Lecture 3 – Drug Targets
Learning objectives
At the end of this lecture you will…
Be able to describe and classify drug targets
Understand the role of target validation in drug discovery
Describe the role and function of the four main classes of drug target
GPCR
Enzymes
Ion Channels
Nuclear receptors
Draw curly arrow mechanisms for peptide hydrolysis by serine, cysteine,
aspartyl and metalloproteases.
Drug targets
Mostly human (for human diseases)
Notable exceptions:
Bacteria, e.g. MRSA, TB
Viruses, e.g. Covid-19, Influenza, H1N1, herpes, HIV
Fungi, e.g. Candida, Aspergillus
Parasites, e.g. P. falciparium (causes malaria), T. Cruzi
(causes Chagas disease)
These pathogens also have biological processes we
can target
Drug targets by therapeutic area
Respiratory Cardiovascular Gastro-intestinal
Cancer Anti-infectives Inflammation
Tissue repair Pain Metabolic diseases
Central nervous
system
Useful to organise a pharmaceutical company this way
Pharmacologists tend to specialise
Chemists contribute across these boundaries
Classification by molecular target
Adrenergic receptors Respiratory
5-HT receptors Cardiovascular
Angiotensin Gastro-intestinal
Kinases Cancer
Protease inhibitors Anti-infectives
Histamine receptors Inflammation
Acetyl choline receptors Tissue repair
Adenosine receptors Pain
Prostaglandins Metabolic diseases
GABA Central nervous
Na+ ion channels, etc, etc system
Drug targets by popularity
From Overington et al, Nature Reviews Drug Discovery, 2017, 17, 19-34
Good target/ bad target?
Assumption: drug + target effect on disease
Frequently found: drug + target NO effect on disease
Why? Lack of understanding about role of biological
target in the disease
Example – asthma (1970-80s)
Histamine Treatment via
Prostaglandins selective blockade?
Leukotrienes
PAF IL
Mast cell activation
cytokines
FA
releases bio-active Inflammation Multiple biochemical
compounds Bronchoconstriction pathways available
Target validation
Target validation is obtaining evidence that intervention at the target has the desired
effect.
Pharmacologists work extensively in identifying and validating new biological targets.
There are multiple approaches to target validation.
CHEMICAL BIOLOGICAL GENETIC
A selective reagent that is
used to interrogate the A selective antibody Knockout: Total removal or
mechanism of a target deletion of a gene through
genetic engineering. Can’t
Y-shaped proteins produced
study proteins essential to life.
Different to a drug by B-cells in response to
antigens. Highly specific to a Knockdown: Temporary
Drugs need balanced single protein. deactivation or suppression of
properties to treat disease a gene using small interfering
Can be tagged with RNA (siRNA). Binds to protein
Probes only need to be fluorescent molecules to mRNA preventing expression.
potent and selective for a visualise protein expression
target as they are only used CRISPR Cas9 – can remove
in vitro. genes from organisms
Target validation – HIV and CCR5
1. Binds to CD4 receptor
2. CD4 receptorbinds to CCR5 or CXCR4
3. Enters cell
CCR5 is undoubtedly the main human
receptor involved in virus entry and cell-
to-cell spread.
There is a natural mutation (CCR5-delta 32)
where an individual has two mutated copies of
the CCR5 gene.
In this mutation CCR5 cannot bind to the CD4 There is one CCR5 inhibitor on the
protein, meaning that the HIV virus cannot market, Maraviroc.
enter the cell. This provides immunity to HIV
infection.
This natural mutation has been replicated in
a knockout model.
GPCRs (7-TM receptors)
G-Protein Coupled Receptors
Extracellular loop
N terminal
domain
7 trans-
membrane
helices
(most common
binding site)
Intracellular loop
C terminal
domain
Located in cell membranes
Activated by extracellular ligand
Lefkowitz and Kobilka, Nobel Prize (Chemistry, 2012)
GPCRs (7-TM receptors)
GPCRs
Actual structure from X-ray crystallography
Extracellular
Each spiral
structure is an
amino-acid
sequence making Region spanning
an -helix: the cell membrane
Form a bundle
Intracellular
-adrenergic receptor
G-protein NOT SHOWN
GPCR signalling
GPCRs as drug targets
465 FDA approved drugs and
108 unique GPCR targets.
>800 human genes for
GPRCs, 50% not
therapeutically explored
Diverse ligand activation
The largest numbers (>40) of
approved agents (x axis) are
seen for analgesics, allergy
and hypertension, whereas
among agents in clinical trials
(y axis) the highest numbers
of listings (>20) are for agents
for diabetes and neoplasms
(cancers).
Enzymes
Enzymes are nature’s catalysts to facilitate the conversion of
substrates to products.
Enzymes have many diverse roles including:
molecule degradation & metabolism
molecule assembly & processing
generation and control of neurotransmitters, etc.
oxidation and reduction
formation and cleavage of amides, esters, carbon-carbon
bonds
activation processes
degradation, assembly and repair of DNA/RNA
Types of enzyme inhibitors
Competitive
Inhibitors that compete for the enzymes natural substrate, this binding
is reversible.
Non-competitive
Do not compete for the natural substrate. Bind to a co-factor or co-
enzyme.
