Antiprotozoal agents I:

Antimalarial chemotherapy
Prof. G. Obeng Adjei
Summary Epidemiology
• endemic through tropics

• 1.2 billion of approx 3.4 billion

exposed worldwide annually at
high risk

• 198 mln symptomatic cases in

2013

• > 80% deaths occur in SSA

Deaths peaked at 1.82 million in

2004; fell to 1.24 mln in 2010;
Summary life cycle
Pre/Erythrocytic stages
 Antimalarial drugs and prophylaxis
• Causal prophylaxis:
prevention of establishment
of infection in the liver 1. Causal
(inhibition of pre-erythrocytic prophylaxis
schizogony)
Blood
schizonticides
• Suppressive prophylaxis: and
suppression of erythrocytic Tissue suppressive
stages (blood schizonticides), schizonticides prophylaxis
protect against clinical illness

• Terminal prophylaxis: Gametocides

prophylaxis administration to
tackle vivax and ovale
hypnozoite stages; after
prolonged exposure (e.g.,
expatriates and long-term
travelers)
 Structural classification of antimalarial drugs
• Cinchona alkaloids – quinine, quinidine

• Synthetic quinoline derivatives:

4 aminoquinolines – chloroquine, amodiaquine, pyronaridine
8 aminoquinolines – primaquine, tafenoquine
Quinolinemethanols – mefloquine, lumefantrine, halofantrine

• Folate synthesis inhibitors:

Biguanides - proguanil
Diaminopyrimidines - pyrimethamine
Sulphonamides – sulphadoxine

• Sesquiterpene lactone endoperoxides – artemisinin & derivatives

• Antimicrobials – doxycycline, clindamycin, atovaquone

Structures and pathways targeted by antimalarial drugs
Haem metabolism:
(chloroquine, amodiaquine quinine, mefloquine, artemisinin)

Electron transport
(primaquine, atovaquone)

Folate metabolism
(sulphadoxine-pyrimethamine, proguanil)

Protein translation
(doxycycline, tetracycline, clindamycin)
 Plasmodium and Heme metabolism
• Plasmodia: limited capacity for de-novo AA synthesis –
(rely on AA released from digested host Hb for metabolism)

• degrade Hb [= protein + haem ]

(protein digested by parasite;] Haem [Fe(II)-protoporphyrin IX)

– not used (toxic; soluble) + protonated AA’s released; AA’s
removed from food vacuole via Pfcrt; Free ferriprotoporphyrin
IX reacts with oxygen → superoxide

Ferriprotoporphyrin IX:
• detoxified by parasite bio-crystallization polymerization →
crystalline haemozoin (if ferriprotoporphyrin does not
polymerize → lysosomal membrane damage → parasite
toxicity]
 Chloroquine mechanism of action
weak base (diffuses into parasite digestive vacuole); converted
to protonated, membrane-impermeable form, (at acid pH of
lysosome)  trapped in the parasite

Protonated CHQ accumulates in

vacuole, (inhibits digestion of Hb by
the parasite) binds heme/ FP →
heme-FP-CHQ complex (highly toxic;
disrupts membrane fxn) → cell
lysis, also haem polymerase

prevents toxic product detoxification Concentrates > x100-fold iRBCs

of haemoglobin catabolism
(parasite poisoned by own metabolic
products)
[Chq] required to alkalinize
mammalian lysosomes > required
to raise pH in mammalian food
vacuoles (relative non-toxicity to
humans
 Chloroquine – mechanism cont’d

(
 Chloroquine – disposition characteristics
• Well absorbed after all routes of administration
• Rapid absorption (Tmax approx. 3 hrs); distribution  [iRBCs]

• Slow distribution into large (>100L/kg) apparent Vd

• Extensive tissue sequestration[liver, spleen, kidney, lungs]
(→plasma level determined by distribution rather than
elimination): requires loading dose to achieve effective [[plasma]]

• Transformation by hepatic CYPs  DECQ and bis-DECQ

• T½  from few days to weeks (as plasma levels decline [reflect

transition from slow distribution to slower elimination from
extensive tissue stores)

• Terminal elimination T½ 30-60 days (trace in urine years after)

 Chloroquine – disposition and pharmacological actions
• ! Rapid entry and slow exit from small central
compartment after parenteral administration → potentially
lethal drug concentrations in plasma!

• ( →parenteral CHQ is administered by constant i.v infusion

or small divided doses via s.c or i.m)

• Absorption and distribution rates more closely matched

after oral administration = safer route!

