Pharmacology of antibiotics

Question 1

What is the difference between gram-negative and gram-positive cells? And which one is more likely to be resistant to antibiotics?

Answer 1

The difference between gram-negative and gram-positive is the composition of the cell wall. To be specific it is the thickness of the peptidoglycan layer. The cell wall in gram-positive bacteria is composed of simple peptidoglycan layer. The surface of the peptidoglycan is highly polar and allows small hydrophilic molecules to get through, this is important in relation to susceptibility of bacteria to antibiotics.

In gram-negative bacteria this is slightly more complex. You still have a layer of peptidoglycan, but it is much thinner, in addition to the thin layer you also have an outer membrane. The outer membrane is like the plasma membrane and is composed of a bilayer, lipopolysaccharides on the outer layer and phospholipids on the inner layer. This makes it harder for hydrophilic and hydrophobic molecules to get through. Therefore gram-negative bacteria tend to be more resistant to antibiotics, it is difficult for antibiotics to gain access into the cell due to the presence of the outer membrane.

Question 2

What is the definition of an ideal antibiotic?

Answer 2

An ideal antibiotic is an antibacterial agent that kills or inhibits the growth of all harmful bacteria in a host, regardless of sight of infection without affecting bacterial gut flora or causing undue toxicity to the host.

An ideal antibiotic does not exist, but antibiotics have selective toxicity, this means they are able to kill or inhibit the growth of bacteria with a minimal adverse effect on the host cell. The basis for selective toxicity is the ability of the antibiotic to bind to targets that are present in bacteria but not in host cells e.g., cell wall. Or bind to targets that are present in both bacteria and host cells but are sufficiently different, allowing selectivity.

Question 3

Define the terms bactericidal and bacteriostatic?

Answer 3

Bactericidal means it kills the pathogen or it kills the bacteria. Bacteriostatic means it holds or prevents the growth of the bacteria making it easier for the host to eliminate it. This however is an oversimplification because these two are not two pure categories.

Question 4

Give examples of Beta-lactum antibiotics

Answer 4

Penicillin, Cephalosporin, Carbapenems and monobactams

Question 5

What is the characteristic feature of beta-lactam antibiotics and what is the mechanism of action?

Answer 5

All B-lactams except for monobactams are composed of a bicyclic-fused ring system containing a B-lactam ring. Monobactams are monocyclic. All B-lactams inhibit cell wall synthesis via binding to a series of enzymes, penicillin binding enzymes (PBP) that are involved in the cross-linking of peptidoglycan. Inactivation of an inhibitor of autolytic enzymes resulting in cell lysis and death.

Question 6

Give examples of Penicillins.

Answer 6

ampicillin, penicillin V, amoxicillin

Question 7

Why are penicillins given in combination with clavulanic acid?

Answer 7

Penicillins can be inactivated by beta lactamases, these are enzymes produced by bacteria that hydrolyse the beta-lactam ring rendering it inactive. Therefore they are used in combination with B-lactamase inhibitors e.g. clavulanic acid, tazobactam.

Question 8

Is penicillin lipid soluble?

Answer 8

Lipid insoluble – do not cross mammalian cells and the cerebral spinal fluid unless the meninges are inflamed in which we use them in the treatment of meningitis

Question 9

How are penicillins eliminated in vivo?

Answer 9

Rapid elimination, predominantly renal. They are found in high concentration in urine so can be used to treat urinary tract infections. We also must exercise caution when using them in patients with renal impairment, dose adjustment needs to be considered.

Question 10

What are the main side effects of Penicillin?

Answer 10

Can cause hypersensitivity reactions, rashes.

Side effects: mainly GI-related due to disturbances in gut flora.

Question 11

What is Benzylpenicillin (Pen G)? How is it administered? What is it used to treat?

Answer 11

Benzylpenicillin (Pen G) – parenteral type of penicillin

This is in activated by gastric acid, so it is given by injection and not orally. It can be used to treat a variety of conditions such as throat infections. It is also used in the prophylaxis of strep B infection in foetuses during labour.

