HU and additional RNR inhibitors inhibit P falciparum growth in vitro in a stage-specific manner. (A) Chemical structure of HU and 3 additional RNR inhibitors (3AP, clof, and gem). (B) One representative inhibitory dose-response curve (with 2 technical replicates) of Pf3D7 strain exposed to indicated drugs for 72 hours (in a standard EC50 assay). (C) EC50 of HU (Pf3D7 n = 35; pfnf54 n = 2; pfdd2 n = 14), 3AP (Pf3D7 n = 15; pfnf54 n = 2; pfdd2 n = 7), clof (Pf3D7 n = 13; pfnf54 n = 2; pfdd2 n = 8), and gem (Pf3D7 n = 15; pfnf54 n = 2; pfdd2 n = 8) as shown. (D) Exposure of Pf3D7 ring, trophozoite, and schizont stages by a 3-hour pulse of 329 μm HU and assessment of their development to subsequent stages (n = 3 each with 3 technical replicates). (E) Morphological analyses of maturation of rings exposed 72 hours to 394.8 μm of HU, as detected by Giemsa staining data shown from 1 representative experiment (from n = 3, each with technical replicates). (F-G) Fold change in HU activity against Pf3D7 infection of sickle (HbSS) compared with normal (HbAA) red cells in (F) EC50 and (G) parasite stage assays. In panels D,F-G, each dot represents an independent biological experiment (with triplicate technical replicates). Detailed information on biological and technical replicates of experiments conducted in HbSS and HbAA blood samples are provided in the legend of supplemental Figure 2B-G. In panels C-D, F-G, the horizontal bar represents the median and the vertical bar represents the interquartile range. For the mean comparisons in panels C-D, 1-way analysis of variance (ANOVA) with a Tukey post hoc analysis was used. EC, effective concentration.
