Download Antimicrobial Agents

Antimicrobial Agents
• Most other drugs to be covered affect body
systems to cure imbalance or malfunction
– E.g. reduce inflammation, relieve pain, etc.
• Antimicrobials used to inhibit or destroy living
organisms which are “uninvited guests”
– Drugs must harm invader without harming host
– Notion of selective toxicity
– Lectures will focus more on interactions between
drug and microbe than drug and host.
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Selective toxicity
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• The more distantly related the invader, the more
targets available for the drug to hit
– The less likelihood of direct toxic effects.
• Prokaryotes biochemically least similar
• Fungi and Protozoa are eukaryotes, so more
closely related to humans.
• Helminths (worms) also animals
• Viruses use our own cell machinery
• Cancer cells ARE our cells.
History of Antimicrobial Therapy
• 1909 Paul Ehrlich
– Differential staining of tissue, bacteria
– Search for magic bullet that would attack bacterial
structures, not ours.
– Developed salvarsan, arsenic derivative used
against syphilis.
http://www.chemheritage.org/EducationalServices/pharm/antibiot/activity/stain
/salvarsa.gif
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Timeline
• 1929 Penicillin discovered by Alexander
Fleming
– Messy lab, cool damp weather, luck
• 1940 Florey and Chain mass produce penicillin
for war time use, becomes available to the
public.
• 1935 Sulfa drugs discovered
• 1943 Streptomycin discovered
• Western civilization fundamentally changed
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Historical distinctions
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• Antibiotics: substances produced by organisms
that have inhibitory effects on other organisms.
– Penicillin, streptomycin
• Synthetic drugs: produced in a lab.
– Salvarsan, sulfa drugs
• Nowadays, most antimicrobials are semisynthetic
– Chemically modified versions of natural products
– Distinction between “antibiotics” and “synthetic
drugs” slowly being abandoned.
Selective toxicity means safer for host
• Antibiotics generally have a low MIC
– Minimum inhibitory concentration
– Effective at lower doses
• Good therapeutic index
– Safer; larger quantity must be administered before
harmful side effects occur.
e.g. Ti = LD50 / ED50
Where LD = lethal dose
ED = effective dose
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Where do antibiotics come from?
• Soil dwelling organisms
– Several species of fungi including Penicillium and
Cephalosporium
• E.g. penicillin, cephalosporin
– Species of actinomycetes, Gram positive
filamentous bacteria
• Many from species of Streptomyces
– Also from Bacillus, Gram positive spore formers
– A few from myxobacteria, Gram negative bacteria
– New sources explored: plants, herps, fish
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Antibiotics are common in nature
• Many are discovered every year, but not all are
useful
– May belong to previously recognized family, so not
really new
– Toxicity to host makes them unusable
– They may have poor chemical properties
• Insoluble, unstable, rapidly metabolized
– No longer effective against resistant organisms
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What and why
• Antibiotics are secondary metabolites
– Substances not essential for the growth of the
organism
– Typically produced at onset of stationary phase
• When growth slows down
– Production inhibited by presence of nutrients
• Why do microbes make them?
– No one is sure
– Habitat guarding: prevents outsiders from
establishing themselves when residents inhibited
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Mostly antibacterial compounds
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• Most infections the result of bacteria or viruses
– Except in Tropical medicine, fungal, protozoal, and
helminth infections of humans less common.
• Viral infections generally untreatable with
antimicrobial drugs
– Our focus is primarily on antibacterial drugs
• Taxonomy of living things: review
– 3 Domains: 2 of Prokaryotes and 1 Eukaryotes
– One prokaryotic Domain, Archaea, not involved in
disease.
Bacteriostatic vs. Bactericidal
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• Antibiotics differ by mode of action
• Bacteriostatic compounds inhibit the growth of
bacteria
– Holds invaders in check; host immune system does
the killing
• Bactericidal compounds directly kill the bacteria
• Location and severity of infection affect choice
of antibiotic
– E.g. CNS infection calls for bactericidal treatment.
Antibacterial agents
• There are 5 principle targets for antimicrobial
agents to work against bacterial cells
– Inhibition of cell wall synthesis
– Inhibition of protein synthesis
– Attack on cell membranes
– Disruption of nucleic acid synthesis
– Interference with metabolism
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Review and Overview of Bacterial Targets
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• Bacterial cell walls
– Except for Mycoplasma and relatives, all bacteria of
the Domain Eubacteria possess peptidoglycan
– Peptidoglycan provides shape and structural
support to bacterial cells
– Bacterial cytoplasm is generally hypertonic
compared to their environment
• Net flow of water: into cell
• Wall under high osmotic pressure
Cell walls continued
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• Chemical structure of peptidoglycan contributes
to its function
– Polysaccharide chains composed of 2 alternating
sugars, N-acetylglucosamine (NAG) and Nacetylmuramic acid (NAM)
– Cross-linked in 3 dimensions with amino acid
chains
– A seamless, bag-like molecule which resists
osmotic pressure
– A breach in peptidoglycan endangers the bacterium
Glycan chains cross-linked with amino acids
•G- and G+ vary w/
DAP vs. lysine and
at the interbridge.
•Note the presence
of unusual “D”
amino acids.
•Peptides attached
to NAM.
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Peptidoglycan is a 3D molecule
Cross links are both horizontal and vertical between
glycan chains stacked atop one another.
http://www.sp.uconn.edu/~terry/images/other/peptidoglycan.gif;
http://www.alps.com.tw/cht/img/anti-allergy_002.jpg
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There is no molecule similar to
peptidoglycan in humans, making
drugs that target cell wall synthesis
very selective in their toxicity against
bacteria.