• Extraordinarily safe drug; however, narrow safety margin

• Dangerous in overdose
 Chloroquine – clinical uses: current and potential
• Treatment of non-falciparum malaria (and sensitive
falciparum malaria – !in areas where resistance has not
yet emerged?)!!!

• Chemoprophylaxis: vivax, ovale and malariae – (does

not eliminate dormant liver forms of vivax and ovale)

• Amoebic liver abscesses [high concentrations in liver]

• Secondary (disease modifying) (RA, SLE etc)
• Antiviral activity ?????
 Chloroquine -adverse effects, contraindications
• Blurred vision

• Pruritus – commoner in black, severity age, fever, parasitaemia

• Cardiotoxicity (QRS widening, T wave abnormalities)

• Ototoxicity , Retinopathy, Peripheral neuropathy (!after

prolonged use for inflammatory conditions)

• Severe hypotension, respiratory and cardiac arrest – (after large

intramuscular injections or rapid intravenous infusions)

• Contraindicated in psoriasis, retinal or visual field abnormalities

 Chloroquine resistance - basis
• Chq-resistant plasmodia accumulate less drug inside parasite
food vacuole

• Protonated amino acids are generated in parasite digestive

vacuole as it degrades haemoglobin

• Protonated amino acids exit lysosome via transmembrane

protein (PFCRT)

• pfcrt mutations (eg substitution of threonine for lysine (K76T),

associated with chloroquine resistance

• The mutated PfCRT probably pumps protonated chloroquine

out of food vacuole
Chloroquine resistance
Rapid absorption,
Amodiaquine
DEAQ - T1/2elimination range from 1-3 weeks –
extensive metabolism → responsible for pharmacological effects
DEAQ +

BIS-DEAQ

AQ
4-5 hrs > 100 hrs
DEAQ
CYP2C8

May be metabolized to chemically reactive

quinone derivative –associated AE’s

Significant first-pass effect

AQ and DEAQ: Large inter-
individual PK variability
OH-DEAQ
(small amount of unchanged
parent drug escape into the
metabolism)
 Amodiaquine toxicity
• Agranulocytosis and hepatotoxicity associated prophylactic use (but
not 3-day treatment regimen)

• Large doses - dystonic reactions + other extra-pyramidal effects

(immunogenic reaction to reactive metabolites?)

• Rare but serious side effects linked to CYP2C8 variant alleles

• CYP2C8*1 = WT allele (acceptable safety profile); CYP2C8*2, *3 and

*4 = "poor metabolizer" phenotypes

• (poor metabolizers display lower treatment efficacy against

malaria, as well as increased toxicity)

• Less pruritus compared to chloroquine

 Quinine
• Cinchona tree-derived alkaloid
• mechanism of action may be similar to CHQ - however,
concentrates to different extent in plasmodial digestive vacuole
• → ? alternative modes of action (e.g., intercalation into DNA
through hydrogen bonding,  plasmodial transcription and
replication)

• Administered orally (7 day course!) or slow i.v., infusion (loading

dose may be required but bolus i.v., administration contra-
indicated due to cardiac dysrhythmia risk)

• Readily absorbed; distributes into apparent volume (approx.

1.5L/kg)
 Quinine – PK characteristics
• varies among different populations:
• plasma conc. higher among malaria patients
• apparent Vd and CL lower among malaria patients
• T1/2 longer in severe malaria patients

• Quinidine - optical isomer with identical actions

• Vd > Quinine
• CL > Quinine
• T½ elim < Qunine
 Quinine - clinical uses
Blood schizonticide against all 4 species;
Gametocidal activity against vivax and ovale

❖ Parenteral treatment of severe falciparum malaria –slow i.v, or

i.m, in dilute solution (divided doses or continuous i.v infusion
• (treatment should begin with loading dose to achieve effective
plasma concentrations; → changed to effective oral agent soon
as patient can tolerate oral medications)

❖ Oral treatment for uncomplicated falciparum malaria

• Pharmacological actions include:

• depressant action on the heart
• mild oxytocic effect on pregnant uterus
• slight blocking action on neuromuscular junction
• weak antipyretic effect
 Quinine – existing preparations
• Hydrochloride; Dihydrochloride; Sulphate; Bisulphate; Gluconate

• each of salt has a different weight – complicates dosing!

