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ANTIBACTERIAL AGENTS
Part 1: Penicillins
PENICILLINS
O C R N O CO2H H N H H S Me
Me
INTRODUCTION
Antibacterial agents which inhibit bacterial cell wall synthesis Discovered by Fleming from a fungal colony (1928) Shown to be non toxic and antibacterial Isolated and purified by Florey and Chain (1938) First successful clinical trial (1941) Produced by large scale fermentation (1944) Structure established by X-Ray crystallography (1945) Full synthesis developed by Sheehan (1957) Isolation of 6-APA by Beechams (1958-60) - development of semi-synthetic penicillins Discovery of clavulanic acid and b-lactamase inhibitors
STRUCTURE
R=
O
CH2
C R
Acyl side chain
H H N
H S N Me Me CO2H
6-Aminopenicillanic acid (6-APA)
O
b-Lactam ring
Thiazolidine ring
Penicillin G
OCH2
CO2H
Shape of Penicillin G
O R C NH H Me S Me O H N H CO2H
..
Biosynthesis of Penicillins
O C R N O CO2H H N S Me
Me
CYS
VAL
Properties of Penicillin G
Active vs. Gram +ve bacilli and some Gram -ve cocci Non toxic Limited range of activity Not orally active - must be injected Sensitive to b-lactamases (enzymes which hydrolyse the b-lactam ring) Some patients are allergic Inactive vs. Staphylococci
Drug Development
Aims To increase chemical stability for oral administration To increase resistance to b-lactamases To increase the range of activity 1
SAR
Amide essential
O C R
H N
Me
N O
bLactam essential CO2H
Me
Carboxylic acid essential
Conclusions
Amide and carboxylic acid are involved in binding Carboxylic acid binds as the carboxylate ion Mechanism of action involves the b-lactam ring Activity related to b-lactam ring strain (subject to stability factors) Bicyclic system increases b-lactam ring strain Not much variation in structure is possible Variations are limited to the side chain (R)
Mechanism of action
Penicillins inhibit a bacterial enzyme called the transpeptidase enzyme which is involved in the synthesis of the bacterial cell wall The b-lactam ring is involved in the mechanism of inhibition Penicillin becomes covalently linked to the enzymes active site leading to irreversible inhibition
H H N H S N O Enz CO2H Me Me
O C R
O C
H H N
H S N H Me Me CO2H
O C R
H H N
H S Me Me CO2H
Nu
R Enz-Nu
-H
O C HN Nu-Enz
NAM
L-Ala D-Glu L-Lys
NAG NAM
L-Ala D-Glu L-Lys
NAM
L-Ala NAG D-Glu
NAM
L-Ala NAG D-Glu L-Lys
NAM NAG
L-Ala D-Glu L-Ala D-Glu L-Lys L-Lys L-Ala D-Glu L-Lys
NAM
L-Lys L-Ala D-Glu
NAM
NAM
L-Lys
SUGAR BACKBONE
PENICI LLI N
D-Alanin e
TRANSPEPTIDASE
NAG
NAG
SUGAR BACKBONE
L-Lys D-Ala
Cross linking
Peptide Chain
D-Ala
OH
Peptid e Chain
D-Ala CO 2H
D-Ala O
Peptide Ch ain
Peptid e Chain
D-Ala
Peptid e Chain
Gly
Gly
OH
Blocked
O R C NH H S N O
H
Peptide Ch ain
Blocked H2 O
O R C O NH H S Me Me CO2H
O
Me Me CO2H
Gly
NH O O H S Me
HN
HN O
Me CO2H
OH
Blocked
R C O O
CO2H
H N
Me H H N H CH3 CO2H
Penicillin
Acyl-D-Ala-D-Ala
R C H N Me H S N O Me Me CO2H
R C O
H N
D- A l a - D - A l a
H S N Me Me CO2H R C H N H S HN O
Blocked
O
HO
Me Me
Acylation
Active Site
OH
CO2H
OH
Resistance to Penicillins
Penicillins have to cross the bacterial cell wall in order to reach their target enzyme But cell walls are porous and are not a barrier The cell walls of Gram +ve bacteria are thicker than Gram ve cell walls, but the former are more susceptible to penicillins
Resistance to Penicillins
Gram +ve bacteria Thick cell wall No outer membrane More susceptible to penicillins
Resistance to Penicillins
Gram -ve bacteria Thin cell wall Hydrophobic outer membrane More resistant to penicillins
L L
Cell membrane
Cell
Resistance to Penicillins
Factors Gram -ve bacteria have a lipopolysaccharide outer membrane preventing access to the cell wall Penicillins can only cross via porins in the outer membrane Porins only allow small hydrophilic molecules that can exist as zwitterions to cross High levels of transpeptidase enzyme may be present The transpeptidase enzyme may have a low affinity for penicillins (e.g. PBP 2a for S. aureus) Presence of b-lactamases Concentration of b-lactamases in periplasmic space Mutations Transfer of b-lactamases between strains Efflux mechanisms pumping penicillin out of periplasmic space 1
Fermentation
H2N
H S N Me Me CO2H
Cl
O
6-APA
O R C
O C R N O CO2H Me H H N H S Me
Semi-synthetic penicillins
PCl5
PhCH2 C N
H2O
PEN
PEN
6-APA
Note - Reaction with PCl5 requires involvement of nitrogens lone pair of electrons. Not possible for the b-lactam nitrogen.
