MSOP1016 Antibacterials Lecture 1 2024-2025 PDF

MSOP1016 Antibacterial Agents Introduction & -lactam antibiotics (part 1) Dr Andrew J Hall Senior Lecturer in Chemistry Room A120, Anson Building [email protected] Recommended text An introduction to medicinal chemistry ISBN: 9780198749691 615.19 PAT (Drill Hall...

MSOP1016 Antibacterial Agents Introduction & -lactam antibiotics (part 1) Dr Andrew J Hall Senior Lecturer in Chemistry Room A120, Anson Building [email protected] Recommended text An introduction to medicinal chemistry ISBN: 9780198749691 615.19 PAT (Drill Hall Library) CHAPTER 19 Antimicrobials Classes to be covered Antimetabolites -lactam antibiotics -lactam inhibitors Carbohydrate-containing antibiotics Tetracyclines Oxazolidinones (Fluoro)quinolones Miscellaneous agents Learning outcomes At the end of this lecture, you should appreciate & understand:- the bacterial cell mechanisms of antibacterial action antimetabolites inhibiting cell wall synthesis -lactam antibiotics (1): penicillins synthesis mechanism of action resistance, acid sensitivity, spectrum of activity A little history… 17th century 19th century Moving forward in time… 1910 1934 1935 The bacterial cell The bacterial cell Gram-positive bacteria Thick cell wall surrounding the membrane The bacterial cell Gram-negative bacteria Thin cell wall covered by a complex outer membrane Antimicrobials Mechanisms of action Inhibition of cell metabolism: sulphonamides Inhibition of cell wall synthesis: penicillins, cephalosporins, - lactamase inhibitors, glycopeptides Interaction with the plasma membrane, e.g. valinomycin, polymixins Disruption of protein synthesis (translation): aminoglycosides, macrolides, tetracyclines Inhibition of nucleic acid transcription and replication: quinolones, aminoacridines + other miscellaneous mechanisms Inhibiting cell metabolism Sulphonamides Sulphonamides are the best examples of such antibacterial agents (antimetabolites) Prontosil discovered to have antibacterial properties only in vivo Sulphanilamide was the first synthetic antibacterial agent active against a wide range of infections. General class proved effective against Gram-positive organisms (pneumococci & meningococci) Problems: ineffective against Salmonella, they yield toxic metabolites Sulphonamides Structure-activity relationship (SAR) para-amino group is essential for activity and must be unsubstituted (exception: when R1 = acyl [amides]) aromatic ring and sulphonamide group are both required sulphonamide and amino groups must be directly attached to the aromatic ring aromatic ring must be para-substituted only sulphonamide nitrogen must be primary or secondary R2 is the only possible site for variation Sulphonamides Reducing toxicity Fatal if it blocks kidney tubules Note: population differences in metabolism Increased solubility due to more acidic sulphonamide NH Sulphonamides Comeback kings? Sulphonamide use decreased on introduction of penicillins Revival: longer lasting sulphonamides, e.g. sulphadoxine Sulphonamides Applications Treatment of urinary tract infections Eye lotions Mucous membrane infections Gut infections (particularly useful here) Sulphonamides Mechanism of action Other antimetabolites Trimethoprim acts against dihydrofolate reductase ➔ inhibition of DNA synthesis & cell growth COTRIMOXAZOLE - combination with sulphamethoxazole Stops incorporation of PABA into dihydropteroate Inhibition of two enzymes in one biosynthetic pathway! “Sequential blocking” Inhibiting cell wall synthesis Penicillins – scientific serendipity Discovery of Penicillin In 1928, Fleming was studying staphylococci bacteria and left some of his culture trays out in the room when he left for his vacation He came back to find that a mold was growing on one of the cultures and noticed that the bacteria surrounding the mold was dead He determined the mold to be part of the Penicillium family and named it Penicillium notatum Inhibiting cell wall synthesis Penicillins – the next steps Isolating Penicillin Problem solved in 1938 using freeze-drying and chromatography First clinical trials on crude extracts in 1941 Structure of Penicillin Established using X-ray crystallography in 1945 Inhibiting cell wall synthesis Penicillins – synthesis 5-step synthesis shown by Sheehan in 1957 Too involved to be used commercially (1% yield!) Isolation of 6-aminopenicillanic acid (6-APA) in 1958 [Beechams] opened the door to semi- synthetic penicillins [MORE LATER!] Inhibiting cell wall synthesis Penicillins G and V Acyl chain (R) depends on fermentation medium Corn steep liquor ➔ Penicillin G Three-dimensional structure Inhibiting cell wall synthesis Synthesis by fermentation? Theoretically, addition of different carboxylic acids to the fermentation medium should ➔ different acyl side chains, e.g. penicillin V This is a limited methodology: only acids of the formula RCH2COOH can be used Semi-synthetic procedure Intermediate= 6-aminopenicillanic acid (6-APA) (from Penicillium chrysogenum) Inhibiting cell wall synthesis Modern production of 6-APA 6-APA now generally synthesised by hydrolysis of penicillin G Note that this is an enzymatic hydrolysis More on why there is/was a drive to make penicillins with different acyl side chains later…. Penicillins Mechanism of action 1 Consider the cell wall structure NAG: R = H NAM: R = CH(CH3)COOH Penicillins Mechanism of action 2 Final step of cell wall synthesis Enzyme responsible for the cross- linking reaction (and inhibited by penicillins) is known as: Transpeptidase Transamidase D-alanyl-D-alanine transpeptidase DD-peptidase Penicillin Binding Protein (PDP) Penicillins Mechanism of action 3 Inhibition of the transpeptidase enzyme Penicillins Mechanism of action 4 Some theory as to why penicillins work Penicillin conformation was thought to be similar to that of the transition state of D-Ala-D-Ala during the cross-linking reaction Problem: why is 6-methylpenicillin