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3.3 Drug Discovery 55
Histamine
Parietal cell
H
2
Histamine receptor
Proton pump inhibitors
H
2
Antagonists
Gastrin receptor
Proton pump
Gastric cell Gastrin
Antibiotics
H Pylori
Gastric mucosal layer
cAMP
Ca
2+
Gastrin
Antacids (OH
–
)
H
+
Figure 3.5 Partial illustration of the control mechanism that stimulates gastric acid
secretion and the intervention points used to treat ulcers. Parietal cells and gastric cells
form part of the epithelial cell lining of the stomach. Histamine release is usually triggered
as part of the enteric nervous system response to distension of the stomach following
eating food. The interaction of food with the gastric mucosal layer is the normal trigger for
gastric cells to release gastrin, which is then carried by the bloodstream to the parietal
cells. Calcium ions and cyclic AMP act as intracellular messengers in the transfer of the
signal from the receptors to the proton pumps of Parietal cells where the acid is generated.
See text for specific detail.
ulcer and a proton pump inhibitor (omeprazole) to temporarily stop acid secretion
so as to allow the ulcer to heal. The various intervention points in the treatment of
ulcers are illustrated in Figure 3.5.
Target discovery had been greatly facilitated by the disciplines of genomics and
proteomics. The term ‘genome’ refers to the entire complement of hereditary infor-
mation (almost always DNA) present in an organism or virus. A genome invariably
consists of genes (discrete sections of the genome, each of which encodes a specic
protein) separated by non-protein-encoding sections. The discipline of genomics
56 3 Drug Discovery, Classification and Early Stage Development
entails the systematic study of the entire genome of a target organism (a human,
a pathogenic microorganism, or virus for example), with the aim of:
● Sequencing the entire DNA complement of the genome and
● Physically mapping the genome arrangement (assign exact positions in the
genome to the various genes and noncoding regions).
While classically genome sequencing was costly and slow, modern gene sequenc-
ing systems can sequence an entire genome in a matter of hours to days and at
a modest cost (as low as US$ 1,000). Such genome data harbors the full amino
acid sequence of every protein the organism can produce (proteins are chains of
amino acids; with each protein being characterized by its own unique amino acid
sequence).
The proteome refers to the entire complement of proteins actually produced
by a specic cell/organism. At any given time in a specic cell, some genes
are actively ‘transcribed’, resulting in the actual synthesis of the protein they
encode. However, some genes are silent (not being transcribed). Proteomics refers
to the large-scale, systematic study of the proteome. It generally involves the
simultaneous/near-simultaneous study of hundreds, and more usually thousands
of proteins that might be present in a particular cell/organism at any given time.
Such studies rely upon ‘high throughput’ analytical techniques/processes, of which
mass spectrometry (MS) is probably best known. Proteomic studies are often
sub-categorized as:
● Functional proteomics, which aims to assign a biological function to each protein
in the proteome.
● Expression proteomics, which entails the analysis of expression of individual pro-
teins in the proteome,andhow these change in responseto stimuli, such as genetic
or environmental factors.
● Structural proteomics, in which the goal is to gain as much information as is pos-
sible relating to the 3-dimensional structure of proteome proteins.
Given that most drug targets are proteins, it becomesobvious that genomics and in
particular proteomics provide a powerful mechanism for identifying potential new
drug targets, and indeed can help in designing drugs to interact with these targets.
For example, expression proteomics can be used to identify proteins whose cellu-
lar concentration increases in a particular disease state, linking the protein to that
disease (Figure 3.6).
Once one or moresuch potential drug targets are identied,it becomes desirable to
validate the target, in other words, to conrm that the target is genuinely important
in driving the disease. For example, even if expression proteomic studies identify a
group of cellular proteins whose concentration increases in a particular disease, for
a proportion of such proteins their increased concentration may be a consequence
rather than a cause of the disease (Figure 3.6). Such proteins will not represent dis-
ease targets (although they may still prove useful in diagnosis as disease markers).