1
O C R
H2O
H N
H S N Me
Acid or enzyme
O C R HO N H N H H S Me
O C R HO2C HN H N H H S Me
Me CO2H
Me CO2H
Me CO2H
O
H
Unreactive
b-Lactam
S
Me
S Me
Me O N H CO2H
N O CO2H
Me
Impossibly strained
Interaction of nitrogens lone pair with the carbonyl group is not possible Results in a reactive carbonyl group
R C O N
H H S N O
S N O
H
S HN O
-H
O
O
Further reactions
Strategy Vary the acyl side group (R) to make it electron withdrawing to decrease the nucleophilicity of the carbonyl oxygen
E.W.G. C O N O H N H S
Decreases nucleophilicity
X HC R
N O
a
C O
H N
H S N O
electronegative oxygen
Better acid stability and orally active But sensitive to b-lactamases Slightly less active than Penicillin G Allergy problems with some patients
Me
b-Lactamase
Me CO2H
Bulky group
R
H N
H S Me
Me CO2H
Enzyme
OMe
Me
Methoxy groups block access to b-lactamases but not to transpeptidases Active against some penicillin G resistant strains (e.g. Staphylococcus) Acid sensitive (no e-withdrawing group) and must be injected Lower activity w.r.t. Pen G vs. Pen G sensitive bacteria (reduced access to transpeptidase) Poorer range of activity Poor activity vs. some streptococci Inactive vs. Gram -ve bacteria
N O Me
Me
Orally active and acid resistant Resistant to b-lactamases Active vs. Staphylococcus aureus Less active than other penicillins Inactive vs. Gram -ve bacteria Nature of R & R influences absorption and plasma protein binding Cloxacillin better absorbed than oxacillin Flucloxacillin less bound to plasma protein, leading to higher levels of free drug
NH2
HO
H C
NH2 H N
Amoxycillin (Amoxil)
H C
NH2 H N
R=
C CH2O CMe3
PIVAMPICILLIN
C O
H S N Me Me
O
R=
TALAMPICILLIN
O CO2R
R=
CH Me
CH2Me
BACAMPICILLIN
Properties Increased cell membrane permeability Polar carboxylic acid group is masked by the ester Ester is metabolised in the body by esterases to give the free drug 1
PEN
O C
PEN
CMe3 O
PEN
H O O
C O CH2 O O
C O CH2
C OH
ENZYME
Formaldehyde
Ester is less shielded by penicillin nucleus Hydrolysed product is chemically unstable and degrades Methyl ester of ampicillin is not hydrolysed in the body - bulky penicillin nucleus acts as a steric shield 1
R=H R = Ph
CARBENICILLIN CARFECILLIN
Carfecillin = prodrug for carbenicillin Active over a wider range of Gram -ve bacteria than ampicillin Active vs. Pseudomonas aeruginosa Resistant to most b-lactamases Less active vs Gram +ve bacteria (note the hydrophilic group) Acid sensitive and must be injected Stereochemistry at the a-position is important CO2H at the a-position is ionised at blood pH
O O
TICARCILLIN
Administered by injection Identical antibacterial spectrum to carbenicillin Smaller doses required compared to carbenicillin More effective against P. aeruginosa Fewer side effects Can be administered with clavulanic acid
Azlocillin
Mezlocillin
MeO2S
HN
O R2N NH H H N H
N O
Me Me
Piperacillin
Et N O
N O
N O
CO2H
Administered by injection Generally more active than carboxypenicillins vs. streptococci and Haemophilus species Generally have similar activity vs Gram -ve aerobic rods Generally more active vs other Gram -ve bacteria Azlocillin is effective vs P. aeruginosa Piperacillin can be administered alongside tazobactam