INACTIVE? Rethink: better to be a closer analogue to the acyl-D-Ala-D-Ala structure (like 6-methoxypenicillin) Penicillins Resistance! Susceptibility to penicillins varies between bacterial strains Streptococcus Pseudomonas Staphylococcus pneumoniae aeruginosa aureus Penicillins Reasons for resistance Physical barriers Presence of -lactamases High levels of transpeptidase Affinity of transpeptidase to penicillin Transport back across outer membrane (Gram-negative) [EFFLUX] Mutations & genetic transfers Penicillins Resistance: physical barriers 1 To inhibit transpeptidase, drug must reach the outer surface of the bacterial cell membrane This means that the drug must pass through bacterial cell wall (Gram-positive & Gram-negative) What stops penicillin reaching the cell membrane? Penicillins Resistance: physical barriers 2 Why aren’t all Gram-negative bacteria resistant? Proteins called PORINS provide a transport “route” Penicillins Resistance: -lactamase enzymes 1 Most important mechanism for bacteria to gain resistance to penicillin These are enzymes mutated from transpeptidases ➔ similarities Difference: they can hydrolyse the ester link formed on opening of the -lactam ring Penicillins Resistance: -lactamase enzymes 2 Gram-positive bacteria Some release -lactamase into surrounding environment interception of penicillin e.g. Staphylococcus aureus – 95% of strains now release enzyme which hydrolyses penicillin G Gram-negative bacteria Most produce -lactamases Enzyme “trapped” in the periplasmic space Penicillin encounters a high concentration of enzyme Why aren’t all Gram-negative bacteria resistant to penicillin? Penicillins Resistance: -lactamase enzymes 3 There are > 1,000 types of -lactamase enzyme Some have selectivity for penicillins (penicillinases), some for cephalosporins (cephalosporinases) and some for both! They can appear in different concentrations and have differing affinities for the different -lactam drugs ➔ varying susceptibility Penicillins Resistance: transpeptidase Gram-negative bacteria Some produce very large amounts of transpeptidase ➔ penicillin unable to inhibit all the enzyme present! But not all transpeptidases are the same! There are differences in affinity There are different susceptibilities Penicillins Resistance: transpeptidase affinity S. aureus Early strains produced transpeptidase with a high affinity for penicillin ➔ inhibition Later strains acquired the transpeptidase PBP2a with LOW affinity for penicillin Note: this is also an issue with enterococci & pneumococci Penicillins Resistance: efflux pumps Gram-negative bacteria Some possess proteins in the outer membrane that can “pump” penicillins (and other drugs) out of the periplasmic space, lowering drug concentration Penicillins Other issues… Resistance is not the only problem with penicillins Sensitivity to acids Limited breadth of activity Do we have any solutions? Penicillins Structure activity relationship (SAR) Can we change the structure to provide solutions? Penicillins Acid susceptibility: origin 1 Ring strain Penicillins Acid susceptibility: origin 2 Highly reactive -lactam carbonyl group Penicillins Acid susceptibility: origin 3 Neighbouring group participation Penicillins Acid susceptibility: only 1 solution! The only option to tackle acid sensitivity in penicillins is to reduce neighbouring group participation Done by adding electron withdrawing groups (ewg) on the side chain amide group (R) Penicillins Improved stability & absorption Penicillins Tackling -lactamases 1 Steric shielding strategy -lactamase resistant penicillins are the “reserve army” Penicillins Tackling -lactamases 2 Isoxazoyl penicillins Isoxazoyl group tackles both acid and -lactamase sensitivity The amide “R” substituents also affect the pharmacokinetic properties of the drugs Penicillins A broader spectrum 1 Spectrum of activity depends on a variety of factors:- Structure Ability to cross the cell membrane (Gram-negative bacteria) -lactamase susceptibility Affinity for target enzyme Rate of efflux Difficult to formulate a clear-cut strategy… Penicillins A broader spectrum: trial & error 1 Hydrophobic groups on the side chain? Gram-positive activity spectrum ↑ Gram-negative activity spectrum ↓ Further increase of hydrophobicity? Gram-positive activity spectrum ≈ Gram-negative activity spectrum ↓↓ Penicillins A broader spectrum: trial & error 2 Hydrophilic groups on the side chain? (NH2, OH, COOH) Gram-positive ≈ ↓ Gram-negative ↑ Gram-negative activity best if hydrophilic group is attached to the carbon  to the side chain C=O Penicillins A broader spectrum: aminopenicillins Common “first line of defence” No steric shield ➔ -lactamase sensitivity… Poor absorption through gut Penicillins Ampicillin pro-drugs Penicillins A broader spectrum: carboxypenicillins Penicillins A broader spectrum: ureidopenicillins Newest class of broad-spectrum penicillins Better properties than the carboxypenicillins Penicillins A broader spectrum: amidinopenicillins Non-acyl amino side chain A single example: mecillinam Considered active only on Gram (-) Very poor oral bioavailability Penicillins A brief word on synergism When a second substance enhances the effect of the first The use of probenecid with penicillins is an example Penicillins Summary Structure: bicyclic, strained -lactam ring Synthesis: fermentation or semi-synthetic procedures Adding EWGs increases acid stability Steric shields can be used to protect against -lactamases Addition of hydrophilic groups on the acyl side chain broadens the spectrum of activity Prodrug preparations are useful Other substances can be co-administered to improve effects of penicillins Learning outcomes You should now appreciate & understand:- the bacterial cell mechanisms of antibacterial action antimetabolites inhibiting cell wall synthesis -lactam antibiotics (1): penicillins synthesis mechanism of action resistance, acid sensitivity, spectrum of activity