• All quinine salts may be given orally or intravenously (IV);

• Quinine gluconate may also be given IM, or rectally

• The ff. amounts of each salt contain equal amounts of quinine!:

• Quinine base = 100 mg
• Quinine bisulfate = 169 mg
• Quinine dihydrochloride = 122 mg
• Quinine hydrochloride = 111 mg
• Quinine sulfate (actually (quinine)2H2SO4∙2H2O) = 121 mg
• Quinine gluconate = 160 mg
 Quinine – adverse effects
• Bitter taste (poor oral compliance)

• Dose-related triad: (Cinchonism, hypoglycaemia, hypotension)

• Visual and auditory abnormalities (prolonged therapy)

• Hyperinsulinaemia hypoglycaemia (stimulate pancreatic  cells)

• Severe hypotension (when administered too rapidly)

• Stimulation of uterine contractions (especially 3rd trimester)

• Blackwater fever – possible hypersensitivity reaction

 Mefloquine
• Similar mechanism of action (chemically related to) quinine

• Slow onset of action; long plasma half life (up to 30days)

(enterohepatic circulation or tissue storage)– (allows
single-dose treatment regimen)

• Terminal elimination T½ approx. 20 days (allows weekly

chemoprophylactic dosing)
 Mefloquine-uses and adverse effects
Blood schizonticidal activity against falciparum and vivax (inactive
against hepatic stages or against gametocytes) → treatment [of
vivax] should be followed by primaquine
Chemoprophylaxis - non-immune travelers staying for brief periods
in endemic areas

Adverse effects:
Neuropsychiatric (vivid dreams, nightmares, hallucinations,
insomnia, anxiety, depression, psychosis, seizures)

Cardiac: conduction abnormalities (bradycardia, QT interval

prolongation, arrhythmias)
Contraindications: hx epilepsy, neuropsychiatric conditions,
arrhythmias, cardiac conduction defects; Pregnancy (long T1/2 and
uncertainty about teratogenic potential)
 Halofantrine
• Effectiveness against erythrocytic (not other) stages of all 4
plasmodia species

• Variable oral absorption; enhanced with food

• Plasma levels peak 16 hours after dosing; T½ - 4 days

• Alters cardiac conduction,→ dose-dependent prolongation of

ECG QT and PR intervals; associated with sudden death

• Cardiotoxic effect potentiated by prior mefloquine therapy

• Embryotoxic in animals (contraindicated in pregnancy)

 Lumefantrine (benflumetol)
• Structurally related to halofantrine
• Mode of action related to parasite detoxification of haem

• Variable oral absorption (markedly improves with food intake)

• Never been used as monotherapy

• Available as fixed dose combination with artemether

• Not associated with the adverse cardiac effects as halofantrine

• Drug levels may be altered by interaction with other drugs,

• Half life when used in combination = 4.5 hours

Lumefantrine
 Inhibitors of electron transport
• Plasmodia: - ATP directly derived from glycolysis
• - rudimentary electron transport system (oxidation of key
enzymes involved in nucleotide synthesis) - (eg DHOD mediates
early step in pyrimidine synthesis, and is reduced during oxidation
of DHOD → orotate)

• Ubiquinone (integral membrane protein in electron transport

chain) - accepts electrons from reduced DHOD (→ regenerating
oxidized form of DHOD necessary for pyrimidine synthesis)

• → interrupting ability of ubiquinone to oxidize DHOD can disrupt

plasmodial DNA replication (de-novo pyrimidine synthesis)
 Primaquine
• 8 aminoquinoline - antimalarial activity derived from:
• i) Quinone metabolite (which interferes with ubiquinone
function as electron carrier in respiratory chain)
• ii) non-specific oxidative damage to plasmodial mitochondria

• acts on exo-erythrocytic stages [in contrast with other

antimalarials], (active against hepatic stages of all human
plasmodia species; (inactive against sporozoites or erythrocytic
stages); Gametocidal against all 4 human malaria species

• Only available agent active against vivax and ovale hypnozoites

• Most effective drug for transmission prevention (does not treat

ongoing attacks of malaria)

• Chemoprophylaxis against all 4 plasmodia species

 Primaquine - disposition
• Administered orally only

• Absorption from GI nearly complete after oral administration

• Plasma conc. peaks within 3 hrs and falls with a T1/2elim of 6 hrs

• Apparent volume of distribution is several times that of TBW

• Major metabolite (8-(3-carboxyl-1-methylpropylamino)-6-

methoxyquinoline – eliminated more slowly and accumulates
with multiple doses
 Primaquine- summary therapeutic uses
• 1. Radical cure of acute vivax and ovale malaria: - CHQ is given
(to eradicate erythrocytic forms) and a 14-day course of
primaquine given (eradicate hypnozoites and prevent relapse)

