Download Topic guide 9.4: The role of biologically active molecules in

Unit 9: Medicinal Chemistry
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The role of biologically
active molecules in
biochemical systems
In this section, you will read about several important drug molecules and
their mechanism of action in biological systems. Some of them (for example,
penicillin-based antibiotics) have already appeared as case studies in the
context of other aspects of medicinal chemistry in Topic guides 9.1–9.3.
You will also revisit some of the ideas about drug toxicity encountered in
Topic guide 9.2, and learn how toxicity is assessed.
On successful completion of this topic you will:
•• understand the role of biologically active molecules in biochemical
systems (LO4).
To achieve a Pass in this unit you need to show that you can:
•• discuss the development and role of selected biologically active
molecules (4.1)
•• explain clinical toxicological terms citing suitable examples (4.2)
•• explain the principles of clinical toxicity (4.3).
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Unit 9: Medicinal Chemistry
1The development of selected drug molecules
Penicillins
Figure 9.4.1: Penicillin G, the
molecule extracted from the
Penicillin mould, has R = C6H5–CH2–.
The β-lactam ring is circled.
H
H
N
R
O
S
O
N
CH3
CH3
COOH
The penicillin group of molecules are antibiotics used to treat a range of bacterial
infections.
The story of the accidental discovery of the antibiotic nature of penicillin is well
known – a Petri dish used to culture Staphylococcus was accidently contaminated
with Penicillium mould. This produces the molecule we now call penicillin as a
secondary metabolite.
All penicillins have the core structure shown in Figure 9.4.1; the nature of the
R group is different for each member of the group.
Mode of action
Penicillins act on bacteria by interfering with the construction of the bacterial
cell wall. This structure consists of a network of polymer chains (known as
peptidoglycan, and consisting of amino acids and sugars bonded together). One
of the key steps in the completion of this strong network is the cross-linking of
the peptidoglycan chains. This occurs when two short peptide chains are linked
by a transpeptidase enzyme (see Figure 9.4.2). If this process of cross-linking is
prevented, the cell wall is therefore weakened and cannot withstand the pressure
within the cell, which therefore bursts, killing the bacterium.
O
Figure 9.4.2: The structure of the
peptidoglycan network present
in bacterial cell walls. The crosslink formed by the transpeptidase
enzyme is shown in the diagram.
O
O
Peptide chains
Key terms
Secondary metabolite: A molecule
produced by a living organism that
has no known role in its biochemistry.
These substances are frequently
used as possible starting points from
which to develop lead compounds.
Nucleophile: Group of atoms with
a lone pair of electrons that can be
donated to an electron deficient
atom.
Polysaccharide chain
O
O
O
O
O
O
This cross-link is formed by the action
of the transpeptidase enzyme
O
O
O
O
O
O
O
O
O
O
Penicillin inhibits the transpeptidase enzyme, which forms the cross-links, by
permanently binding to the active site. The key structural feature of the penicillin
molecule is the β-lactam ring. This is attacked by nucleophiles, such as –OH
groups, and attaches to a serine residue in the active site.
9.4: The role of biologically active molecules in biochemical systems
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Unit 9: Medicinal Chemistry
Figure 9.4.3: The mechanism by
which the β-lactam ring is opened
up, involving nucleophilic attack
by the OH of a serine residue.
proton
transfer
N
O
–
O
OH
ser
N
hydrolysis
–O
+
OH
ser
O
N
H
+
NH
O
O
ser
NH
O
OH
+ OH
ser
ser
Activity
Research how chemists have
developed new penicillins to
overcome bacterial resistance.
Explain how the structural features of
these adapted molecules achieve this.
Key terms
Hydrolysis: A reaction in which a
molecule is broken down by the
action of water.
Peptide (or oligopeptide):
Molecules consisting of short chains
of amino acid residues.
Figure 9.4.4: The angiotensin-converting
enzyme catalyses the formation of
angiotensin II from angiotensin I,
by removing two amino acids.
ACE inhibitors
angiotensin I
0000000000
ACE
angiotensin II
00000000 + 00
hypertension
This opens up the β-lactam ring and the serine residue is later regenerated by a
final hydrolysis step (see Figure 9.4.3).
Antibiotic resistance
Some bacteria possess a gene that enables them to synthesise an enzyme called
penicillinase or β-lactamase. This enzyme causes the hydrolysis of the amide group
in the β-lactam ring, inactivating penicillin. These bacteria are therefore resistant
to penicillin.
