High-Throughput Assay and Discovery of Small Molecules that Interrupt Malaria Transmission

Abstract

SummaryPreventing transmission is an important element of malaria control. However, most of the current available methods to assay for malaria transmission blocking are relatively low throughput and cannot be applied to large chemical libraries. We have developed a high-throughput and cost-effective assay, the Saponin-lysis Sexual Stage Assay (SaLSSA), for identifying small molecules with transmission-blocking capacity. SaLSSA analysis of 13,983 unique compounds uncovered that >90% of well-characterized antimalarials, including endoperoxides and 4-aminoquinolines, as well as compounds active against asexual blood stages, lost most of their killing activity when parasites developed into metabolically quiescent stage V gametocytes. On the other hand, we identified compounds with consistent low nanomolar transmission-blocking activity, some of which showed cross-reactivity against asexual blood and liver stages. The data clearly emphasize substantial physiological differences between sexual and asexual parasites and provide a tool and starting points for the discovery and development of transmission-blocking drugs.

Discovering New Transmission-Blocking Antimalarial Compounds: Challenges and Opportunities

“…Primary hit rates (>80% inhibition at 5 μM) of 4.5-24% [20,23,27,29] were achieved when screened against stage IV/V gametocytes; this increased to 33% against stage I-III gametocytes [23,26]. Furthermore, although the rank-order of hits were similar, there was not complete overlap between MMV Malaria Box screens conducted with different assay platforms or between laboratories using the same assay platform [24,25].…”

“…Hits should be tested against asexual parasites to confirm specific activity or to identify a dual-active compound. If a compound is only active against gametocytes (>10-fold selectivity [20]) it may be used to eliminate the reservoir of gametocytes in semi-immune asymptomatic carriers with a predictable 'reduced-risk' of inducing resistance. However, strategies that will allow efficacy, dosing and safety testing in vivo have not been clearly defined for transmission-blocking only compounds compared to those outlined for compounds active against asexual parasites and this could complicate their clinical development.…”

“…Similar approaches have been proposed to discover transmission-blocking compounds by screening against primarily mature gametocytes but this has been very difficult compared to screening asexual blood stages, particularly because the apparent quiescence of mature gametocytes has hampered the development of cell-based screening assays specific to stage V gametocytes [19]. This limited metabolic repertoire restricts the 'druggable' pool of biochemical activities in these parasites [20].…”

“…The Medicines for Malaria Venture (MMV) Malaria Box of 400 compounds (selectively active against asexual parasites [22]) has been screened worldwide for activity against stage IV/V gametocytes [20,[23][24][25][26][27][28][29][30][31]. Primary hit rates (>80% inhibition at 5 μM) of 4.5-24% [20,23,27,29] were achieved when screened against stage IV/V gametocytes; this increased to 33% against stage I-III gametocytes [23,26].…”

“…Some chemical signatures for compounds targeting both asexual and gametocyte stages have been described [24] including endoperoxides, certain quinolines (including primaquine and mefloquine), anthracyclines, dihydroergotamine-type adrenergic agents and inhibitors of kinases, protein biosynthesis, the proteasome, protein modification / membrane trafficking and ion homeostasis [20,32]. Additionally, certain chemical classes (e.g.…”