Question 12

What is Phenoxymethylpenicillin (Pen V))? How is it administered? What is it used to treat?

Answer 12

Phenoxymethylpenicillin (Pen V) – oral type of penicillin

Similar activity to Pen G but is more acid stable so can be given orally. Treats similar infections to Pen G, but also used in:

• Pneumococcal infection prophylaxis in patients with splenectomy or sickle cell disease
• Streptococcal infection prophylaxis in patients following rheumatic fever

Question 13

What penicillin group is active against beta lactamase producing staphylococcus?

Answer 13

Penicillinase-resistant penicillin’s
- Flucloxacillin, Temocillin

They are not inactivated by beta lactamase and that is why it is effective against beta lactamase producing staphylococcus. Temocillin is also active against gram negative bacteria.

Question 14

What are examples of broad spectrum Beta lactam penicillins?

Answer 14

Broad spectrum penicillin’s
- Ampicillin, Amoxicillin

These are highly susceptible to inactivation by beta-lactamase and Staphylococcus species is resistant to ampicillin. Ampicillin has poor oral bioavailability, less than 50% of the medication given orally is absorbed and the presence of food reduces the absorption further. Therefore we tell patients to take the medication on an empty stomach or at least 30 minutes before food. Amoxicillin is more widely prescribed as it has better oral bioavailability and food does not interfere with its absorption.

Maculopapular rash is associated with the use of ampicillin and amoxicillin, these are usually not due to penicillin allergy but due to glandular fever which is a viral infection. Therefore we don’t use ampicillin and amoxicillin blindly to treat sore throats in patients.

Question 15

What is Co-amoxiclav?

Answer 15

Co-amoxiclav: Amoxicillin with clavulanic Acid

Clavulanic Acid is a beta-lactamase inhibitor and as a result you can use co-amoxiclav to treat infections due to beta-lactamase producing strains in infections such as respiratory tract infections, skin and soft tissue infections and animal bites.

Antipseudomonal penicillin’s and only available combined with a beta-lactamase inhibitor

• Piperacillin with tazobactam – Tazocin
• Ticarcillin with clavulanic Acid – Timentin

Ticarcillin has very good activity against gram-negative bacteria, and they are used in the treatment of a wide range of hospital infections or hospital acquired infections such as pneumonia or more serious UTIs.

Question 16

What are Mecillinams?

Answer 16

Mecillinams
- Pivmecillinam is a prodrug hydrolysed to mecillinam
It has significant activity against gram-negative bacteria in the treatment of salmonellosis and UTIs.

Question 17

What are the key characteristic of Cephalosporins?

Answer 17

Cephalosporins – 2nd group in the beta lactam family
Pharmacologically they are like penicillin, they act by inhibiting the synthesis of the cell wall, but they are also poorly lipid soluble, so they penetrate poorly into the cerebrospinal fluid unless the meninges are inflamed and that is why it is used in the treatment of meningitis. They are mainly excreted through the kidneys so again adjustment considerations need to be considered with patients with renal impairment.

We need to be aware of hypersensitivity reactions. If patients are allergic to penicillin, then they’re most likely allergic to Cephalosporins so their use should be limited. Their use is associated with increased risk of Clostridium difficile in both hospital and community settings, so we restrict the use of them.

Question 18

What are examples of Cephalosporins and what are they effective against?

Answer 18

1st generation: Cefradine, Cefadroxil, Cefalexin: used to treat UTI, RTI, STI, Otitis media
2nd generation: Cefaclor, Cefuroxime: used to treat H influenza and in surgical prophylaxis
3rd generation: Cefotaxime, Cefixime, Ceftazidime: used to treat serious infection
- meningitis
- pneumonia
- septicaemia

1st generations are effective against gram positive bacteria
2nd generations are effective against gram negative bacteria
3rd generations are effective against severe infections

Question 19

What are the key characteristic of Carbapenems?