ACE inhibitors
Angiotensin (more correctly known as angiotensin II) is a small peptide hormone
involved in the system that regulates blood pressure. When present in raised
concentrations it causes high blood pressure (hypertension) which, if left
untreated, has serious health consequences.
Angiotensin II is formed by the removal of two amino acids from the inactive
compound angiotensin I (see Figure 9.4.4). The reaction is catalysed by the
angiotensin-converting enzyme (ACE). Inhibition of this enzyme will therefore
decrease the rate of formation of angiotensin II.
Development of ACE inhibitors
The development of this group of drug molecules is a good example of how
structure-activity relationships can be used in the development of an effective
drug.
The importance of ACE first came to be realised from a study of the venom of
the Brazilian Pit Viper; its fatal effects were due to a massive and very rapid drop
in blood pressure caused by inhibition of ACE. However, this inhibitor was itself
a peptide. This meant it would not be suitable as an orally-administered drug
because it would be rapidly hydrolysed in the stomach.
This peptide had a proline amino acid at the carboxyl end of the peptide chain:
H2N–Glu–Trp–Pro–Arg–Pro–Gln–Ile–Pro–Pro–COOH
Researchers created thousands of molecules based on proline and evaluated their
effectiveness using animal-based tests.
Initially, a succinyl derivative was found to be the most effective inhibitor and
became the lead compound (see Figure 9.4.5).
Figure 9.5.5: The structures of
(a) proline (b) the general structure
of the proline derivatives that were
investigated (c) succinyl proline.
= a single amino acid
(a)
(b)
HN
(c)
R
N
OH
9.4: The role of biologically active molecules in biochemical systems
HO
OH
N
OH
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Unit 9: Medicinal Chemistry
Further screening suggested that adding a methyl group in the 2-position of the
succinyl group further increased inhibitory strength by a factor of about 15. This
created a chiral carbon at the 2 position and only one enantiomer showed the
enhanced inhibitory effect.
Figure 9.4.6: The structure of captopril.
The presence of the methyl and SH group
increases the inhibitory effect enormously
compared to succinyl proline.
HS
N
O
O
OH
At this point, structure-activity relationships were used to optimise the lead
compound. ACE functions in the presence of zinc ions, which were thought to
complex to the penultimate amino acid of the substrate. Replacing the second
carboxyl group from succinic acid with an SH (sulfhydryl) group would increase
the strength of binding to this zinc ion.
The result of this drug development process was to produce the first marketable
ACE inhibitor, captopril (see Figure 9.4.6).
Figure 9.4.7 shows the binding of captopril to the active site of ACE.
CH3
H
S
CH2
CH
C
O
:
:
Figure 9.4.7: Captopril binds to the
active site of ACE using covalent bonds
to the Zn2 ion as well as hydrogen
bonding and ionic bonding.
N
C
O–
+
NH3
O
H
Zn2+
N
Activity
Describe the similarities and
differences in the structures of
captopril and succinyl proline.
Take it further
The Wikipedia entry for ACE inhibitors, found at
http://en.wikipedia.org/wiki/ACE_inhibitors_drug_design
tells the story of the development of ACE and gives an excellent insight into the practical
application of structure-activity relationships.
Anticancer agents
Cancer, the uncontrolled division of cells, is not a single disease but a spectrum of
more than 100 identifiable types, usually specific to a particular tissue or organ.
The different types of cancer respond in different ways to drugs and, as a result,
the number of anticancer drugs that have been developed is huge.
The majority of drug treatments are designed to disrupt the processes that occur
during cell division. This makes it very difficult to achieve high levels of potency
without also observing very significant toxic effects, since the drugs will also
disrupt the activities of other cells that undergo rapid cell division such as skin
cells, blood cells in the bone marrow and cells lining the gastrointestinal tract,
creating unpleasant and potentially fatal side effects.
More recently, drugs have been developed that target specific receptors present
in cancer cells and offer hope for effective treatment without the risk of toxicity to
normal cells.
A discussion of the full range of anticancer agents is beyond the scope of this
publication; two case studies will illustrate some of the key principles.
9.4: The role of biologically active molecules in biochemical systems
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Unit 9: Medicinal Chemistry
Methotrexate
Methotrexate disrupts the synthesis of the base thymine, which is one of the four
bases present in the nucleotides that make up DNA.
