Antibiotics targeting energy metabolism Flashcards
Current treatment tuberculosis
- Isoniazid
- Ethoniamide
- Pyrazinamide
- Rifampin
New antibiotics for tuberculosis should:
- Have rapid reaction
- Be selective to kill bacteria (not patients)
- Bioavailability intermittent treatment
- Active on (almost) all individuals in all bacterial populations
- In a bacterial population, bacterias different (but genotype can be the same)
- Specific, well understood mechanism of action
- Synergistic action in drug conditions
Working mechanism Isoniazid
- Interfere with cell enveloppe
Rifampin (fluorquinolones/ Aminoglycoside)
Back up drugs
Potential target mycobacteria tuberculosis?
- Proton motive force
- Complexes of respiratory chain (use proton motive force)
- ATP synthase
Phenothiazines: (inhibitor proton motive force)
- Interfere with membrane functions
- Ineterfere with oxygen consumption
- Block Type II NADH > But bacteria will than use type I to survive
- Change redox state of respiratory chain
Clofazimine (NADH dehydrogenase inhibitor)
Same target as phenothiazines:
Interferes with membrane functions
- Its a repurpuse drug for M. Leprae
- Promising in combination therapy against tuberculosis
- Accepts e- from NADH type II and transfers them to oxygen»_space; Create peroxide (ROS > kills bacteria)
- In normal essay you will miss this drug because is causes only 10-20% of inhibition.
Imidazopyridines: Cytochrome bc1 complex inhibition:
- Cytochrome bc1 inhibition
- bc1 is important for growth
- Found by large screening and slowly improved last years
DARQ (diarylquinolines)
- ATP synthase as target:
- Prevents growth of M. tuberculosis
- Impact biochemical pathway
- Impact on bacterial growth
- Impact cellular metabolite
- Approval drug of last resort against MDR-tuberculosis. In phase II no serious side effects.
- Promising new antibiotic :
> Strongly active as drug target (Low [drug] can inhibit) : IC50 < 10 nM
> Strongly active against M. tuberculosis
> Sensitivity correlated with point mutation in ATP synthase. Stronly inhibit mycobacterial ATP synthesis. Is a strong argumentation that it was a target.
> Acceptable for human volunteers
Two types of NADH dehydrogenase (target)
Type I: Complex in mitochondria and homologous enzymes in bacteria
«_space;Also target anti-cancer drugs
Type II: Found in microorganism (bacteria/yeast) : Has massive side effects but is a target for antibacterial drugs.
Repurpuse drug
Drug proved for another disease
Cytochrme bc1 complex inhibition
- Not in our body
- Kown as anti-parasitic drugs
Bc1: Important for growth > Needs to be validated as target.
Imidazopyridines against tuberculosis ontdekking
- Make resistance mustations : One difference with wild type was a point mutation in bc1.
- Offered more of target to the bacteria (up-regulation) > sensitivity was decreased
- Did functional assay > Cellular ATP levels went down > So action imidazopyrine is related to energy metabolism
Cyt Bc1 as target needs validation
- Maybe the point mutation makes cytbc1 more active and the real target is somewhere else
- Need binding information
ATP synthase as target
- Strange target because we use it ourselfs in mitochondrien. And bacteria don’t need it, they can grow without.
- But DARQ is very specific, so low effect to human mitochondria
DARQ: Slowly inhibits mycobacterial ATP synthesis
DARQ binding experiment:
- DARQ binds ATP synthase: Find binding partners.
- Two proteins identified > Strongly indication that it is the target
ATP synthase subunits identified as DARQ binding partners.
DARQ activity test
DARQ bind ATP synthase
- Overexpression system for tuberculose: ATP synthase lacking
- Sensivity correlated with point mutation in ATP synthase.
Dormant bacteria
Same genome as bacteria that are killed but these ones will survive. We don't know how. Maybe: - Down regulate their metabolism - Inert to most antibiotics - Low metabolic activity - Maybe survive chemotherapy
Biggest advantage of drug blocking energy synthase metabolism
- These kill also dormant bacteria
Dormant bacteria are biggest challenge of antibiotics.
Why the delay in energy metabolism inhibitors?
- You see the first days no killing, this is why their working is underestimate.
«_space;This because the ATP level need to go down under a certain [] level before it will effect the bacteria. - But after three weeks lots of killing
How do bacteria survive the first three weeks after energy synthase inhibitors
- Upregulation of ATP syntheses
- Upregulation dormancy response and don’t need much ATP anymore
- Upregulate: Alternative energy pathways (cytochrome B)
- Downregulation: Biosynthesis pathways (DNA, proteins, lipids)
How does M. tuberculosis respond to blocking energy metabolism?
- Transcriptosome, proteasome response
- Observe level of target protein, pathway, metabolites and whole bacteria
- Upregulation of ATP synthese > Upregulation of dormancy response.
Influence energy source:
- Tested sensitive for ATP syntheses
- Do they use gluocose of fatty acids?
> Kill was stronger with fatty acids as only energy source
> M. tuberculosis can use human fatty acids as human energy source
> When you knock out these sources, is dependent for working of ATP synthase inhibitors
Underestimate working of ATP synthase inhibitors
- Bacteria used other sources than ATP synthase
- Work after three weeks
- We use ATP synthase ourself
- Bacteria don’t need ATP synthase to survive
Problem tuberculosis now a days:
- New drugs needed: 6 months antibiotic cocktail
- Increasing resistance strains
- 2-3 miljoen dead/year
Proton motive force:
- Pyrazinamide: Front line drug
But target not known.
Act on proton motive force not it’s main action. - Prodrug»_space; Pyrazinoic acid [] high at infection site.
Mechanism:
- Decrease membrane potential in M. tuberculosis
- Also proton motive force in our mitochondrien.
NADH dehydrogenase as target (Proton motive force)
- Phenothiazines
- Clofazimide
