B5 Therapeutic Classes of Drugs: Folate

Antagonists (Mode of Action and Infections)

B31BFI Bacterial and Fungal Infections
Dr Felicity Rose (UK)
Dr Chee-Mun Fang

Objectives
At the end of this part of the lecture you should .
Understand and be able to describe how bacteria utilise folate
and how this differs for mammalian cells.
Understand how antibiotics can selectively interfere with this
process in bacteria.
Understand the microbiology of infections that this class of
antibiotics are used to treat using urinary tract infection as an
example.
Describe the mechanisms of resistance to folate antagonists.
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B31BFI Bacterial and Fungal Infections

Classes of Antibiotics

Bacterial Cell

KEY
No brackets anti-bacterial agents
() anti-mycobacterial agents
[] anti-fungal agents
{} anti-protozoal agents
From Hugo and Russell Pharmaceutical Microbiology 8th Edition Chapter 12, Figure 12.1: p 201
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B31BFI Bacterial and Fungal Infections

Folate
Folate (aka folic acid & vitamin B9) is
an important co-factor in all living
cells.
Folate is not biologically active itself,
but it provides important molecules
upon its metabolism.
In mammals , it is important during
phases of rapid cell division, such as
in infancy and pregnancy.
The reduced form, tetrahydrofolate
(THF) functions as a carrier of single
carbon fragments used synthesise
adenine, guanine, thymine &
methionine for DNA synthesis.

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Folate synthesis in bacteria:

1. Synthesis of dihydropteroic
acid from one molecule of
pteridine & one molecule of paminobenzoic acid (PABA).
2. Glutamic acid then added to
form dihydrofolic acid.
3. Dihydrofolic acid reduced to
tetrahydrofolate by
dihydrofolate reductase.

B31BFI Bacterial and Fungal Infections

Folate Utilization
Folate utilization in bacteria
and mammalian cells is
fundamentally different.
Bacteria (& protozoa) are
unable to take up exogenous
folate and so they must
synthesise it themselves.
Mammalian cells do not
synthesise dihydrofolate, and
so it is taken up through the
diet and converted to
tetrahydrofolate in the liver.
From Hugo and Russell Pharmaceutical Microbiology 8th Edition Chapter 12, Figure 12.7: p 213
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B31BFI Bacterial and Fungal Infections

Folate Metabolism
Addition 2 protons

Addition C from
conversion of serine to
glycine

H
H

DHFR

Folic acid
NADPH+H+

CH2

DHFR

THF

DHF
NADP+

NADPH+H+

Ser

Gly

Loss of
methyl
group
to
dUMP

5,10-methylene
THF

NADP+

SHT

Conversion of tetrahydrofolate back to dihydrofolic

acid the reaction repeats
DHF - dihydrofolate
DHFR - dihydrofolate reductase
THF - tetrahydrofolate reductase
SHT serine hydroxymethyl transferase
dUMP - deoxyuridine monophosphate
dTMP - deoxythymidine monophosphate
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dUMP
H3 C

DNA synthesis
B31BFI Bacterial and Fungal Infections

ThymidylateSynthetase

dTMP
6

Antibiotics:
sulfonamides
Sulphonamides (aka sulfo drugs) are structural
analogues of pABA.
Inhibit incorporation of pABA into
dihydropteroic acid.
Their importance in clinical use has decreased
due to bacterial resistance.
New antibiotics are also generally more active
and less toxic further supporting their
replacement.

The first step in the synthesis of folate is the

formation of dihydropteroic acid from one
molecule of pteridine & one molecule of paminobenzoic acid (PABA).

Example drugs in this class are

sulfamethoxazole and sulfadiazine.

Sulphonamides inhibit this reaction by acting

as pABA analogues.

Sulfamethoxazole can be administrated with trimethoprim as they exhibit synergistic

activity (co-trimoxazole).
Co-trimoxazole only prescribed for treating respiratory and urinary tract infections
where there is good evidence for its use.
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B31BFI Bacterial and Fungal Infections

Antibiotics:
trimethoprim
Trimethoprim is a selective inhibitor of bacterial
DHFR.
Acts as a structural analogue of dihydropteroic
acid portion of the dihydrofolate substrate.
Trimethoprim fits into active site of DHFR
normally occupied by DHF forming strong
hydrogen bonds with amino acid residues &
water molecules lining the site.

Trimethoprim
Trimethoprim is a selective inhibitor of
bacterial DHFR by acting as a structural
analogue of dihydropteroic acid portion (blue
box below) of the dihydrofolate substrate.

