Additional file 5: Figure S3. of Fluorescence/luminescence-based markers for the assessment of Schistosoma mansoni schistosomula drug assays

Fluorescence generated from culture medium as measured by (A) Omnicathepsin, (B) DAPI and (C) Hoechst 33258. Since our standard culture medium could also contribute to high fluorescence, RPMI medium was also tested. Graphs presented here correspond to optimal marker concentrations and incubation times are indicated in the main text. (PPTX 79 kb

Boletín de Segovia: Número 131 - 1838 noviembre 8

Copia digital. Madrid : Ministerio de Cultura. Subdirección General de Coordinación Bibliotecaria, 200

Chemical structures of the five lead compounds selected for <i>in vivo</i> studies.

<p>Chemical structures of the five lead compounds selected for <i>in vivo</i> studies.</p

Worm burden reductions observed for the five lead candidates in <i>S. mansoni</i> infected mice.

<p>Mice harbored a patent <i>S. mansoni</i> infection. Different dosage regimens were used (1×400 mg/kg, 4×80 mg/kg on four consecutive days or 4×100 mg/kg every 4 hours).</p><p>WBR: Worm burden reduction.</p><p>p-value<0.05.</p><p>p-value<0.005.</p

Adult worm IC₅₀ and IC₉₀ values of five <i>in vivo</i> candidates compared to praziquantel 72 hours post drug exposure.

<p>PZQ: Praziquantel.</p

Adult worm IC₅₀ values of five <i>in vivo</i> candidates over time post drug exposure.

<p>Values were determined 1, 2, 4, 7, 10, 24, 48 and 72>33.3 µM are indicated with (<b>//</b>). PZQ: praziquantel.</p

Screening flow.

<p>Screening was conducted at the Swiss TPH (screening cascade <b>I</b>; steps <b>A</b>–<b>E</b> and Quality control): Primary screening steps (yes/no filters) of 100 µM and 33.3 µM resulted in 179 and 72 hits, respectively. Active compounds (n = 72) moved on to Step <b>C</b> and IC₅₀ values were evaluated on NTS. Thirty-four compounds showed activities with IC₅₀ values <10 µM and pre-screening was conducted on adult schistosomes (Step <b>D</b>). Active compounds (n = 16) with schistosomicidal effects at 33 µM compound concentration were further characterized (step <b>E</b>). The quality control represents randomly selected compounds from compounds classified as non-active from the pre-screening steps (step <b>A</b>/<b>B</b>) on NTS which were re-evaluated in step <b>B</b>. In parallel, all compounds (n = 400) were studied at the LSHTM in London (screening cascade <b>II</b>): step <b>A</b>, all 400 compounds were screened on <i>S. mansoni</i> adults at 15 µM. Step <b>B: 44</b> compounds were active and these were then tested for IC₅₀ determination on adult worms. From both screening cascades, five compounds were selected for <i>in vivo</i> testing based on pharmacodynamic and pharmacokinetic properties as well as toxicity.</p

Characterization of five lead candidates selected for <i>in vivo</i> testing.

<p><i>In vitro</i> activity on <i>P. falciparum</i> 3D7, NTS, adult <i>S. mansoni</i>, cytotoxicity on MRC5-cells, and pharmacokinetic parameters* of 5 active compounds selected for <i>in vivo</i> studies identified in 2 parallel screens at the Swiss TPH and LSHTM.</p><p>PK parameters are unpublished data, <i>In vitro</i> activity on <i>P. falciparum</i> 3D7 and cytotoxicity on MRC5-cells can be found at <a href="http://www.mmv.org/research-development/malaria-box-results" target="_blank">http://www.mmv.org/research-development/malaria-box-results</a>.</p

Malaria Box Heatmap.

<p>Shown are selected data from the HeatMap (<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005763#ppat.1005763.s002" target="_blank">S1 Table</a>) for the 400 Malaria Box compounds. Each column represents an assay (grouped by category), compounds are represented in rows. The red-green gradient represents higher to lower activity. Favorable PK activities are scored green. <i>Pf</i>: <i>Plasmodium falciparum</i>, <i>Pb</i>: <i>Plasmodium berghei</i>, PK: pharmacokinetics, sol.: solubility, hERG: human ether-a-go-go channel inhibition, DDI: drug-drug interactions (predicted).</p

Antiprotozoal Malaria Box compounds with activity in biological assays and lacking toxicity at therapeutic levels.

<p>Selectivity Index, SI, is toxicity level/activity level; p, probe-like; d, drug-like.</p