an increase dose of the drug needed to achieve an effect or
dependence (symptoms may occur when acutely stopped with the drug).
question 8a
Het kan geen antagonist zijn, want bij een antagonist zou de cAMP concentratie hetzelfde
blijven en niet minder worden, want de receptor zou dan volledig geblokkeerd worden.
Inverse agonist zorgt ervoor dat de receptor van de active state naar de inacive state gaat.
Receptor signaal zal verminderen, dus cAMP concentratie zal minder afnemen en dichter bij
een rechte lijn komen.
Agonist zorgt voor inhibitie van cAMP. De agonist stimuleert de receptor wel maar zorgt wel
voor een inhibiting signaal naar de cAMP productie.
Question 11
, Nathalie Deen
Biology, Radboud University
27-10-2019
Potency
- 50% of the maximum effect is reached, doesn’t have to mean that 50% of the
receptors is occupied.
Efficacy
- Maximal effect> ACD have the same efficacy
- Partial agonist ( compound B) is that binding of the compound does give a signal but
does not have the same high response (effect). Binding of an antagonist would result
in no response.
SSA 1.5
2. New drug targets are identified by using the knowledge of the human genome,
proteome and metabolome. > using bioinformatics, gene expression data etc. this
can be done by examining expressed mRNA levels in diseases and overall gene
expression.
o Phenotypic screening could be used> which is identifying the substances like small
molecules that alter the phenotype of a cell or an organism in a desired manner.
o A target is considered sufficiently promising when it has at least a causal role in the
disease.
o A receptor is a target for cancer and therefore also a target for an anticancer drug
when
▪ The receptor is dysfunctional In cancer cells> behaves different than in normal
cells (?)
▪ Or the target is essential for survival of the cancer cell> the receptor becoming
inactive will lead to cell death.
▪ Or Other macromolecules cannot compensate for the action of the unnormal
functioning of the target.
▪ Or when the target is only presented in tumour cells.
o To discover ligands for a new drug target receptor, you can express the target in a
reporter cell system that will fluorenes when the target is stimulated, the stimulated
receptor will increase camp or ca production or gene transcription. Testing whether
the ligand has an agonist or antagonist action, you can use an known ligand with a
known function on the receptor and use that as a starting point.
o They appear to be toxic or have a low efficacy.
3. Experimenten die uitgevoerd kunnen worden om eigenschappen van drug X te
achterhalen ( see lecture 2.2 )
a. To find out how powerful drug X is and how it changes with different doses or
way of administering, or time of administration, you could determine the
efficacy and potency from a concentration- response curve from an organ
bath experiment with isolated tissue. Also check for side effects.
b. To find out what the anatomical localization is of the target of X, you should
incubate the lunch tissue with radiolabelled and make microscopic tissue
slices. Then you find where its bound, than relate this to the histology of the
of the tissue> cell types that are found there etc.
i. Trying to let X bind to other tissues can give an idea of what the
binding affinity (Kd) is of X to the different tissues and having known
receptor types in the different tissues also will give an idea with which
receptor X interacts.
, Nathalie Deen
Biology, Radboud University
27-10-2019
c. The molecular mechanism of a drug can be found by determining the signal
transduction mechanism> measuring second messenger, gene expression,
enzyme activity, phosphorylation of proteins etc.
d. Drugability= usefulness of a medicine> measure the bioavailability after
inhalation or oral intake (or intravenously, not in this trachea experiment),
measure halflife (time of elimination), plasma-protein binding and distribution
Exam question
What is the Kb of kaitocephalin. The affinity of kaitocephalin for the receptors> the Kb
(binding constant)
- First pA2 uitvinden. = is the intersection with the xas bij Y=0.0
o in grafiek kijken waar op de Yas 0.0 zit. Dan de lijn bij 0.0 kijken waar hij bij de
xas zit (in grafiek kunnen x en y as ook kruisen bij een negatieve y NML)
o the Pa2 should always be a positive number. Dus stel er staat -6.8. dan is Pa2 =
6.8
- it is a competitive compound because there is a parallel shift to the right.
- Which agonist is more potent > there need to be a higher concentration of
kaitocephalin to get the same response so NMDA (the natural neurotransmitter) is
more potent.
- What type of agonist is kaitocephalin at high concentrations?
o A higher concentration of the agonist NMDA makes sure there is a lower
maximum resonpse value.
o NMDA is the agonist and inc
o Kaitocephalin is has the features of a non competitive antagonist at higher
concentration because highering the concetration of NMDA does not result in a
same/ higher response.
o Non competitive > the max value of the curve goes down> typical for non
competitive antagonist
▪ The maximum of dose-response curves decreases.
Lecture 2.2: Disposition of drugs
ADME
- Absorption, distribution, metabolism (biotransformation ) and elimination.
o Absorption: the transportation of the drug from the administration site to the
general circulation.