Answer 19

Carbapenems – 3rd group in the beta lactam family
These are not orally active so are given by injection. Imipinem is susceptible to degradation by enzymatic activity in the kidney so is used in combination with cilastatin (which blocks its renal metabolism). Side effects: like other B-lactams: Hypersensitivity reactions should be considered.

Carbapenems penetrate the central nervous system better than other antimicrobials and therefore are associated with neurotoxicity that can cause symptoms like seizures especially in patients with renal impairment.

Question 20

What are the key characteristic of Monobactams?

Answer 20

Monobactams – 4th group in the beta lactam family
Aztreonam: Activity limited to Gram-negative aerobic e.g. Pseudomonas aeruginosa. They are not active against gram-positive bacteria so should not be used blindly to treat infections.

Question 21

What are the key characteristic of Glycopeptides? Give an example.

Answer 21

Glycopeptides e.g., vancomycin, are naturally occurring antibiotics; they are large hydrophilic molecules composed of Glycosylated cyclic or polycyclic peptides synthesised non-ribosomally. By Glycosylated cyclic we mean sugar residues attached to them. In terms of the mechanism of action they are like beta-lactam is in that they inhibit cell wall synthesis however they do this differently.

Vancomycin mechanism of action: It is effective mainly against Gram positive bacteria. It covalently binds to the terminal dipeptide, D-alanine-D-alanine of the peptidoglycan precursors thus preventing transpeptidation and trans-glycosylation.

In terms of peptidoglycan, we said that they are composed of chains of polysaccharides and these polysaccharides contain alternating sugar residues of N acetal glucosamine and N acetamide muramic acid to each residue. Each residue of N acetamide muramic acid is attached to an oligopeptide chain. This chain terminates with a D-alanine-D-alanine moiety. Glycopeptides bind to this moiety preventing the transpeptidation which is important in the cross-linking of oligopeptide chains. The binding to this moiety also interferes with trans-glycosylation which is an important step in the elongation of the polysaccharide chain linearly.

Question 22

Why are Glycopeptide e.g., vancomycin, not effective against gram-negative bacteria?

Answer 22

They are not effective against gram-negative bacteria. This is because in gram-negative bacteria the membrane has a unique lipid composition that acts as a barrier that excludes both hydrophilic and hydrophobic molecules. This membrane also contains transmembrane proteins that act as water filled channels called porins that allow hydrophilic molecules like penicillin to diffuse through but would exclude larger molecules like Glycopeptides. So the reason they are ineffective against gram-negative bacteria is because they are excluded by the outer membrane and prevented from reaching the peptidoglycan layer where they exhibit their mechanism of action.

Question 23

What are Glycopeptide e.g., vancomycin used to treat? list side effects.

Answer 23

Glycopeptides are mainly used for the treatment of MRSA and severe staphylococcal infections if penicillin allergy. Poor oral bioavailability so is given intravenously. Teicoplanin can be given intramuscularly. The only indication for using glycopeptides orally is for the treatment of C. difficile because they will act locally in the gut.

Teicoplanin compared to Vancomycin is highly protein bound and as a result has a long elimination half-life in the region of about 150 hours. This allows for once daily dosing but, as you know, for drugs that have long elimination half-life you need to give a loading dose initially to achieve a rapid steady state concentration.

For Teicoplanin initially you would give it 12-hourly for the first 3 to 5 doses then it becomes once daily.

In terms of side-effects you have:

• ototoxicity and nephrotoxicity
• red man syndrome and severe hypertension caused by vancomycin if administered rapidly

Teicoplainin may be better tolerated.
Telavancin, Dalbavancin

Question 24

What are the key characteristic of Lipopeptides? Give an example.

Answer 24

It’s like vancomycin in the aspect that it is active against gram-positive bacteria. It has a novel mode of action, although not very well understood, it is thought that it inserts itself into the cytoplasmic membrane forming oligo metric structures that would act like pores in the cytoplasmic membrane leading to leakages of ions, particularly potassium, which leads to disruption of membrane potential and inhibition of cellular pathways. Important cellular pathways like DNA and RNA synthesis.