Some of the other proteins however may be causing/driving the disease (and thus
would represent legitimate drug targets).
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3.3 Drug Discovery 57
1
2
3
4
5
1
2
3
4
5
Proteome profile,
healthy tissue
Proteome profile,
diseased tissue
(a) (b)
Figure 3.6 Diagrammatic representation of a quantitative proteomics approach to target
identification. In this simplified example, the proteome consists of just five different
proteins, separated from each other in 2 dimensions by a technique known as
2-dimensional gel electrophoresis. The concentration of protein number 5 is substantially
higher in the diseased tissue sample, as compared to the healthy tissue, indicating that it
may represent a potential disease target. In reality, potential target identification is more
complex than shown below. Tissue samples analysed would generally contain thousands of
different proteins and, for example, an increase in a particular protein concentration may be
a consequence of the disease state, rather than causing it.
Target validation is essential to the successful development of an eective drug.
Validation studies normally entail the application of various disciplines/techniques,
to provide an overall, comprehensive picture of the target and its relationship to the
disease status. Biochemical studies are required to characterize the target in full, to
understand the biochemical processes underlining the disease state, and to contex-
tualize the role of the target in the disease process. Molecular biology is also often
applied to ‘knock out’ or ‘silence’ the gene which encodes the target from a cell or
indeed from whole animals. The resulting absence of the target protein provides
further insight into its normal role in the body and in the disease.
3.3.2 Lead Discovery, Validation and Optimisation
Once a satisfactory target has been discovered and validated, the next step in the
drug discovery process is to identify substances that can interact with the target and
hence modulate target activity in a way that is likely to counteract the disease. Such
substances are called leads (i.e. lead drug candidates). In most instances, leads are
molecules that inhibit target’s biological function (e.g. antibody-based inhibition of
TNF-α activity in the treatment of certain inammatory conditions; Figure 3.4), but
for some disease states target activation as opposed to inhibition is required (e.g.
insulin, when it binds to its cell surface receptor, triggers the activation of a receptor
associated enzyme).
58 3 Drug Discovery, Classification and Early Stage Development
A popular approach to lead discovery entails the screening of large libraries of
compounds for their ability to bind to the target. This requires the development of
suitable assays to measure binding. These binding assays may be based on the mea-
surement of a responsethat is triggered by binding or on using compounds labeled in
such a fashion that they can be detected e.g. dye, uorescence, enzyme, or radioac-
tive carbon-14 labelling methods.
Traditionally, discovery studies usually focused upon the screening of large
libraries of natural compounds i.e. chemical substances generally extracted from
plant or microbial sources. A variation of such blind screening approaches is the
use of synthetic chemical libraries. Large synthetic libraries of chemical substances
can now be generated using modern chemical biosynthetic approaches such as
combinatorial chemistry.
As discussed earlier, modern drug discovery has come to rely more and more on
a knowledge-based approach in which an understanding of the disease process at a
molecular level is essential. Once potential drug compounds have been identied, a
long and arduous process of drug development and evaluation ensues.
In the rational approach, focus is put on identifying the amino acid sequence
and in particular the 3D structure of potential binding sites (structural proteomics;
Section 3.3.1) with the view of predicting the structure of molecules that will give a
best-t interaction with these sites (see also Figure 3.1). To determine an accurate
3D structure requires the application of X-ray crystallography, NMR spectroscopy
or Cryo-Electron Microscopy to a puried sample of the potential target. Com-
puter modelling and simulation programmes can then be used to ‘design’ virtual
molecules that should interact with the binding sites. Finally, it is up to the organic
chemists to actually synthesise these molecules for testing.