One of the key steps in the synthesis of thymine is methylation of a uracil base (see
Figure 9.4.8). This methylation step requires the involvement of a molecule called
tetrahydrofolate (shown in the diagram as tetrahydrofolic acid, THF), which is
eventually oxidised to dihydrofolate (shown as dihydrofolic acid, DHF). In order for
synthesis of thymine to continue, the tetrahydrofolate must be regenerated from
dihydrofolate by the action of dihydroreductase.
Dihydroreductase
Figure 9.4.8: Synthesis of a uracil
group requires regeneration of THF.
THF
DHF
Methylene THF
+ CH3
N
O
CH3
N
N
O
Uracil monophosphate
N
Thymine monophosphate
It is this latter step that is targeted by methotrexate – it acts as a competitive
inhibitor of dihydroreductase. It is easy to see why when you examine the
structures of methotrexate and DHF in Figure 9.4.9.
Within a constant regeneration of DHF, the synthesis of thymine will cease.
Figure 9.4.9: The structures of
dihydrofolate (top) and methotrexate
(bottom) show how similar in
structure the competitive inhibitor
is to that of the normal substrate.
O
O
O
N
N
H
N
N
N
H
H
N
NH3
Cl
Pt
Cl
NH3
O
H
O
OH
O
OH
N
N
N
OH
N
NH2
Figure 9.4.10: The structure of cisplatin.
The ‘cis’ prefix in the structure indicates
that the chloride ligands are adjacent to
each other on the same side of the Pt ion.
OH
N
H
O
N
Cisplatin
Cisplatin is an example of a transition metal complex, in which small molecules
and ions (in this case ammonia and chloride ions) form dative covalent bonds to a
central transition metal ion (platinum) – see Figure 9.4.10.
It was discovered to have anticancer properties in the 1960s. The mechanism of
its action is now known to involve disrupting both replication and transcription
of DNA.
9.4: The role of biologically active molecules in biochemical systems
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Unit 9: Medicinal Chemistry
Activity
Write a short report describing the
mechanism of action of cisplatin,
making it clear how the mechanism
relates to its structure.
Take it further
More details of the action of cisplatin can be found at
http://chemcases.com/cisplat/cisplat12.htm, including some excellent graphics showing the
disrupted structure.
Antiviral drugs
Viruses, which cause diseases such as influenza (‘flu), chickenpox and AIDS, have
proved far more difficult to target by drug therapy than bacterial agents. The main
reason for this is that viruses use their host cell’s metabolism to produce all the
molecules required for replication and assembly of new viruses, hence it is very
difficult to target specific metabolic reactions of viruses without also affecting the
host’s metabolism.
Vaccines
Vaccines provide a way of protection against infection by viruses, by stimulating
the body’s immune system to recognise virus particles.
Case study
Viruses mutate rapidly, and this is particularly significant in the production of seasonal ‘flu vaccine,
often taken by at-risk groups of patients at the onset of each winter. Each year in February, the
World Health Organization (WHO) recommends the three strains of ‘flu virus that should be
included in that year’s vaccine. From this point, small laboratories, such as that run in the UK by the
National Institute for Biological Standards and Control, create a ‘seed’ vaccine strain. They incubate
small amounts of the virus mixture in a hen’s egg, creating a sample large enough to work with.
Surface proteins from the viruses are attached to a harmless virus particle to create the seed
vaccine strain. From this point it may take three to five months to produce and market sufficient
quantities of the vaccine to meet demands. Clearly the time frame of this process is very different
from that of the development of a new drug; clinical safety trials must also be carried out within
the same time frame.
1 How do you think the WHO decides which strains to include in the annual vaccine?
2 Why is it difficult for the producers of vaccines to produce vaccines rapidly enough to combat
pandemics (such as the swine flu pandemic in 2009)?
Activity
Research the mechanism by which
oseltamivir prevents the infection of
host cells and write a short report to
explain it.
Tamiflu (oseltamivir)
Whereas vaccines make use of the body’s immune system, antiviral drugs work by
targeting the activity of proteins specific to viruses. One such protein is known as
viral neuraminidase, which has a role in allowing the virus to infect host cells.
Antiretroviral drugs
Link
The structure of DNA is covered
in detail in Unit 1: Biochemistry
of Macromolecules and Metabolic
Pathways.
