Antibiotics and Antifungals (13.03.2020)

Question 1

General Information about bacteria

Answer 1

• Single-cell microorganisms - cell wall & cell membrane
• An entire phylogenetic domain
• ~ 1/3 are pathogenic

Question 2

Membrane characteristics of different bacteria + name an example for each of the three categories

Answer 2

Gram Positive Bacteria

• Prominent peptidoglycan cell wall, cell membrane below
• E.g. Staphylococcus Aureus

Gram Negative Bacteria

• Outer membrane with lipopolysaccharide (then a peptidoglycan layer and another inner membrane)
• E.g. Escherichia Coli

Mycolic Bacteria

• Outer mycolic acid layer (reasonable peptidoglycan layer below, they don’t fall under g+ or g-)
• E.g. Mycobacterium Tuberculosis

Question 3

Prokaryotic protein synthesis

Answer 3

1. Nucleic Acid Synthesis
   a) Dihydropteroate (DHOp)
   - Produced from paraaminobenzoate (PABA)
   - Converted into dihydrofolate (DHF)
   b) Tetrahydrofolate (THF)
   - Produced from DHF by DHF reductase
   - THF -> Important in DNA synthesis
2. DNA replication
   - DNA gyrase (same as topoisomerase, uses tension)
   - Topoisomerase - releases tension
3. RNA synthesis
   - RNA polymerase
   - Produces RNA from DNA template
   - Differ from eukaryotic RNA polymerase
4. Protein synthesis
   - Ribosomes
   - Produce protein from RNA templates
   - Differ from eukaryotic ribosomes

Question 4

Ribosomes in eukaryotes and prokaryotes

Answer 4

P: 30-50s

E: 40-60s

-> protein synthesis

Question 5

Nucleic acid synthesis in baacteria

Answer 5

1. PABA

(via DHOp synthase) ->

2. Dihydropteroate DHOp
   - >
3. DHF

(via DHF reductaasae) ->

4. THF (Tetrahydrofolate)

Question 6

DNA replication in bacteria

Answer 6

• uses DNA gyrase

- it is a topoisomerase -> releases tension

Question 7

RNA synthesis in bacteria

Answer 7

• RNA polymerase -> different in eukaryotes

- makes RNA from DNA template

Question 8

Bacterial protein synthesis inhibitors - where do they interfere?

Answer 8

• Sulphonamindes -> inhibit DHOp synthase
• Trimethoprim -> inhibits DHF reductase
• Fluoroquinolones inhibit DNA gyrase and topoisomerase IV
• rifamycins -> inhibit bacterial RNA polymerase
• Macrolides, Aminoglucosides, Chloramphenicol and Tetracyclines inhibit bacterial ribosomes.

Question 9

Rifamycins

Answer 9

• only used to treat TB

- The rifamycins (e.g. Rifampicin) inhibits bacterial RNA polymerase

Question 10

Sulphonamides

Answer 10

• Sulphonamides inhibit DHOp synthase (nucleic acid synthesis)
• not much used anymore

Question 11

Trimethoprim

Answer 11

• inhibits DHF reductase

- used sometimes but in combination moslty

Question 12

Fluoroquinolones

Answer 12

• inhibit DNA gyrase and topoisomerase IV

- e.g. Ciprofloxacin

Question 13

Macrolides

Answer 13

• e.g. erythromycin

- inhibit prokaryotic ribosomes

Question 14

Which drugs inhibit prokaryotic ribosomes?

Answer 14

Aminoglycosides (e.g. Gentamicin)
Chloramphenicol
Macrolides (e.g. Erythromycin)
Tetracyclines

Question 15

Which drugs inhibit bacterial intracellular processes?

Answer 15

Sulphonamides
Tetracyclines
Trimethoprim
Fluoroquinonoles
Rifaamycins
Aminoglycosides (e.g. Gentamicin)
Chloramphenicol
Macrolides (e.g. Erythromycin)
Tetracyclines

Question 16

Bacterial Wall synthesis

Answer 16

PtG synthesis -> transportation -> incorporation

1. Peptidoglycan (PtG) synthesis
   - A pentapeptide is created on N-acetyl muramic acid (NAM)
   - N-acetyl glucosamine (NAG) associates with NAM forming PtG
2. PtG transportation
   - PtG is transported across the membrane by bactoprenol
3. PtG incorporation
   - PtG is incorporated into the cell wall when transpeptidase enzyme cross-links PtG pentapeptides

PtG is the most important component of the cell wall.

Question 17

List the antibiotics that interfere with the bacterial cell wall and where they interfere

Answer 17

1. PtG synthesis
   - Glycopeptides (e.g. Vancomycin) bind to the pentapeptide preventing PtG synthesis
2. PtG transportation
   - Bacitracin inhibits bactoprenol regeneration preventing PtG transportation
3. PtG incorporation
   - beta-lactams bind covalently to transpeptidase inhibiting PtG incorporation into cell wall
   - beta-lactams include:
   - Carbapenems
   - Cephalosporins
   - Penicillins
4. Cell wall stability
   - Lipopeptide - (e.g. daptomycin) disrupt Gram +ve cell membranes
   - Polymyxins - binds to LPS & disrupts Gram -ve cell membranes

Question 18

Name different beta lactams

Answer 18

• penicillins
• carbapenems
• cephalosporins

Question 19

Simply list the antibiotics that interfere with the bacterial cell wall

Answer 19

• beta lactams (carbapenems, penicillins, cephalosporins)
• glycopeptides
• bacitracin
• lipopeptide
• polymyxins

Question 20

What do glycopeptides do?

