title: Activity of clinically relevant antimalarial drugs on Plasmodium falciparum mature gametocytes in an ATP bioluminescence "transmission blocking" assay.
abstract: BACKGROUND
  Current anti-malarial drugs have been selected on the basis of their activity against the symptom-causing asexual blood stage of the parasite. Which of these drugs also target gametocytes, in the sexual stage responsible for disease transmission, remains unknown. Blocking transmission is one of the main strategies in the eradication agenda and requires the identification of new molecules that are active against gametocytes. However, to date, the main limitation for measuring the effect of molecules against mature gametocytes on a large scale is the lack of a standardized and reliable method. Here we provide an efficient method to produce and purify mature gametocytes in vitro. Based on this new procedure, we developed a robust, affordable, and sensitive ATP bioluminescence-based assay. We then assessed the activity of 17 gold-standard anti-malarial drugs on Plasmodium late stage gametocytes.
  
  
  RESULTS
  Difficulties in producing large amounts of gametocytes have limited progress in the development of malaria transmission blocking assays. We improved the method established by Ifediba and Vanderberg to obtain viable, mature gametocytes en masse, whatever the strain used. We designed an assay to determine the activity of antimalarial drugs based on the intracellular ATP content of purified stage IV-V gametocytes after 48 h of drug exposure in 96/384-well microplates. Measurements of drug activity on asexual stages and cytotoxicity on HepG2 cells were also obtained to estimate the specificity of the active drugs.
  
  
  CONCLUSIONS
  The work described here represents another significant step towards determination of the activity of new molecules on mature gametocytes of any strain with an automated assay suitable for medium/high-throughput screening. Considering that the biology of the forms involved in the sexual and asexual stages is very different, a screen of our 2 million-compound library may allow us to discover novel anti-malarial drugs to target gametocyte-specific metabolic pathways.
authors.affiliation: GlaxoSmithKline R&D, Tres Cantos Medicine Development Campus, Malaria Discovery Performance Unit, Madrid, Spain. Joel.l.lelievre@gsk.com
authors.forename: Joël
authors.initials: J
authors.lastname: Lelièvre
authors.forename: Maria Jesus
authors.initials: MJ
authors.lastname: Almela
authors.forename: Sonia
authors.initials: S
authors.lastname: Lozano
authors.forename: Celia
authors.initials: C
authors.lastname: Miguel
authors.forename: Virginia
authors.initials: V
authors.lastname: Franco
authors.forename: Didier
authors.initials: D
authors.lastname: Leroy
authors.forename: Esperanza
authors.initials: E
authors.lastname: Herreros
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry*; D000818:Animals; D000962:Antimalarials / Q000494:pharmacology*; D005854:Germ Cells / Q000187:drug effects*; D008163:Luminescent Measurements / Q000379:methods*; D021261:Parasitic Sensitivity Tests / Q000379:methods*; D010963:Plasmodium falciparum / Q000166:cytology* / Q000187:drug effects*
chemical_list: D000962:Antimalarials; D000255:Adenosine Triphosphate