3.4 Drug Development
Before embarking on to the wide variety of activities and studies that are required,
it is normal practice to validate the lead identied. Validation approaches include
detailed biochemical studies, which study lead binding to the target in detail, as
well as binding specicity (exploring the ability of the lead to binding to cellular
molecules other than the target – which could ultimately result in unwanted side
eects). It is also standard practice to examine structurally related compounds, par-
ticularly in the case of small-molecule drugs. There are a number of reasons for this.
It is possible that an analogous compound could actually be more potent than the
candidate compound initially identied. Secondly, by examining a broad range of
compounds, it reduces the chances of a competitor being able to bring similar drugs
to market.
At this early stage, once initial compounds have been characterised and shown
to have potential therapeutic value, it is advisable to prepare a patent application.
Patents grant legal protection to applicants to exclusively commercialise their
‘invention’ in exchange for making a detailed description of the ‘invention’ available
to the public. To obtain a patent, the subject of the invention must be novel, have
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3.5 Drug Delivery 59
utility and should advance the ‘state of the art’ beyond that which previously
existed. The standard term of protection that is granted is twenty years from the
date a patent application is led. Drug developers may apply for an extension of the
protection period to take account of time lost while conducting clinical trials and
obtaining marketing authorisations. The standard maximum extension permitted
is ve years, subject to the restriction that it does not result in a total protection
period post-authorisation of more than fteen years in Europe or fourteen years in
the US. Specic paediatric studies can add a further six months of protection.
This protection provides a vital incentive to drug development companies, as
patented medicines are usually able to command a premium price – the pre-
sumption being that the new drug is more eective than any existing alternative
therapies. The patent holder may also gain revenue by charging a licensing fee to
other companies that wish to manufacturethe drug or use it aspart of a combination
therapy. In this way, the developer has an opportunity to recoup the substantial
costs that go into drug development.
3.4.1 Manufacture and Control
While early-stage studies require only small quantities of the putative drug,
non-clinical and clinical studies will require greater drug quantities and, if the drug
is nally approved for general medical use, still greater product quantities will be
required to satisfy market demand. It is therefore essential to optimize the drug
manufacturing process as early as possible during drug development. Optimization
of the drug’s manufacturing route must take not only technical chemical issues into
account, but also regulatory requirements (e.g. likely maximum impurity levels
acceptable) and the suitability of the manufacturing scheme developed for scale-up.
The latter point is important as major manufacturing changes in later-stage
clinical evaluation can eectively invalidate earlier-stage studies and the regulatory
authorities will demand proof from the company that any manufacturing changes
introduced at any stage of the drug’s life cycle has not aected the drug properties
in any negative way. Such validation studies can be costly and dicult and, if not
undertaken to the regulator’s satisfaction, can even result in removal of the drug
from the market. As part of manufacture development, the company will begin
to develop analytical tests (for e.g. drug identity and purity) which will become
essential elements in the quality control of the drug’s eventual manufacturing
process.
3.5 Drug Delivery
A key element in the development process is to select and optimise the most appro-
priate method for delivery of the drug. Drugs may be administered by:
● injection,
● ingestion,
60 3 Drug Discovery, Classification and Early Stage Development
● inhalation or
● application to dermal or other accessible body surfaces.
Development of the most appropriate route for administration is inuenced by
a number of factors including location of the target within the body, desired speed
and duration of the therapeutic eect, and properties and characteristics of the drug
substance.
3.5.1 Location
Obviously, if you wish to treat a skin condition or infection, a preparation that can
be applied topically will be the preferred option. Similarly, inhalation will be the rst
choice if trying to treat a pulmonary or bronchial condition, such as asthma. Dermal
application will also be the rst choice for localized tissue treatments (e.g. muscle
injury), provided the drug can be adsorbed throughthe skin. However, in most other
situations it is necessary for drugs to enter the bloodstream in order for them to be
transported to their site of action. This is most commonly achieved by ingestion, or
intravenous (i.v.), intramuscular (i.m.), or subcutaneous (s.c.) injection where the
oral route is not suitable.