Question 25

What are the pharmacokinetic parameters of Lipopeptides e.g., Daptomycin in vivo?

Answer 25

It is highly protein-bound and therefore has a long elimination half-life in the range of nine hours allowing for once daily dosing. Inactivated by pulmonary surfactant so should not be given to treat respiratory tract infections. Dose-dependent muscle toxicity is its main side-effect. The patient is advised to look out for muscle pain or muscle weakness and their creatine kinase levels should be monitored as that acts as an indicator of muscle damage. Daptomycin should not be prescribed with any drugs that could increase the risk of myopathy such as statins.

Question 26

Daptomycin binds to cytoplasmic membranes. How do you explain its low toxicity for mammalian cells as they also have cytoplasmic membranes?

Answer 26

It is thought that the reason for that is the difference in the nature of the lipids that constitute bacterial membranes cytoplasmic membrane and the ones that constitute the cytoplasmic membrane of mammalian cells. Bacterial cytoplasmic membrane is thought to be rich in acidic lipids which are rare in mammalian cells. These sorts of acidic lipids induce are confirmational change in Daptomycin molecules allowing their oligomerisation in the cytoplasm membrane and the formation of the pores that disrupt or permeabilise the cytoplasmic membrane.

Question 27

What are the key characteristics of Tetracyclines? Give an example.

Answer 27

• Examples e.g. doxycycline, tetracycline

They work by inhibiting bacterial protein synthesis by binding to the 30 S ribosomal subunit blocking access tRNA to its binding site. Inhibit bacterial protein synthesis by inhibiting the binding of aminoacyl t-RNA to the mRNA-ribosome complex. Passively diffuse through outer membrane porins of Gram-negative bacteria, accumulate in the periplasm, uptake into the cytoplasm is partially energy dependent. The outer membrane of gram-negative cells contains water field channels called porins these allow small hydrophilic molecules to get through. Tetracyclines diffuse through as positively charged cations through formation of complexes with magnesium ions. By forming complexes they travel as positively charged cations through the channel in the outer membrane and when they get into the periplasmic space they accumulate there. They dissociate from magnesium ions and now are weakly lipophilic and get transported across the plasma membrane in an energy dependent manner that would involve active transporters. They are selected for bacteria as we do not have the same ribosomes as bacterial cells and we do not have the same active transporters involved in the uptake of tetracycline so they cannot accumulate inside of our cells. Lack of major anti-eukaryotic effects due to selectivity towards 70S ribosomes and poor accumulation within mammalian cells.

Question 28

In terms of Tetracyclines, is resistance common? and how is this antibiotic affected by food? and what are some common side effects?

Answer 28

Resistance is high but they are still the treatment of choice for a variety of relatively uncommon infections.
Given orally – absorption improved in the absence of food. Strong chelating agents – absorption reduced in the presence of milk, antacids, and iron preparations. As strong chelating agents they bind to polyvalent cations like calcium in aluminium forming insoluble complexes which are poorly observable from the gut.

Side effects: diarrhoea, nausea and vomiting, skin reactions, photosensitivity, tooth discolouration.
Effects on skeletal development, deposition in growing bone and teeth (Should be avoided in children <12 years, pregnancy, and breastfeeding).

Question 29

What are the key characteristics of Aminoglycosides? Give an example.

Answer 29

Aminoglycosides

These are broad spectrum antibiotic and inhibit protein synthesis by binding to 30 S ribosomal subunit. They don’t always bring translation or protein synthesis to an immediate stop but sometimes they induce a conformational change in the complex between messenger RNA and the amino acid tRNA leading to mismatching so that the incorrect amino acid is incorporated into the peptide chain. When the chain is inserted into the plasma membrane it would cause a disruption in it and allow for permeabilisation of molecules including glycosides themselves?