Could cause conformational change so the natural substrate cannot
bind.
Irreversible
Bind covalently to the active site to permanently deactivate catalytic
function.
Examples of enzyme inhibitors
Kinases
A kinase is a type of enzyme
Its role is to phosphorylate molecules, i.e.
ATP (adenosine triphosphate) ADP
NH2 NH2
N N N N
O O O O O
P P P N P P N
O O O O N O O O N
O O
O O O O O
HO OH HO OH
kinase
O O
P O
Alcohol substrate R OH R O Phosphorylated
substrate
Messages to cells
Classes of kinases
Protein kinases & lipid kinases
Protein kinases phosphorylate OH groups in proteins
Serine, threonine and tyrosine
Histidine kinases phosphorylate the NH group in
histidine
Peptide substrate
Lipid kinases phosphorylate phosphatidylinositols
O
O - OH OH
O
O
P
O
O OH
O
HO OH
O
Phosphatidylinositol substrate
Kinase cascades and cross-talk
Kinases often exist in cascades
Not always linear
Different cascades may activate different pathways
Proteases (proteinases)
Proteases are enzymes that cleave proteins & peptides
Four main mechanistic classes
Aspartyl, e.g. renin, HIV protease
Serine, e.g. thrombin, urokinase, trypsin
Cysteine, e.g. cathepsin K, transglutaminase
Metalloproteases, e.g. matrix metalloproteases,
angiotensin converting enzyme
Aspartyl
Mechanism
O Asp
H
P1 O P1 O O
H H
N N
N N
O H P1' O H P1'
O H O
Asp H
O
O O P1
H Asp Asp
O O H O NH
P1 H OH H O
P1 H O H
N H
N N
N
O P1' OH
O P1' O
O H H
O N
Asp H HO
O Asp
O P1'
Serine
Mechanism
Ser Ser
His His
N N
O H O H N
H Asp H
Asp N O N H
O N N
H P1 O O
H O H H
O N P1 N
N N
O H P1' H P1'
O
peptide substrate
tetrahedral intermediate
Ser Ser
His His
N N
N O H N
O H H Asp H O
Asp N H H
O N H O N N
HN O
O O
O O H O H H P1'
N N
H O
P1
P1' H P1'
NH covalently bound
O H intermediate
Cysteine
Mechanism
Cys Cys
His His
N N
S H S H
H
N N H
N N
H P1 O O
H H
N P1 N
N N
O H P1' H P1'
O
peptide substrate
tetrahedral intermediate
Cys Cys
His His
N N
S H S H O
N H H
N N N
HN H O
O O
O H H P1'
N N
H O
P1
P1' H P1'
NH covalently bound
O H intermediate
Metalloproteases
mechanism
L
L L = part of protein that can bind
Zn2+ L zinc ion, e.g. His, Glu, Asp
P1 O P1 O
H H
N N
N N
O H P1' O H P1'
O H O
H
O
L
L L P1
2+ L L
Zn L
O Zn2+ NH
P1 H OH H P1 H O O H
N H
N N
N
O P1' OH O
O P1' H
O H N
H O HO
O P1'
O
Ion Channels
Ion channels control the passage of ions (Na+, K+, Ca2+) in/out of
cells. Voltage-gated & ligand-gated.
Ligand-gated: e.g. GABA, nicotinic acetylcholine, NMDA receptors
have small polar compounds as endogenous ligands (cf GPCRs)
but are ion channels
How voltage-gated channels work How ligand-gated channels work
Uses of ion channel modulators
Na+ channels important K+ channels important for
for nerve signal regulating heart beat
conduction
Local anaesthetics Anti-arrhythmics
NHSO2Me
Me
H O
N N
NEt2 lidocaine Me
dofetilide
O MeO2SNH
Me
Reduction of H
N Me
Epilepsy insulin secretion N diazoxide
Cl S
Cl O O
Cl
NH2
Cl- channels important
Cl N
N in cystic fibrosis
N NH2
lamotrigine
ivacaftor
Nuclear receptors
Receptors that exist in cell nuclei
Role is to bind to DNA and regulate gene expression, e.g.
I binds to
I O
NH2
HO I CO2H DNA strand
I
thyroxine
Thyroid hormone
receptor (protein) Altered gene
Slow response time expression & protein
production
Altered metabolic rate
Ligand examples include testosterone, oestrogen, thyroxine.
Protein-protein interactions - undruggable?
Protein-protein interactions are large surface area
contacts between two or more proteins.
Until recently considered ‘undruggable’
New technologies in gene-editing
and proteomics allow interactions to
be explored with extreme precision.
Will lead to new drugs against new
targets
Summary
• There are many ways to tackle the treatment of a disease, and these will
have different molecular targets.
• Pharmacologists work to validate targets as a potential therapeutic
approach.
• Medicinal chemists work to design and synthesise molecules that bind to
these targets.
• There are four main types of drug target
GPCR
Enzyme
Ion Channel
Nuclear Receptor
• Understanding how these targets work is critical to developing drugs where
their dysfunction is implicated in the disease.
• Novel targets are needed to develop better, safer drugs. Protein-protein
interactions represent a new developing field of drug targetting.
Next time we will consider how drugs bind to their targets.
