week 6 - drugs of malaria Flashcards
what are the 2 main types of drugs
quinine and non-quinine
when treating malaria the drugs generally are aimed towards which stage of the parasite?
RBC trophozoite asexual stage
quinine, chloroquine and mefloquine target which part of the parasite?
the food vacuole
what was the first drug used to treat malaria and where did it come from?
quinine
plant in peru
what sort of structure is quinine?
4-methanol quinoline
what is one side effect of quinine?
it rarely causes cinchonism that can lead to blindness
against which stage is quinine effective?
the RBC asexual trophozoite stage
can quinine be used against chloroquine resistant malaria?
yes the mechanisms of resistance gained by plasmodium sp. to quinine and chlorquine differ
how is quinine administered?
orally
how much of quinine is blood bound and how much is protein bound?
70% protein
30% blood
how is quinine eliminated from the body?
readily metabolised by the liver
20% excreted in urine
problem with this drug is that the group right at the end can be metabolised in the liver and by removing this ethylene group stops the drug from working
what is the half life of quinine?
8-14 hours
what is the structure of chloroquine like?
4-aminoquinoline
where does chloroquine come from?
it is an artificial analogue of quinine
against which stage is chloroquine effective?
against the RBC trophozoite stage
Chloroquine was used extensively int he 1960s-1070s in an eradication campaign in conjunction with DDT however now _______
resistance is a major problem
how is chloroquine taken?
orally
what is the distribution of blood and protein like for chloroquine and what sorts of tissues does it prefer?
50-60% protein
preference for adipose tissue
how is chloroquine eliminated from the body?
partially metabolised in liver to active de-ethylated metabolites - these still work against the parasite unlike quinine where chopping off end group stops the drug working
excreted in urine unchanged (45%) but very slowly
what is one of the reasons chloroquine is so effective at treating malaria?
due to its long half life of 1-2 months and hence can be maintained at therapeutic levels within the blood plasma for a long time
quinine and chloroquine target for the mode of action is the food vacuole but specifically which process?
the method of feeding
As the parasite feeds the haemoglobin from the RBC is taken up by ____ and then through a modified structure called the ___ in the parasites cell surface
phagocytosis
cytosome
Haemoglobin is broken down via proteolytic processes. Releasing ___ and ______
amino acids
haematin
Haematin is toxic, what happens to it?
crystallised to form haemozoin
which part of haematin is capable of generating free radicals and damaging membranes?
the iron atom (Fe 3+)
What helps detoxify the haematin?
the haematin molecules stick together and the iron and propionate groups stick together. They neutralise each other
What is haemozoin formed of?
4 beta haematin molecules are joined by hydrogen bonding
Quinine and chloroquine mode of action: drugs enter the parasite how?
drugs enters cell by diffusion and accumulates in food vacuole
what do quinine and chloroquine prevent?
formation of haemozoin, so haematin forms and kills the parasite by generating free radicals
How do the quinine based drugs interact with the haematin?
the quinoline aromatic ring will interact with the aromatic porphyrin ring of haematin.
this interaction blocks the iron and prevents haemin detoxication.
What stops the chloroquine once in the food vacuole coming out again?
the pH
acidity in the food vacuole causes chloroquine / quinine to accumulate - they become protonated.
Malaria eradication was regarded as possible how?
prior to resistance of parasite to chloroquine
malaria eradication regarded as feasible using a combination of DTT to attack mosquito and chloroquine to kill the parasite
Describe the spread of Chloroquine resistant malaria.
Chloroquine resistant started to emerge in SE asia and S.America in the 1960s probably because of the misuse of the drug. Mainly it’s the parasite in these regions have the ability to adapt to drugs.
Most resistance starts around vietnam, laos.
In 1959 it emerges a problem in south america, and whether this was an imported case? Or was it completely separate event.
Chloroquine has spread to all of the regions of malaria in the world.
Chloroqune is more or less redundant now a days
what suggests that the mechanism of resistance to chloroquine is complex?
because the resistance develops slowly. can get spectrum of resistances - low to high
where is the resistance to chloroquine in terms of drug or target?
unlikely to be resistance at target level as the drug target is haematin which is synthesised by the human host and cannot be altered by the parasite.
more likely to be resistance at the drug level - less chloroquine in food vacuole of resistant parasites could be due to reduced uptake, and drug effluz.
chloroquine resistance involves _____ protein. A protein which contains ___ transmembrane domain and is localised to the membrane of the ______. It functions as amino acid/peptide transporter
PfCRT
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food vacuole
How was the actual mechanism to chloroquine discovered?
The parasites can undergo genetic exchange in mosquito. Took wild type populations mixed with a non resistant population in the lab and having them pass through the mosquito and then into a RBC environment and looking at the offspring at that point and monitoring the offspring for chloroquine resistance.
They managed to identify 1 loci of around 35kb associated with chloroquine resistance.