• 2. Terminal prophylaxis of vivax and ovale malaria – (Since

standard chemoprophylaxis does not prevent vivax or ovale
malaria relapse, primaquine may be used after travel to an
endemic area to diminish the likelihood of relapse

• 3. Chemoprophylaxis - Daily treatment may provide good

levels of protection against falciparum and vivax malaria

• 4. Gametocidal action – Single primaquine dose can be used

as a control measure to render P. falciparum gametocytes
noninfective to mosquitoes (no clinical benefit to patient)
 Primaquine – notable AE’s
• Significant oxidative stress (formation of oxidized compound 
haemolysis in G6PD deficiency→ should not be administered
without confirming G6PD activity

• (RBCs unable to regenerate NADPH, [in G6PD] which is depleted

by oxidant metabolic derivatives of primaquine → impaired RBC
metabolic function → haemolysis)

• Can induce fatal haemolysis in foetal erythrocytes independent

of maternal G6PD status - should not be administered to
pregnant women

• Methemoglobinemia (in NADH methemoglobinobin reductase)

 Atovaquone
• Hydroxynapthoquinone - structural analogue of ubiquinone

• Inhibits interaction between reduced ubiquinone and cytochrome

bc1 complex (→ disrupting electron transport)
• [Plasmodia depend on e- transport to regenerate oxidized DHOD;
atovaquone disrupts pyrimidine synthesis, preventing
(plasmodial) DNA replication]

• Selectivity of atovaquone for plasmodia (100-fold) relies on

differences in AA sequences between human and plasmodial
ubiquinone

• However, selectivity is easily disrupted (single point mutation in

cytochrome bc1 complex can render plasmodia resistant to
atovaquone!) - atovaquone is not administered as single agent
 Atovaquone
• Active against tissue and erythrocytic schizonts (allows
chemoprophylaxis to be stopped 1 week after end of exposure)

• Exhibits erratic bioavailability; absorption  with fatty food;

• Plasma level-time profiles shows double peak (1st peak 1-8
hours; 2nd peak 1-4 days) - enterohepatic circulation
• Both treatment and prevention of malaria

• Atovaquone-Proguanil- fixed combination – exhibits synergistic

antimalarial activity (unrelated to action of proguanil as
antifolate [ other DHFR inhibitors do not have synergistic
effects with atovaquone!]
• Proguanil may also act as uncoupling agent in mitochondrial
membranes (when administered with atovaquone), - enhancing
atovaquone-mediated mitochondrial depolarization
 Inhibitors of folate metabolism
• Folate:
• in humans, essential vitamin; must be ingested in diet

• (must be synthesized de-novo in parasites and bacteria =

(useful target for selective drug action)

• Sulphonamides and sulphones inhibit folate synthesis by

competing with p-aminobenzoic acid

• Pyrimethamine and proguanil inhibit dihydrofolate reductase

(which prevents folate utilization in DNA synthesis)

• Together, they block folate synthetic pathway at different

points (synergistic action)
 Sulphadoxine-Pyrimethamine
• Pyrimethamine: T1/2elimination = 4 days
• Sulphadoxine, long-acting, T1/2= 170 hrs (7-9 days)

• Long T1/2 elim – permits 1x daily dosing (long T1/2 provides high selective
pressure for resistance !)

• Long T1/2 elim provides selective pressure for resistance development

• Resistance against P. falciparum common (dhfr  dhps mutations)

• Effective against blood schizonticidal stages of falciparum

• Less effective against other malaria species

• Most serious drug reaction - hypersensitivity to sulphonamide

component (Stevens-Johnson syndrome)
 Clinical uses of folate synthesis inhibitors
• Intermittent preventive treatment of falciparum malaria (IPTi, IPTp)
– Sulphadoxine-pyrimethamine

• Treatment of chloroquine-resistant falciparum malaria

• Sulphadoxine-pyrimethamine – no longer recommended (resistance)

• Chlorproguanil- dapsone – withdrawn on account of toxicity

• 4. Toxoplasmosis treatment
– Sulphadiaxine – pyrimethamine

• 5. Pneumocystosis treatment
– Sulphamethoxazole-trimethoprim
 Artemisinin (qinghaosu) and derivatives
• Most rapidly acting antimalarials known (potency in vivo = 10-
100-fold greater than that of other antimalarials)

• Herbal antipyretic >2000 years (Artemisia annua)

• Requirement for endoperoxide moiety for antimalarial activity

• Artemisinin – poorly soluble chemical extract; not used orally

(T½ 2 – 4 hrs)

