Amebiasis is a common worldwide diarrheal disease, caused by the protozoan parasite, Entamoeba histolytica.
Metronidazole has been a
drug of choice against
amebiasis for decades despite its known side effects and low efficacy against asymptomatic
cyst carriers. E. histolytica is also capable of surviving sub-therapeutic levels of
metronidazole in vitro. Novel drugs with different mode of action are therefore urgently needed. The
sulfur assimilatory de novo
L-cysteine biosynthetic pathway is essential for various cellular activities, including the proliferation and anti-oxidative defense of E. histolytica. Since the pathway, consisting of two reactions catalyzed by
serine acetyltransferase (SAT) and
cysteine synthase (CS,
O-acetylserine sulfhydrylase), does not exist in humans, it is a rational
drug target against
amebiasis. To discover inhibitors against the CS of E. histolytica (EhCS), the compounds of Kitasato Natural Products Library were screened against two recombinant CS
isozymes: EhCS1 and EhCS3. Nine compounds inhibited EhCS1 and EhCS3 with IC50 values of 0.31-490 μM. Of those, seven compounds share a
naphthoquinone moiety, indicating the structural importance of the moiety for binding to the active site of EhCS1 and EhCS3. We further screened >9,000 microbial broths for CS inhibition and purified two compounds,
xanthofulvin and
exophillic acid from fungal broths.
Xanthofulvin inhibited EhCS1 and EhCS3.
Exophillic acid showed high selectivity against EhCS1, but exhibited no inhibition against EhCS3. In vitro anti-amebic activity of the 11 EhCS inhibitors was also examined. Deacetylkinamycin C and
nanaomycin A showed more potent amebicidal activity with IC50 values of 18 and 0.8 μM, respectively, in the
cysteine deprived conditions. The differential sensitivity of trophozoites against deacetylkinamycin C in the presence or absence of
L-cysteine in the medium and the IC50 values against EhCS suggest the amebicidal effect of deacetylkinamycin C is due to CS inhibition.