In clinical chloroquine resistant strains, mutation detected in the ____ gene
pfcrt
key chloroquine resistance conferring mutation is localised in a region of the protein implicated in _______
substrate activity
what is the point mutation called which is implicated in chloroquine resistance?
K76T
___ is a diagnostic tool for chloroquine resistance
K76T
What occurs in chloroquine sensitive strains?
chloroquine is protonated (due to low pH) hence is +vely charged
side chain on lysine (K) amino acid is a +vely charged lysine repels the +vely charged chloroquine (ie it is retained in food vacuole)
What occurs in chloroquine resistant strains?
chloroquine is protonated (due to low pH) hence is +vely charged
lysine (K) has been altered to threonine (uncharged) chloroquine can now interact with PfCRT and is transported out of the food vacuole (efflux pump)
In addition to the K76T mutations, some strains show high levels of chloroquine resistance have acquired other mutations in ___
pfCRT
What could lead to higher levels of chloroquine resistance?
resistance in K76T PfCRT strains can be enhanced by mutations in a second gene – pfmdr
(pfmdr = P. falciparum multi-drug resistance )
PfMDR is an ATP-binding cassette transporters (ABC-transporter) group of membrane proteins
MDRs on other organisms function to export hydrophobic drugs & indirectly regulate ionic gradients
MDRs responsible for drug resistance in cancer cells
the calcium channel blocker verapamil which reverses resistance of mammalian cells to anti-cancer cells reverses chloroquine resistance
pfMDR is localised to the membrane of the ______
food vacuole
Precise mechanisms underlying PfMDR in resistance remains unknown but what is known?
that different geographical regions show different mutations
Resistance to chloroquine develops ____
slowly
mutation in 2 genes (___&____) have been implicated in resistance
pfcrt
pfmdr
All chloroquine resistance strains contain a mutation in ______ leads to low / medium levels of resistance
pfcrt (K76T)
high levels of chloroquine resistance can occur in which 2 ways?
acquisition of other mutation in pfcrt
acquisition of mutations in a 2nd gene pfmdr
Mefloquine is another derivative of quinine. How is it administered?
orally
what is the trade name of mefloquine?
Lariam
When was mefloquine used and why is it not used much anymore?
used in the treatment of chloroquine resistant falciparum malaria.
crosses the blood brain barrier
severe and permanent side effects including neurological disorders
what is the mode of action of mefloquine?
food vacuole
Is there resistance to mefloquine?
resistance is emerging in south east asia - but different mechanism to chloroquine
resistance to mefloquine depends on _____ pfmdr1
wildtype
mutation of pfmdr confers mefloquine sensitivity
high levels of mefloquine resistance is associated with what?
over expression of wildtype PfMDR
so why is mefloquine resistance different to chloroquine resistance?
amplification of efflux pump = mefloquine
mutation of efflux pump = chloroquine
Sulfadoxine pyrimethamine has the trade name ______
combinational therapy
fansidar
which life stage does sulfadoxine pyrimethamine target?
active against the asexual RBC cycle
when is sulfadoxine pyrimethamine used?
second line drug combination where chloroquine resistance is prevalent.
not used in UK since 2008
what are the side effects of sulfadoxine pyrimethamine?
fatalities have occured due to drug induced, dermatological conditions.
skin rash, photosensitivity, blood disorders
Pharmacokinetics of sulfadoxine pyrimethamine = absorption?
single oral dose
adult - 3 tablets
1 tablet contains 500mg sulfadoxin and 25mg pyrimethamine
Pharmacokinetics of sulfadoxine pyrimethamine = distribution?
90% drug binds to proteins
cross placental barrier and pass into breast milk
pyrimethamine concentrates in red/white blood cells and crosses into CNS fluids
Pharmacokinetics of sulfadoxine pyrimethamine = elimination
low levels of each drug is metabolised
both excreted in urine - unmetabolised drug in urine
long half lives- sulfadoxine = 170hrs
pyrimethamine = 110hrs
What is the mode of action of sulfadoxine-pyrimethamine?
drugs target different enzyme in folate biosynthesis pathway
drugs act synergistically (cooperative)
function to reduce folate levels
many bacterial and anti cancer drugs target enzymes in this pathway
why does reducing folate levels kill parasite?
folate is essential for DNA synthesis and metabolism of certain amino acids
why is the folate biosynthesis pathway particularly attractive to target drugs at in malaria parasites?
because we do not have a folate biosynthesis pathway as we have to obtain folate in our diet
Sulfadoxine component is responsible for _____
most side effects observed in treatment
Sulfadoxine is an analogue of _____
p-amino benzoic acid
what is the target of sulfadoxine and how?
dihyropteroate synthase - this enzyme is missing in humans. Its a competitive inhibitor
pyrimethamine is a _______ drug that targets???
pyrimidine containing
targets dihydrofolate reductase this is present in humans
also competitive inhibitor
is there resistance for sulfadoxine pyrimethamine?