• Artesunate – water soluble derivative; p.o., i.m., i.v., and rectal

use (T½ 45 min)

• Artemether – lipid soluble synthetic analogue; p.o., i.m., rectal,

(T½ 4-11 hours)
sesquiterpene lactone cyclic endoperoxides
Artemisinin - mechanism
 Artemisinin – some proposed mode(s) of action
Formation of carbon-centred free radical (when activated by Fe
(free or haem-bound) - Carbon-centred free radical has ability to
alkylate many proteins (as well as haem) [Fe-catalyzed cleavage
of endoperoxide bridge in parasite food vacuole → free radical
production → parasite toxicity

II. Inhibition of plasmodial sarco-endoplasmic reticulum

calcium-dependent ATPase [SERCA]
mechanism for specificity for plasmodia-infected
erythrocytes may derive from:
a) artemisinin requirement for haem for free radical
formation; b) artemisinin preferential accumulation in
plasmodia
 Artemisinin and derivatives
• Blood Schizonticidal against all plasmodial forms

• High recrudescence rate when used as monotherapy

• Drug levels decrease after several days (auto-induction)

• Among the safest antimalarials known

• High dose parenteral administration of lipophilic derivatives

associated with dose-dependent brainstem toxicity in animals

• Dose-related reversible changes in reticulocyte and neutrophil

counts and transaminase levels

• Avoid in pregnancy (1st trimester) – reproductive toxicity

 Combination therapy
• Simultaneous use of 2 or more blood schizonticidal drugs with
independent modes of action and different biochemical targets

• Concept based on probability that rate of resistance

development to two drugs in combination = product of
development of resistance to each of the drugs

• [e.g. if 1 in 106 parasites are resistant to drug a, and 1 in 106 are

resistant to drug B, then the probability of a parasite developing
resistance to AB is 1/106 × 1/106 = 1/1012]

• Based on the assumption that resistance could be delayed or

prevented by combining drugs
 Malaria chemotherapy: Monotherapy no longer indicated

Antimalarial combination therapy

Available ACTs
Artemether-lumefantrine
Artesunate-amodiaquine
Artesunate-sulphadoxine-pyrimethamine
Dihydroartemisinin-piperaquine
Artesunate-mefloquine
Artesunate-pyronaridine
Artesunate-sulphamethoxypyrazine-pyrimethamine
Artesunate-atovaquone-proguanil
Artemisinin combination therapy
Simultaneous use of an artemisinin
with another blood schizonticide

Rapidly acting artemisinin substantially

reduces parasite burden; partner drug
wipes out residual parasites

Rationale is to exploit synergistic potential

of the individual drugs – without
compromising safety

Overall aim of strategy:

improve efficacy,
delay onset of resistance,
reduce malaria transmission

PK mismatch may leave the longer-acting drug unprotected

 Selected characteristics-available ACT
T1/2 Efficacy Tolerance Complia Child Pregna
nce ncy

Artemether- 3 3-6 d ++ + +/- ++ ?

Lumefantrine
Artesunate- 3 9-18 d + + + ++ ?
Amodiaquine
Artesunate- 1/ 7/4 d + ++ + ++ +
Sulphadoxine- 3
Pyrimethamine
Dihydroartemis 3 9d ++ + + ++ ?
inin-
Piperaquine
Artesunate- 3 6-33 d ++ + + ++ +-
Mefloquine
 Summary classification - Antimalarials
Drug Class Use(s)
Chloroquine 4-aminoquinoline Treatment/prophylaxis

Amodiaquine 4-aminoquinoline Treatment of chq-R strains; combined

with artesunate
Piperaquine Bisquinoline Treatment in combination with
dihydroartemisinin
Quinine Quinoline-methanol Oral and i.v., Treatment of
uncomplicated and severe disease
Quinidine Quinoline methanol i.V treatment of severe disease

Mefloquine Quinoline-methanol Chemoprophylaxis and treatment

Primaquine 8-aminoquinoline Radical cure and terminal prophylaxis of

P. vivax and ovale; chemoprophylaxis for
all species
Lumefantrine Amyl alcohol Treatment, in combination with
artemether
Halofantrine Phenanthrene methanol treatment

Sulphadoxine-pyrimethamine Folate antagonist Treatment and IPT

Atovaquone-proguanil Quinone-folate antagonist Treatment and chemoprophylaxis

Artemisinin derivatives Sesquiterpene lactone Treatment for both uncomplicated and

severe disease
endoperoxides