trans

DRUG RESISTANCE IN APICOMPLEXAN PARASITES 405
failure rates by inclusion of pfmdr1 status in
association with pfCRT.
The biochemical function of PfCRT (and
Pgh1) are unknown. Biochemical evidence indicating
reduced access to heme as the basis of
resistance is consistent with any mechanism
that keeps heme and chloroquine apart within
the parasite. It has been argued that mutations
in PfCRT decrease the pH of the terminal vacuole.
It was claimed that this reduction in pH,
rather than increasing drug accumulation by
ion trapping of the weak base, reduces heme
solubility, thereby reducing the availability of
the drug binding site. Unfortunately the experimental
approach used to measure the pH in
these experiments was fundamentally flawed.
The pH probe used was localized exclusively
in the cytosolic compartment rather than in
the food vacuole and therefore could not be
used to report the true vacuolar pH. Ongoing
experiments are aimed at determining if PfCRT
alters drug accumulation at the target site indirectly
by influencing ion gradients within the
parasite, or whether it can move the drug (or
less likely heme) directly. This information is
central to any strategies involving resistance
reversal and rational drug design.
Chloroquine resistance is becoming an
increasing problem in Plasmodium vivax.
P. vivax, although not generally fatal, is responsible
for considerable morbidity with some
75–90 million cases annually. The drug of choice
against the erythrocytic stage is chloroquine.
Chloroquine resistance in P. vivax was first
reported in 1989 in Papua New Guinea. Despite
comparable chloroquine use, clinical resistance
in P. vivax has taken much longer to appear
than resistance in P. falciparum. Little is known
about the chloroquine-resistance phenotype in
P. vivax due to difficulties in maintaining these
parasites in continuous culture, although the
drug is assumed to have the same mechanism
of action. Analysis of the PfCRT homolog in
P. vivax failed to identify any mutations in this
gene that were associated with chloroquine
resistance.
Amodiaquine
The 4-aminoquinoline amodiaquine is a structural
analog of chloroquine (Figure 16.2).
When used clinically amodiaquine undergoes
efficient dealkylation to desethyl-amodiaquine.
Both parent drug and metabolite contribute to
antimalarial action in vivo. All of the available
evidence supports the argument that amodiaquine
and chloroquine share a common
mechanism of action based on an interaction
with heme. Amodiaquine is more active than
chloroquine in vitro. This increased activity
may be related to the drug’s greater lipophilicity,
but is independent of its weak base qualities
(amodiaquine is a poorer weak base than
chloroquine) or its ability to interact with
heme (as determined from inhibition of heme
crystalization). Amodiaquine and chloroquine
share cross-resistance in vitro, and drug activity
correlates with drug accumulation. Absolute
parasite sensitivity to amodiaquine is always
superior to chloroquine, and a verapamil effect
has never been demonstrated. In comparison,
the principal circulating metabolite desethylamodiaquine
shares greater cross-resistance
with chloroquine and there is a slight verapamil
effect. Based on these observations it is probable
that mutations in PfCRT also influence
in vitro parasite susceptibility to amodiaquine
and desethyl-amodiaquine. This argument
has not yet been formally tested. However,
the selection of the K76I mutation under
chloroquine pressure in an isolate carrying the
complete complement of additional PfCRT
mutations resulted in a reduced susceptibility
of the selected isolate to amodiaquine and
quinacrine in addition to chloroquine. Despite
these in vitro associations, there is compelling
MEDICAL APPLICATIONS