The influenza virus is able to be replicated and assembled because its RNA
genome is able to be replicated and transcribed in the host cell’s nucleus (usually
it is only DNA that undergoes these processes). Some viruses have a different
mechanism of replication. This is known as reverse transcription, whereby the viral
RNA is transcribed into a DNA molecule that then directs protein synthesis in the
conventional way.
9.4: The role of biologically active molecules in biochemical systems
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Unit 9: Medicinal Chemistry
Take it further
If you are unfamiliar with the structure and function of DNA, and particularly the processes of
replication and transcription referred to in this section, there are some useful animations with
descriptions available at websites such as www.johnkyrk.com/DNAanatomy.html.
These types of viruses, known as retroviruses, can be treated by antiretroviral
drugs that target the stages in this process.
AZT (azidothymidine, zidovudine)
Figure 9.4.11: The structures of AZT
(top) and thymidine (bottom) show
how similar the two molecules are.
O
NH
HO
–
O
N
O
AZT acts as a competitive inhibitor of the reverse transcriptase molecule because
it bonds strongly to one of the active sites of the enzyme. As a result, reverse
transcription is stopped and the virus is inactivated.
Nitric oxide
+
N=N=N
O
H3C
HO
The nucleotides in DNA consist of three groups: a phosphate, sugar and a base.
AZT has a structure similar to a naturally occurring combination of sugar and
base known as thymidine – it differs only by having an azido (–N=N+=N−) group
attached in place of a hydroxyl group (see Figure 9.4.11).
NH
O
OH
N
O
Nitric oxide (NO) is produced by all cells in the body and has a range of functions,
mostly connected with signalling between cells; being so small it rapidly diffuses
across cell membranes and within the cytoplasm.
There specifically, it has a direct role in neurotransmission. One important effect is
to cause dilation of blood vessels (vasodilation); nitric oxide stimulates the enzyme
guanylyl cyclase to produce cyclic GMP (guanosine monophosphate) – another
important signalling molecule that causes smooth muscle fibres to relax.
Because of this effect, it has important medical uses (and it must surely have the
simplest structure of any drug molecule!) and can be used directly in emergency
treatment of heart failure.
In cases of severe angina (chest pains), nitroglycerin is given, which reacts in the
body to form NO in situ.
Portfolio activity (4.1)
Research a drug molecule, which could be one of the examples from the activity sections in this
topic guide. Write a detailed report describing:
•• the disease or condition against which it acts, including any significant biochemical details
•• how the drug was discovered and developed
•• the mechanism of the drug’s action.
9.4: The role of biologically active molecules in biochemical systems
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Unit 9: Medicinal Chemistry
2 Clinical toxicology
Toxicology is the study of the toxic effects of substances; clinical and preclinical
tests are used to assess the toxicity of drugs and the same principles are used to
assess toxicity of other chemical substances.
Toxicity
You met the concept of toxicity (the ability of a substance to cause harm to a
person) in Topic guide 9.2.
The toxic effects can be short term or long term; hence several types of toxicity
need to be considered.
Acute toxicity
This is toxicity due to the effects of a single exposure to a substance, or several
exposures over a short period (for example, 24 hours).
A measurement of the acute toxicity of a drug must be made before proceeding
to clinical trials – this is done by carrying out studies in at least two animal species.
These studies will lead to a calculation of the LD50 value for a drug, which is the
dose required to cause mortality in 50% of a group of test animals (frequently rats
or mice).
The LD50 is expressed as the number of grams of a substance per kilogram of the
animal being tested.
Some values of LD50 for a range of drugs that have been discussed in this unit are
given in Table 9.4.1 – the smaller the number, the more toxic the substance.
Table 9.4.1: Table of some LD50 values.
Activity
Use research to find LD50 values for
three other drugs encountered in
this unit. Compare the values from
different sources – what do you
notice?
Drug
LD50 g/kg–1 for mice, oral route of administration
paracetamol
1.9
captopril
3.1
AZT
3.0
methotrexate
0.13
For comparison, the Botulinum toxin type A (used in cosmetic treatment) has an
LD50 value of around 5 × 10–8 g/kg–1.
There is considerable controversy about how valid these animal-based LD50
values are in predicting the acute toxicity of a drug in humans, as well as the
ethical issues raised by the use of animals in this way.
Chronic toxicity
This is toxicity due to the effect of repeated or continuous exposure, sometimes
lasting for the entire life of the exposed organism (effects of exposure of less than
one year are sometimes referred to as sub-chronic toxicity).