Entry of aminoglycosides into the cells involved three stages:

• Electrostatic binding between the aminoglycoside and the negatively charged components of the lipid bilayer for example the polysaccharide in the outer membrane or the phospholipids
• Magnesium ions in a membrane parallel become displaced, magnesium ions are important for cross bridging of lipids and stabilisation of the plasma membrane so by destabilising them and removing them we cause disruption of the membrane structure leading to the initial uptake of aminoglycosides
• The uptake of glycosides requires respiration and so it is ineffective against anaerobic bacteria whether uptake of aminoglycosides is impaired under anaerobic conditions.

Question 30

Why are combination treatments in terms of aminoglycosides and beta-lactams more effective than mono-therapy?

Answer 30

Synergistic effects with other antibiotics e.g. B-lactams.
The combination treatment can be more effective than using each antibiotic individually. When aminoglycosides are used in combination with beta-lactams the efficacy improves greatly. Beta lactams are known to disrupt cell walls, so this leads to an increase of uptake aminoglycosides, more uptake then if aminoglycosides were working alone.

Question 31

What are the key characteristics of Macrolides? Give an example.

Answer 31

Macrolides – 3rd family of protein synthesis inhibitors
- Erythromycins, clarithromycin, and azithromycin.

They inhibit synthesis by binding to the 50 S ribosomal subunit blocking access of the tRNA. They are big so they sterically hindered the access of it. They inhibit the translation of proteins. They are also able to concentrate within leukocytes so they get transported to the site of infection because leukocytes will travel to the site and then they play a major role in killing bacteria. Pathogens that survive intracellularly cannot survive against macrolides.

In terms of clinical use they have a slightly broader spectrum than penicillin so that is why they are an alternative if you have a penicillin allergy. They also widely used for the treatment of respiratory tract infections and for skin and soft tissue infections. They are available in both oral and intravenous form and they are metabolised by the liver and predominantly secreted in bile.

They are inhibitors of cytochrome P450, so drug interactions always need to be checked especially because they affect the metabolism of drugs that are metabolised by the cytochrome. Inhibiting enzyme that metabolises them Will increase their plasma levels.

Question 32

Specifically in relation to Erythromycins, clarithromycin, and azithromycin, what are they used to treat?

Answer 32

Erythromycin:
- Inactive against most Gram-negative bacteria except H. influenza and N. gonorrhoeae.
- Side effects: diarrhoea, nausea, and vomiting
Clarithromycin:
- Converted into an active metabolite in the liver.
- Slightly greater activity than erythromycin against H. influenza.
- Given twice daily
- Less side effects
- Eradication of H. Pylori
Azithromycin:
- Once daily
- Treatment of gonorrhoea, Lyme disease, typhoid

Question 33

What are Quinolones? their mechanism of action and adverse effect?

Answer 33

Quinolones

Bind and stabilise topoisomerase II or IV- DNA complexes resulting in the formation of double stranded DNA breaks.

They are not protein synthesis inhibitors, but they inhibit DNA replication and transcription. They are broad-spectrum antibiotics:
- Only fluoroquinolones are available in the UK.

They have high oral bioavailability for the absorption is reduced by antacids, so we must leave two hours between them. Adverse effects:

• Convulsions (higher risk when taken with NSAIDs)
• Tendon damage (rare)
• Aortic aneurysm
• Irreversible musculoskeletal effects in children

Ciprofloxacin inhibits the metabolism of theophylline.

Question 34

Define empirical therapy

Answer 34

Empirical therapy – use the information available to make the best guess to which antibiotic to use

Underlying disease:

• Clinical indication
• Severity
• Most likely causative organisms

Antibacterial agent:

• Spectrum of activity
• Pharmacodynamics and pharmacokinetics properties
• Adverse drug reactions profile
• Drug interactions
• Formulation
• Cost-effectiveness

Patient’s characteristics:

• Age
• Allergy status
• weight
• Medical conditions
• Renal and hepatic function
• Pregnancy and breastfeeding
• Previous antibiotics exposure
• Other medications
• Social history