3.5.2 Drug Characteristics
The characteristics and properties of the drug will have a big bearing on its ability
to enter the body and reach its target, particularly with respect to its size and rel-
ative solubility in polar versus nonpolar solvents. All body uids are water-based
and are thus compatible with polar solutes (hydrophilic substances). Cells on the
other hand, are surrounded by cell membranes, which are based on a lipid bilayer
structure that creates a nonpolar core that is not very compatible with polar solutes.
Excluding injection, drugs must be able to pass through cellular tissues to enter the
body. Small molecule drugs with signicant nonpolar character (hydrophobic sub-
stances) are able to passively diuse through membranes. More polar or charged
molecules will have greater diculty in passing through the lipid bilayer but may
cross cell membranes with the assistance of appropriate transmembrane proteins
that provide channels for passive diusion or active transport. The mechanisms are
illustrated in Figure 3.7. Relative polarity will also have a signicant bearing on the
solubility of the drug and may dictate the level of ‘free’ drug circulating in the blood-
stream and the distribution pattern between various tissues. A nonpolar drug will
display slower dissolution rates and solubility in aqueous solutions, such as encoun-
tered in the gastrointestinal tract, while most of it will be bound to proteins such as
albumin when circulating in the bloodstream. Such a drug is also likely to show a
tendency to deposit in fat cells, which could reduce the eectiveness of a therapy
in obese people. Chemists may have the opportunity to manipulate the polarity of
the drug substance during development so as to favourably inuence its behaviour.
Hydrophobic character is determined by the amounts of nonpolar carbon–carbon
and carbon–hydrogen bonds that are present in a molecule. Polarity/watersolubility
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3.5 Drug Delivery 61
Transmembrane protein channel
Lipid bylayer
Non-polar core
(hydrophobic)
Polar/charged drug
Non-polar drug
Figure 3.7 Passage of drugs across cell membranes.
Cell
Endosome
Movement of drug across cell
Cell membrane
Figure 3.8 Passage of large molecule drugs across cells.
can be improved by introducing groups such as hydroxyl, carboxyl,amino, sulphate,
or phosphate into a compound. Thus the drug developer may be able to optimise the
properties of a drug substance by adding chemical groups at locations remote from
the site that is involved in interaction with the target.
Large molecule drugs have greater diculty in traversing cell membranes and
to date, most biopharmaceutical products are administered via injection (i.e.
parenterally), either intravenously (i.v.), intramuscularly (i.m.), or subcutaneously
(s.c.). However, as any diabetes suerer will attest, this is not the most pleasant
prospect when frequent administration is required. Thus, drug companies are keen
to develop alternative routes for administration of biopharmaceuticals. Although
large molecule drugs have great diculty crossing cell membranes, they can enter
many cells by a process of pinocytosis, through which the substances are enveloped
in a portion of the cell membrane. See Figure 3.8 for an illustration. Despite
this, ingestion is not a viable option for biopharmaceuticals, as they will face
denaturation by the acid environment of the stomach, and degradation by digestive
enzymes in the gut. Adsorption via the lungs has shown greater promise and the
rst such product (an inhalable insulin, tradename “Exubera”) was approved for
general medical use in 2006.
62 3 Drug Discovery, Classification and Early Stage Development
3.5.3 Speed and Duration of Therapeutic Effect
The desired speed and duration of a therapeutic eect will also inuence the
development of the nal drug product. The standard treatment of meningitis
with antibiotics is via intravenous administration so as to achieve an immediate
eect, while rapid pain relief is promoted as a feature of many common analgesic
remedies, which are invariably manufactured for oral administration. On the other
hand, treatment of blood pressure orirregular heartbeat requires the achievement of
reasonably constant levels of drug in the circulatory system. In such circumstances,
a slower release form of the drug is required, normally as an oral dosage form.