Answer 20

Glycopeptides (e.g. Vancomycin) bind to the pentapeptide preventing PtG synthesis

Question 21

What does bacitracin do?

Answer 21

Bacitracin inhibits bactoprenol regeneration preventing PtG transportation (bactoprenol transports PtG across the membrane)

Question 22

What do beta-lactams do?

Answer 22

• bind covalently to transpeptidase
• inhibit PtG incorporation in the cell wall
• transpeptidase enzyme cross-links PtG pentapeptides

Question 23

What do lipopeptides do?

Answer 23

• disrupt gram+ve cell membranes
• interfere with cell wall stability
• e.g. daptomycin

Question 24

What do polymyxins do?

Answer 24

• interfere with cell wall stability

- bind to LPS and disrupt gram-ve cell membranes

Question 25

Why is AB resistance a threat?

Answer 25

- ‘CATASTROPHIC’, ‘APOCALYPTIC’, ‘AS BIG A RISK AS TERRORISM’ UK Chief Medical Officer - ~ 70% of bacteria developed resistance - 25000 yearly death rate - Europe & US

Question 26

What are the causes of AB resistance?

Answer 26

Unnecessary prescription -> ~ 50% of antibiotic prescriptions not required Livestock farming -> ~ 30% of UK antibiotic use in livestock farming (and reducing) Lack of regulation -> OTC availability in Russia, China, India Lack of development -> Very few antibiotics in recent years (it is not very profitable for pharmaceutical companies to make drugs which are made to keep in case of emergency and try no t to use to prevent resistance) -> financially not favourable

Question 27

Name the types of AB resistance

Answer 27

- destruction enzymes - additional targets - alteration of target - alteration in drug permeation - hyper production

Question 28

Production of destruction enzymes as a resistance mechanism

Answer 28

- beta-lactamases hydrolyse C-N bond of the beta-lactam ring - Example: - Penicillins G & V -> Gram +ve - Flucloxacillin (naturally resistant due to stereo hindrance) & Temocillin -> beta-lactamase resistant - Amoxicillin -> Broad spectrum AB - Gram -ve activity - Co-administered with Clavulanic acid (then it is resistant to the beta-lactamases but not by itself)

Question 29

What does broad spectrum mean in terms of antibiotics?

Answer 29

an antibiotic that acts on the two major bacterial groups, gram-positive and gram-negative, or any antibiotic that acts against a wide range of disease-causing bacteria.

Question 30

Resistance mechanism: additional target

Answer 30

- Bacteria produce another target that is unaffected by the drug - Example: E Coli produce different DHF reductase enzyme making them resistant to trimethoprim

Question 31

Resistance mechanism: alterations in target enzymes

Answer 31

Alterations in target enzymes - Alteration to the enzyme targeted by the drug. - Enzyme still effective but drug now ineffective - Example: S Aureus - Mutations in the ParC region of topoisomerase IV confers resistance to quinolones

Question 32

Resistance: alteration in drug permeation

Answer 32

- Reductions in aquaporins - & increased efflux systems Examples - Primarily of importance in gram –ve bacteria

Question 33

Resistance: hyperproduction

Answer 33

- Bacteria significantly increase levels of DHF reductase | - Example: E Coli produce additional DHF reductase enzymes making trimethoprim less effective

Question 34

What are the main categories of anti-fungal drugs?

Answer 34

- Azoles: Fluconazole | - Polyenes: Amphotericin

Question 35

Fungal Infections

Answer 35

- Can be classified in terms of tissue/organs: - Superficial - Outermost layers of skin - Dermatophyte - Skin, hair or nails - Subcutaneous - Innermost skin layers - Systemic - Primarily respiratory tract -> down the list the infection becomes more serious

Question 36

Azoles

Answer 36

- Inhibit cytochrome P450-dependent enzymes involved in membrane sterol synthesis - Fluconazole (oral) -> candidiasis & systemic infections

Question 37

Polyenes

Answer 37

- Interact with cell membrane sterols forming membrane channels - Amphotericin (I-V) -> systemic infections

Question 38

Drug details (antifungals)

Answer 38

15 anti-fungal drugs licensed in the UK Two most common categories: - Azoles: Fluconazole - Polyenes: Amphotericin

Question 39

LO1: Antibiotic classes: identify the main classes of antibiotic drugs and distinguish between them in terms of mechanism of action

Answer 39

Intracellular targets - Nucleic acids - Sulphonamides (DHOp), Trimethoprim (DHFR) - DNA gyrase - Fluoroquinolones e.g. Ciprofloxacin - RNA polymerase - Rifampicin - Bacterial ribosomes – Macrolides (Aminoglycosides, Tetracyclines) Cell membrane targets - Peptidoglycan (PtG) synthesis - Vancomycin inhibits pentapeptide - PtG incorporation - Carbapenems, Cephalosporins & Penicillins inhibit transpeptidase - Membrane stability - Lipopeptides & Polymyxins

Question 40

LO2: Antibiotic resistance: recall the mechanisms commonly used by bacteria to become resistant to drugs

Answer 40

- Destruction enzymes - beta-lactamases - Additional target - Different DHFR enzyme - Hyper-production - More DHFR enzyme - Changes to target - Changing DNA gyrase - Alterations in permeation - Increased efflux mechanisms

Question 41

LO3: Anti-fungals: differentiate between the drugs used to treat fungal infections

Answer 41

- Azoles - Inhibit ergosterol production | - Polyenes - Bind to ergosterol and create pores