Gram positive & Gram Negative
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• Gram positive bacteria have a thick cell wall
– Peptidoglycan directly accessible from environment
• Gram negative bacteria have a different wall
– Thin layer of peptidoglycan
– Surrounded by an outer membrane composed of
lipopolysaccharide, phospholipids, and proteins
– OM is a barrier to diffusion of molecules including
many antibiotics
• Some hydrophobic antibiotics may diffuse in.
• Porins allow passage of only some antibiotics
Gram negative cell structure
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Ribosomes: site of protein synthesis
• Prokaryotic ribosomes are 70S;
– Large subunit: 50 S
• 33 polypeptides, 5S RNA, 23 S RNA
– Small subunit: 30 S
• 21 polypeptides, 16S RNA
• Eukaryotic are 80S
Large subunit: 60 S
• 50 polypeptides, 5S, 5.8S, and 28S RNA
– Small subunit: 40S
• 33 polypeptides, 18S RNA
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Differences in structure between
prokaryotic and eukaryotic ribosomes
make antibiotics that target protein
synthesis fairly selectively toxic
against bacteria.
Drug entry into host cells and into host
mitochondria (w/ prokaryotic ribosomes) apparently
not a major problem.
Interference with nucleic acid synthesis
• Bacterial DNA is negatively supercoiled
– Supercoiling is maintained by gyrase, a type II
topoisomerase.
– Inhibition of gyrase and type IV topoisomerase
interferes with DNA replication, causes cell death
– Eukaryotic topoisomerases differ in structure
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Cell membranes as targets
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• Bacterial cell membranes are essentially the
same in structure as those of eukaryotes
– Antibiotics also affect Gram neg. cell walls, ie. Outer
membrane together with cell membrane
– Anti-membrane drugs are less selectively toxic than
other antibiotics, i.e, worse therapeutic index
– Many antifungal drugs make use of cell membrane
differences.
Other Targets
• Metabolic inhibitors
– Mostly target the folic acid synthesis pathway
– Many bacteria can and do synthesize a large
proportion of needed cofactors
– Humans require folic acid in the diet (a “vitamin”),
thus folic acid synthesizing enzymes are not an
available target in humans
• Selectively toxic
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Drug selection
• Identity of infectious agent
– Many infections viral, antibiotic administration is a
dangerous waste.
– Identification of bacterium provides rational basis
for choice
– Administration depends on severity of situation
– Choice of drug influenced by location of infection
• Certain bacteria are more likely at certain
anatomical sites, directing “blind” choice
• Not all drugs reach compartments equally
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Spectrum
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• When specific testing is not done or delayed,
antibiotic with a broad spectrum is administered
– Broad spectrum antibiotics can penetrate Gram –
outer membranes, resist inactivation, etc.
– Shotgun: better chance of inhibiting pathogen
• Downsides
– Normal microbiota found in ecological balance
– Death of normal microbiota results in overgrowth of
native, resistant bacteria (endogenous infection;
“superinfection”) or allows invasion by outside
opportunists.
Drug administration
• Antibiotics administered oral, i.v., i.m.
– Same caveats apply, i.e. acid instability, delayed
absorption with food for oral
– i.v. gives higher, quicker concentrations, reaches
more compartments with sufficient dose quickly
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Problem sites
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•
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Endocarditis: fibrin from inflammation
CSF because of blood-brain barrier
Osteomyelitis
Artificial joints, valves
– biofilms make it difficult for drug to reach target
• Abscesses: inflammatory barrier restricts
access; also bacteria stop growing
• Intracellular infections
– Penetration into host cells, not just microbe
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Host factors and toxicity
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• Antibiotics have been so successful because of
their selective toxicity
– But “any substance that has a biological effect will
have a biological side effect”
• Allergies and intolerance
– Immediate and delayed type hypersensitivities
when drug acts as a hapten
– Intolerance: e.g. erythromycin makes me puke.
• Age: renal function, development, type of
pathogen
Host factors and toxicity-2
• Renal
– Many antibiotics cleared through renal action, so
renal function affects dose, choice.
• Liver: Good hepatic function needed for
metabolism of some antibiotics
• Pregnancy
– Beware of developmental effects, teratogenesis
• Host defenses
– Influences choice of bactericidal vs. bacteriostatic
• Genetic background, metabolic factors
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Combination therapy
• Some valuable reasons why combination
therapy is used
– Synergistic effects between two drugs
– Polymicrobial infections, e.g. abdominal injuries
– Avoid or circumvent microbial resistance
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Antibiotic resistance
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• Inherent: Outer membrane of Gram negative
bacteria, wall-less bacteria.
• Mutations: change in transport protein,
ribosome, enzyme, etc. Normally harmful
mutations are selected FOR in the presence of
antibiotic.
• Plasmids: through conjugation, genetic
information allowing cell to overcome drug.
http://www.mun.ca/biochem/courses/3107/
images/Stryer/Stryer_F32-13.jpg
Mechanisms of drug resistance
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• Alteration of target: active site of enzyme
changes, ribosome changes.
• Alteration of membrane permeability: transport
protein changes, drug no longer enters; drug
that does enter is actively pumped out.
• Enzymatic destruction of drug: penicillinases
(beta lactamases)
• “End around” inhibitor: bacteria learns to use
new metabolic pathway, drug no longer
effective.