The discovery of a novel series of compounds with single-dose efficacy against juvenile and adult Schistosoma species.

Treatment and control of schistosomiasis depends on a single drug, praziquantel, but this is not ideal for several reasons including lack of potency against the juvenile stage of the parasite, dose size, and risk of resistance. We have optimised the properties of a series of compounds we discovered through high throughput screening and have designed candidates for clinical development. The best compounds demonstrate clearance of both juvenile and adult S. mansoni worms in a mouse model of infection from a single oral dose of < 10 mg/kg. Several compounds in the series are predicted to treat schistosomiasis in humans across a range of species with a single oral dose of less than 5 mg/kg

Orally active antischistosomal early leads identified from the open access malaria box.

BACKGROUND: Worldwide hundreds of millions of schistosomiasis patients rely on treatment with a single drug, praziquantel. Therapeutic limitations and the threat of praziquantel resistance underline the need to discover and develop next generation drugs. METHODOLOGY: We studied the antischistosomal properties of the Medicines for Malaria Venture (MMV) malaria box containing 200 diverse drug-like and 200 probe-like compounds with confirmed in vitro activity against Plasmodium falciparum. Compounds were tested against schistosomula and adult Schistosoma mansoni in vitro. Based on in vitro performance, available pharmacokinetic profiles and toxicity data, selected compounds were investigated in vivo. PRINCIPAL FINDINGS: Promising antischistosomal activity (IC50: 1.4-9.5 µM) was observed for 34 compounds against schistosomula. Three compounds presented IC50 values between 0.8 and 1.3 µM against adult S. mansoni. Two promising early leads were identified, namely a N,N'-diarylurea and a 2,3-dianilinoquinoxaline. Treatment of S. mansoni infected mice with a single oral 400 mg/kg dose of these drugs resulted in significant worm burden reductions of 52.5% and 40.8%, respectively. CONCLUSIONS/SIGNIFICANCE: The two candidates identified by investigating the MMV malaria box are characterized by good pharmacokinetic profiles, low cytotoxic potential and easy chemistry and therefore offer an excellent starting point for antischistosomal drug discovery and development

Comparison of Microscopy and Alamar Blue Reduction in a Larval Based Assay for Schistosome Drug Screening

Only one drug, praziquantel, is widely available for treating schistosomiasis, a disease affecting an estimated 200 million people. Because of increasing usage there is concern about development of praziquantel drug resistance and a perceived need to develop new schistosomicides. Possible sources of these are large collections of compounds held by pharmaceutical companies and academic institutions. Anti-schistosome activity can be detected in vitro by visually assessing damage to cultured adult schistosome worms, but these are large and are recovered from mice which somewhat limits screening throughput. By contrast, schistosomula can be produced in vitro and used for screening in microwell plates, thus allowing medium throughput screening. High throughput screening (HTS) would require automated readout of schistosomulicidal action rather than manual microscopy. Here we report on the use of Alamar blue (AB), a fluorescent indicator of cell viability which can be measured rapidly and automatically. The AB assay was readily able to detect compounds causing death or severe damage to the larvae but was less reliable than microscopy for more subtle morphological changes including those induced by some known schistosome drugs. It is concluded that an automated HTS would benefit from integrated use of both AB and automatic image-based morphology assays

Integrated dataset of screening hits against multiple neglected disease pathogens.

New chemical entities are desperately needed that overcome the limitations of existing drugs for neglected diseases. Screening a diverse library of 10,000 drug-like compounds against 7 neglected disease pathogens resulted in an integrated dataset of 744 hits. We discuss the prioritization of these hits for each pathogen and the strong correlation observed between compounds active against more than two pathogens and mammalian cell toxicity. Our work suggests that the efficiency of early drug discovery for neglected diseases can be enhanced through a collaborative, multi-pathogen approach

Praziquantel analogs with activity against juvenile Schistosoma mansoni.

Six amide and four urea derivatives of praziquantel were synthesized and tested for antischistosomal activity against juvenile and adults stages of Schistosoma mansoni in infected mice. Only one of these had significant activity against adult worms, but, unlike praziquantel, six of these had low to modest activity against juvenile worms. A praziquantel ketone derivative had the best combination of activity against juveniles and adults, but it had no effect on the motility of adult S. mansoni in ex vivo culture. Cytochrome P450 metabolic stability data support the hypothesis that the major trans-cyclohexanol metabolite of praziquantel plays an important role in the antischistosomal activity of this drug

Comparison of microscopy and Alamar blue assays during routine screening.

<p>See Legend to <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0000795#pntd-0000795-g002" target="_blank">Figure 2</a> for most details. Results are shown for the screening of 558 compounds (i) by microscopy at day 3 of culture, larvae being reported as DEAD, morphologically DAMAGED or NORMAL and (ii) by addition of Alamar blue on day 7 and reading 24 h later.</p

Comparison of microscopy and Alamar blue to assess activity of known anti-schistosome drugs.

<p>Replicate 96-well microtitre plates were set up containing DMSO at 0.5% and either 10 µg/ml (Figure A) or 1 µg/ml (Figure B) of Oxamniquine (OX), Praziquantel (PZ), Meclonazepam (MCZ), Clonazepam (CZ), Ro15-5458 (Ro15), Oltipraz (OPZ) or Dihydroartemisisin (DHA). 100 schistosomula were added per well. Triplicate wells were set up for each treatment. The % dead or damaged schistosomula were counted on each of days 1, 3, 5 and 7. Alamar blue was added on these days and fluorescence read in Arbitrary Units (AU) 24 h later. Mean fluorescence values from wells containing AB and cM169 without parasites were subtracted from the values for test wells. The results for each of the triplicate wells are shown in the graphs with the horizontal bars representing the mean values. The dotted horizontal line on each AB graph corresponds to an AU value 25% lower than the mean for the triplicate DMSO controls. Values for medium alone controls were very comparable to the DMSO values (data not shown).</p

Alamar blue fluorescence development following culture with schistosomula.

<p>Wells contained 0, 100 (500/ml) or 1000 (5000/ml) schistosomula. # Wells containing schistosomula but no Alamar blue at time 0 (these values did not alter with time in culture). * P<0.01, ** P<0.0001 relative to wells with no schistosomula added. The mean ± standard deviation of fluorescence, expressed in arbitrary units (AU), is based on 5 replicate wells.</p