Arose in s.e.asia then spread rapidly. it is still used but not as widely as it used to be.
Sulfadoxine-pyrimethamine basis for resistance due to differences in _____activities.
antifolate
Type II antifolates (pyrimethamine) are _____
Type I antifolates (sulfadoxine) not as __
highly active
active
Sulfadoxine-pyrimethamine resistance occurs due to point mutations in which genes?
Firstly in DHFR-TS (target for type II)
then in PPPK-DHPS (target for type I)
Explain DHFR-TS mutation further
Only DHFR domain is affected. A diagnostic mutation in DHFR (S108N) is always seen first confers 100 fold increase in resistance to pyrimethamine.
Acquisition of secondary DHFR mutations progressively enhances resistance.
Explain PPK-DHPS mutation further
After alteration of DHFR at all 4 positions, mutations in DHPS arise to give resistance to sulfadoxine
What is the trade name of atovaquine and what sort of drug is it?
malarone
part of a combinational therapy
Which stage is atovaquine active against?
active against the liver and erythrocyte stages.
active against strains resistant to other anti malarials
very few side effects
Atovaquone pharmacokinetics - absorption
oral
1 tablet = 100mg proguanil and 250mg of atovaquone
Atovaquone pharmacokinetics - distribution
highly lipophilic
99% drug binds to
serum albium
Atovaquone pharmacokinetics - elimination
not metabolised
excretes slowly in faeces, little in urine
half life - 48-72hours
what is the active site of atovaquone?
the ubiquinone structure which is a quinone base structure
what does ubiquinone do?
shuttles electrons from dehydrogenase that form complex I and II of the oxidative phosphorylation pathway onto complex III (cytochrome b)
Is there resistance to atovaquone?
resistance with atovaquone alone can easily arise by a point mutation in the gene encoding cytochrome b.
Resistance is rare when using the Malarone combinational therapy. First failure reported in 2002
Atovaquone targets what?
mitochondrial electron transport
Which drug targets the apicoplast?
doxycycline
What is doxycycline?
tetracycline antibiotic
it is a broad spectrum antibiotic
prophylaxis to prevent malaria
which stage does doxycycline target?
active against the asexual cycle
what are the side effects of doxycycline?
stomach upset and nausea
vaginal yeast infection
Pharmacokinetics of doxycycline - absorption
oral
100-200mg/day for 7-14 days
Pharmacokinetics of doxycycline - distribution
90% drug remains in plasma
cross placental barrier and pass into breast milk
Pharmacokinetics of doxycycline - elimination
not metabolised
excreted in urine / faeces
half life 18hrs
what is the mode of action of doxycycline?
target in apicoplast - non photosynthetic organelle related to plastids such as chloroplasts that evolved via secondary endosymbiosis
The apicoplast is the bacterial ______ mechanism, doxycycline inhibits cell growth by inhibiting ________
translation
translation in the apicoplast
How does doxycycline inhibit translation in the apicoplast?
Doxy blocks the acceptance site in the ribosome and so the charged tRNA cannot come in and bind effectively with the ribosome so the protein doesnt form
What is the problem with doxy?
it prevents parasite growth (static agent) rather than killing (-cidal agents) and therefore the immune system is required to activate and wipe the parasite out
Is there resistance to doxy?
no but matter of time.
3 mechanisms of resistance reported in bacteria - enzyme inactivation, efflux, and ribosomal protection
Which part of the parasite do the artemisinins affect?
no idea!
what is artemisinin?
from leaves of a plant that have been used by Chinese herbalists, the active compound was purified in 1972
Artemisinin is made from a series of _______units that contain a ______bond
isoprene
peroxide
Which stage is artemisinin active against?
asexual ring stage
When is artemisinin used?
to treat multi drug resistant plasmodium strains
Is there resistance to artemisinin?
documented in south wast asia.
The molecular marker for resistance is KELCH protein
what are the 3 suggested mode of actions for artemisinin?
inhibition of food vacuole cysteine protease activity
damage parasites electron transport chain in the mitochondria to oxidative stress
irreversible inhibition with an ATP-ase that pumps Calcium from the cytoplasm into the ER
What are the problems with Artemisinin?
problems with supply cannot grow enough plant material
semi-synthetic compounds developed
artemisinin use as a monotherapy explicitly discouraged by the WHO treatment failure because it has been used on the Thai Laos boarder and it should always be a part of a combinatorial therapy
what has been suggested as the way forward with Artemisinin?
Artemisinin Combinational Therapies
What is Artesunate and Amodiaquine?
ASAQ
Active against all forms of malaria and against all chloroquine resistant strain of Plasmodium falciparum
What has ACT accomplished?
in a short period ACTs main treatment targeting malaria in africa.