Unlike acute toxicity data, no single measure of chronic toxicity is used; the
research data will highlight specific issues of toxicity to certain tissues or systems.
Again, some of the data will be obtained from animal studies but clinical trials will
also monitor the chronic effect of drug treatment if this is appropriate.
9.4: The role of biologically active molecules in biochemical systems
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Unit 9: Medicinal Chemistry
Take it further
More details of the testing regimen used to establish chronic toxicity and other long-term effects of
drugs can be found at www.meti.go.jp/english/information/data/TESTmammal.html.
A database of toxicological studies for a wide range of drugs can be accessed at
http://toxnet.nlm.nih.gov/cgi-bin/sis/htmlgen?HSDB.
Specific chronic tests
Table 9.4.2 summarises some specific chronic tests.
Table 9.4.2: Specific chronic tests.
Teratogenic toxicity tests
To assess the potential of the drug to cause foetal abnormality
(teratogenicity)
Reproductive toxicity
tests
To assess the potential of the drug to cause fertility problems or
developmental problems in offspring
Mutagenicity
To assess the potential of the drug to cause genetic mutations
Activity
A well-known example of a drug that was widely used before its teratogenicity was fully
appreciated was thalidomide. Research the effects that this drug had and the mechanism of its
teratogenic effect.
3 Clinical toxicity
Faced with the toxicological information about the toxicity of a drug, what
principles will a medical practitioner use to manage or evaluate the use of a drug
in a clinical setting?
Risk assessment
This approach is common to any setting where a hazardous substance or process
is being used. Essentially it seeks to answer three key questions:
•• What is the hazard which might be encountered?
•• What is the risk of that hazard occurring?
•• How can the risk be reduced or managed?
In the case of a drug, the possible toxic effects (as covered above) will be the
hazard that may be encountered – the toxicological studies will give some
quantitative information about the probability of harm occurring.
The management of the risk is the point at which the practitioner will need to take
decisions – about dosage, route of administration, length of treatment, and so on.
In many cases there will be recommendations available on databases but, in some
cases, the practitioner’s own experience and judgement will be called into play.
9.4: The role of biologically active molecules in biochemical systems
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Unit 9: Medicinal Chemistry
Risk-benefit analysis
In many cases a decision may need to be taken about whether the benefit of the
drug treatment outweighs the risks of harm that may result from the treatment.
In the case of life-threatening diseases such as cancer, where the risks of harm
from treatment are high and the potential for benefit may be measured only
in terms of a short extension of life, such analyses may be difficult for both
practitioner and patient and may raise difficult ethical issues about the rights of
patients to access such treatment against the judgement of the practitioner.
Portfolio activity (4.2, 4.3)
Choose a drug (or drugs) that you have been reading about in this topic guide. Use this drug (or
these drugs) to illustrate the principles of clinical toxicity. In your report:
•• provide information about the acute and chronic toxicity of the drug, clearly distinguishing
between the two terms
•• explain what data would be needed to assess teratogenic toxicity, reproductive toxicity and
mutagenicity, providing any data for your chosen drug
•• include a risk-benefit analysis of the use of the drug by comparing the known hazards with the
benefits it can provide.
Checklist
At the end of these two sections you will understand the following ideas:
 acute toxicity is measured by the LD50 value
 assessing chronic toxicity requires detailed long-term studies
 these studies will also look at the potential for causing birth defects, reproductive problems
and genetic mutations
 in taking decisions about the use of a drug, a risk assessment should be carried out and the
benefits considered against the risks of identified hazards.
Acknowledgements
The publisher would like to thank the following for their kind permission to reproduce their
photographs:
Shutterstock.com: isak55
All other images © Pearson Education
In some instances we have been unable to trace the owners of copyright material, and we would
appreciate any information that would enable us to do so.
About the author
David Goodfellow studied Natural Sciences at Cambridge and spent 20 years teaching A-level
Chemistry in a sixth-form college. He was lead developer for the OCR AS Science in 2008 and for
several years was chief examiner for the course. He now works as a freelance writer and examiner
alongside part-time work as a teacher. Publications include a textbook for the AS Science course,
teaching materials to accompany Chemistry GCSE courses and contributions to textbooks for BTEC
First Applied Science.
9.4: The role of biologically active molecules in biochemical systems
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