As already mentioned, the properties of the drug substance will have a bearing
on the ability to achieve this. The rate of release can be further controlled by the
way in which the drug is mixed or formulated with other ller substances, termed
excipients. A list of common excipients is shown in Table 3.2.
Ingestion remains the most popular route for drug delivery due to its convenience
and the high level of patient compliance generally attained. However, there are
potential obstacles that the drug developer may need to address. The acid environ-
ment in the stomach may prove detrimental to some drugs. Fortunately, this can
be overcome by encapsulating the drug in a protective enteric coating that only
allows it to be released when it reaches the intestine. A second disadvantage, that
may not be as easily overcome, is that once adsorbed through the intestine, the
drugs are carried rst to the liver, which is the principal organ for drug metabolism.
Consequently, there is the risk that considerable activity may be lost before the drug
reaches its site of action. This is termed the hepatic rst-pass eect.
In summary, the characteristics of the drug and the disease or condition may sug-
gest a likely route and modeof delivery. Considerable work is requiredtodevelop and
optimise an appropriate drug presentation. The suitability of any particular strategy
Table 3.2 Common excipients use in the formulation of drug products.
Excipient
Magnesium stereate Lactose
Microcrystalline cellulose Starch (corn)
Silicon dioxide Titanium dioxide
Stearic acid Sodium starch glycolate
Gelatin Talc
Sucrose Calcium stearate
Pregelatinized starch Hydroxypropyl methylcellulose
Hydroxypropyl cellulose Ethylcellulose
Calcium phosphate
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3.7 Further Reading 63
must be veried by undertaking the extensive pharmacodynamic and pharmacoki-
netic studies that form part of the preclinical and clinical trials (Chapters 4 and 5).
3.5.4 Stability
As discussed above, stability of the drug as it enters the body can have a signicant
bearing on how it is presented. The inherent stability of the drug mayalso decide the
nal form in which the drug product is presented. Because of stability limitations,
many biopharmaceutical products are presented in a freeze-dried form that is only
re-constituted with water prior to use. The stability of solid tablet forms and the type
of packaging used may be dictated by the tendency to absorb moisture. It may be
possible to reduce this hydroscopic behaviour or improve stability by using dierent
salts or combinations of excipients.
3.6 Chapter Review
This chapter looked at various ways by which drugs may be categorised, such as by
prescription status, mode of action, physical properties and therapeutic use. Basic
strategies used in the discovery of drugs were examined. These ranged from simple
observations to the collection of traditional cures and screening of animal and plant
substances to today’s research-based strategies that rely ongaining an understanding
of disease processes at a molecular level. Finally, the chapter considered the drug
development process in terms of choosing an appropriate drug form and method of
delivery.
3.7 Further Reading
Drug Discovery and Development
Hughes, J.P. et al. (2011) Principles of early drug discovery. British Journal
of Pharmacology, 162, 1239–1249.
Sowjanya, K. and Girish, C. (2019) Structural genomics in drug discovery: an overview.
Journal of Pharmacology & Pharmacotherapeutics, 10 (1), 1–6.
Puhl, A.C. et al. (2019) New targets for HIV drug discovery. Drug Discovery Today, 24
(5), 1139–1147.
Hedl, T.J. et al. (2019) Proteomics approaches for biomarker and drug target discovery
in ALS and FTD. Frontiers in Neuroscience, 13, Article Number: 548. https://doi.org/
10.3389/fnins.2019.00548.
An overview of the drug development process: https://www.fda.gov/patients/learn-
about-drug-and-device-approvals/drug-development-process
64 3 Drug Discovery, Classification and Early Stage Development
Patenting
The United States Patent and Trademarks oce: https://www.uspto.gov
The European patent oce: https://www.epo.org/index.html
Patenting versus exclusivity: https://www.fda.gov/drugs/development-approval-
process-drugs/frequently-asked-questions-patents-and-exclusivity
ATC
ATC Code: https://www.whocc.no
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