▪ When taken in a tablet, the dosage form has to be disintergrated and
dissolved, before it can pass through the cell membrane of the enterocytes to
the blood circulation.
▪ The rate of dissolution (absorption (?)) is dependent on the physical
properties of the dosage form> particle size, hardness, etc.
o Distribution: the drug gets distributed into tissues and organs. Only a small
fraction of the dose reaches the site of action, the rest may cause side effects or
gets eliminated.
▪ Want to concentrate most of the drug at site of action.
o Elimination (clearance): the disappearance of the drug from the body, by
excretion into urine or faeces, or by biotransformation.> after
, Nathalie Deen
Biology, Radboud University
27-10-2019
biotransformation, the metabolites are excreted. The metabolites can also still
be pharmacologically active compounds.
▪ Elimination is metabolism and excretion
- Biotransformation compounds can protect the body against drugs.
- When it passes the liver, than it will be in the general circulation, the enzymes in the
liver determine the bioavailibity.
- Liver most important for biotransformation as it contains the compounds.
Elimination
Drug disposition and plasma protein binding
- Binding to a plasma protein> less free available so there is less effect. It can not
easiliy be filtered by glomerulus. Elimination will be less if its bound to a plasma
protein.
- Plasma protein binding: the degree to which drugs attach to proteins in the blood.
The less bound a drug is, the more easily it can travel through cell membranes or
diffuse.
o Blood proteins are ie. Albumin, lipoprotein, glycoprotein and alpha, beta and
gamma globulins
Drug transport across cell membranes
- Balance tussen hydrofillie and lipofillie For diffusion
o Carriers and channels also say something about
bioavailability
o Lipophilic > passive diffusion is high, with
passive diffusion there is no limitation.
o Hydrophilic compound, the passive diffusion
would be low. So dia 6 is a lipophilic.
o With carrier mediated transport, there is a
maximum, because of their max katabolism
(metabolism etc)
o The ionization also plays a role, ionized
compounds can only cross the membrane with
carrier molecules
o Acids can not be easily removed from acidic environments
▪ With a high pKA environment it can be easily removed.
▪ Changing the pH of the urine for excretion.
▪ This is the effect of urnary pH on drug excretion
Effect of drug meabolsim on excretion > biotransformation process
o Very lipophilic compounds cannot be easily excreted. Make them more
watersolible. Phase 1
o Phase 2> add a hydrophilic group to it, than the compound becomes more
hydrophilic.
o Pahse 3> excretion of compound
o Not biotransformed drug will (can) stay in the body
The biotransformation process
- Phase 1:
, Nathalie Deen
Biology, Radboud University
27-10-2019
o Oxidation reduction, hydrolysis of polar products (ions?). oxidation is losing
electrons.
o Introduction of a functional group like oh NH2, COOH or SH
o Most important phase 1 compound is cytochrome p450 (CYP)> it does oxidation
and that is the most important phase 1 reaction. The compound contains iron. >
this way it inserts an oxygen.
o The main reactions of phase 1 biotransformation are oxidation, reduction and
hydrolysis.
o Esters are most easily hydrolysed in the blood (through esterases).
o The enzyme systems that are involved are cytochrome p450 (CYP),
monooxygenases, alcohol dehydrogenases peptidases etc.
- Phase 2
o Adding hydrophilic compounds > so introduction of largeass functional groups
kind of (?)
▪ Conjugation with endogenous substrate
• Methyl, acetyl, glucuronide, sulphate and glutathione
• It’s a glucuronide conjugate when there is glucuronide acid is added
or
o Vb of conjugation with Acetate dia 15
o Phase 2 is coupling to the hydrophilic compound, getting hydrophilic products.
o Phase 2 does not necessarily happen after phase 1.
o The main reactions in phase 2 biotransformation are glucuronidation,
sulphation, conjugation, methylation etc. and include the enzymes
Glucuronyltransferase, sulphotransferase, amino acid N-transferase,
methyltransferase etc.
- Summary and difference biotransformation phases : drugs are compounds that are
foreign to the body (xenobiotics). The purpose of biotransformation is to convert the
drugs into more polar (water soluble ) molecules, so they can get easily excreted. The
other purpose is to convert the drugs into less active molecules, so detoxification.
Phase 1 will make the molecule more polar by introducing it to a functional group.
Phase 2 biotransformation will make the molecule more water soluble because these
phase 2 reactions involve conjugation of an endogenous substrate to the functional
groups in the drug molecule. Therefore the molecule will become better water
soluble.
- Toxification and detoxification during the different phases: The main features of
phase 1 and 2 reactions> 1: the products of phase 1 can be pharmacologically less
active but also more active (through i.e. bioactivation of a prodrug), they can also be
toxic. Phase 2 is considered a detoxification process in which the conjugates are
pharmacologically inactive. Phase 3 also contributes to detoxification as the drug or
its active metabolites are excreted.