Trimethoprim can be used alone for urinary and

respiratory tract infections amongst others.

Gram positive
MRSA

Staphylococci

Streptococci

Gram negative
Ef

Anaerobes

Coliforms

Resp

Pyo

ESBL

Atypicals

Trimethoprim

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B31BFI Bacterial and Fungal Infections

Urinary Tract Infection

Urinary tract infection (UTI) is an infection that
affects the urinary tract.
An infection of the lower urinary tract (bladder) is
known as a cystitis. Symptoms include:
painful urination and either frequent urination
or the urge to urinate (or both).
An infection of the upper urinary tract (kidneys) is
known as pyelonephritis. Symptoms include:
fever and lower back pain in addition to the
symptoms of a lower UTI.
The urinary organ system (the kidneys, the
ureters, the bladder and the urethra)
produces, stores, and eliminates urine from
the body. The urinary tract in males and
females are similar except that the urethra
in women is shorter making women more
susceptible to infection.

The main causative micro-organism of both upper

and lower UTI is Escherichia coli (can be caused by
other bacteria, viruses or fungi but this is rare).
UTI occur more commonly in women than men and
recurrences are common.

Image taken from http://universalium.academic.ru

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B31BFI Bacterial and Fungal Infections

Escherichia coli
Escherichia coli is a gram-negative, coliform, rodshaped enteric bacteria.
They form part of the natural flora of mammalian
gastrointestinal tract.
E. coli often causes UTI (from entry from the
intestinal tract to the urinary tract) and
gastroenteritis (from eating infected food
contaminated with pathogenic strains).
Pathogenicity mediated by three antigens:
Polysaccharide shell.
O-polysaccharide.
Lipid A (causes toxic shock).
Picture taken from Bauman Microbiology with
Diseases by Taxonomy; 3rd Edition, Chapter 20:
p 580.
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Escherichia coli cont

Virulence factors include:
Outer capsule - protection from phagocytosis.
Fimbriae and adhesins attachment to
epithelial cells.
T3SS needle complex

Exotoxins
Iron-binding proteins (siderophores) acquire
iron needed for growth.
Haemolysins release iron from blood cells.
Type III secretion system builds and
insertion needle to inject proteins into host
cells.

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Antibiotic therapy for UTI

Gram positive
Pyelonephritis /
Upper UTI
Trimethoprim

MRSA

Staphylococci

Streptococci

Ef

Gram negative
Anaerobes

Coliforms

Resp

Pyo

ESBL

Atypicals

Nitrofurantoin
Gentamicin
Co-amoxiclav
Cefuroxime
Ertapenem

Trimethoprim usual choice for simple UTI (nitrofurantoin 2nd line)

Urosepsis/pyelonephritis IV co-amoxiclav /cefuroxime or gentamicin

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Resistance to trimethoprim

Resistance to trimethoprim occurs

as a result of:

Over-production of DHFR by
bacteria.

Acquisition of the dfr gene

encoding a resistant form of
the enzyme most commonly
DHFR type I, II or V (amongst
enterobacteria).

This image shows antibiotic sensitivity testing of E. coli isolated from a urine sample from a patient with a UTI. It
can be seen that the bacterium is resistant to trimethoprim (W; blue arrow), the first line antibiotic for UTI, but
sensitive to nitrofurantoin (F; red arrow), the second line antibiotic for UTI).
Watch the video in Moodle that shows how the causative microorganism of a UTI is identified in a clinical
microbiology laboratory and antibiotic sensitivity determined.
Image used with permission from the Clinical Microbiology Laboratories, University Hospitals Trust, Nottingham.
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Key Points
Folate provides small carbon fragments (via an intermediate tetrahydrofolate)
essential for DNA synthesis.
Bacteria synthesise folate; mammals take up folate in the diet.
Sulphonamides are structural analogues of pABA; inhibit incorporation of pABA
into dihydropteroic acid.
Reduced clinical use of sulphonamides due to bacterial resistance.
Trimethoprim fits into active site of DHFR normally occupied by DHF.
Trimethoprim predominantly used to treat UTIs (mainly caused by E. coli)
Resistance to trimethoprim arises due to increased production of DHFR or a
resistant form of the enzyme.
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References and Further Reading

Bauman Third Edition

Chapter 20

Hugo and Russell

Eighth Edition
Chapters 7, 11 -13

BNF latest edition

5. Infections

Thanks to Tim Hills Lead Pharmacist Antimicrobials and Infection Control, Nottingham University
Hospitals for the antibiotic spectrum charts.
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