MOESM1 of Selective inhibition of PfA-M1, over PfA-M17, by an amino-benzosuberone derivative blocks malaria parasites development in vitro and in vivo

Additional file 1. In vitro and in vivo profile of compound T5. Description of data: The data, detailed in this additional file, comprises a summary table for T5 pharmacokinetic properties, followed by descriptions of the various corresponding assays

Sensitivity and accuracy of HHPRED+CODD and HMMER+CODD using the known Pfam domain occurrences for certifications.

<p>This figure reports the number of new domains (x-axis) certified by HHPRED+CODD (in orange and green for the phylum specific and non-specific approaches, respectively) and HMMER+CODD (blue) using local (left) and global (right) alignments for various FDR thresholds (y-axis).</p

Number of sequenced genomes and domain coverage in the Eukaryote tree.

<p>This figure reports the number of genomes entirely sequenced in each of the 5 supergroups of the Eukaryote tree <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0095275#pone.0095275-Keeling1" target="_blank">[58]</a>. In each group, a few sequenced genomes are provided as example, along with statistics relative to Pfam domains (release 26): the proportion of proteins where at least one Pfam domain has been identified using recommended Pfam score thresholds (above), and the proportion of amino acids covered by a Pfam domain (below). Most of the genomes sequenced to date belong to the Unikont (241) and plant (60) super-groups. We can see that there is a marked difference in the protein domain coverage between these groups and the three other groups: while the proportion of proteins where at least one known Pfam domain is usually above 70% in Unikonts and plants, it lies between 50% and 60% in the other groups. Similarly, while the proportion of amino-acids covered by a Pfam domain is often above 40% in plants and Unikonts, it is around 22% in the other supergroups.</p

New Pfam domains (release 26) identified at 5% and 10% FDR.

<p>The table reports the number of new domains identified by HHPRED (local mode, phylum non-specific approach) and CODD for the three certification types: known Pfam domains (Pfam), known InterPro non-Pfam domains (Interp), potential domains (Pot). “All”: results achieved when combining the 3 types. “# dom.”: number of new domains identified. “new fam.”: number of domain families that were not previously known in any protein of the organism. In each cell, the left and right numbers report the result at 5% and 10% FDR, respectively. Column “All”: The number in parenthesis reports the proportion of already known domains or family this represents. ^*For the certifications by Interpro domains, this is the number of domains identified at 12% FDR because no FDR below 10% can be achieved by this certification type.</p

New GO annotations at 5% FDR.

<p>“# known GO” is the number of known GO annotations from EuPathDB; “# GO known dom.” is the number of GO annotations that can be deduced from already known domains; “#GO new dom.” is the number of new GO annotations that can be deduced from new domains. Numbers in parenthesis report the number of annotations that confirm already known annotations or annotations deduced from known domains.</p

Sensitivity and accuracy of HHPRED and HMMER for <i>P. falciparum</i> and <i>L. major</i>.

<p>Number of new domains (x-axis) identified by HHPRED (green) and HMMER (blue) using local (left) and global (right) alignments for various FDRs (y-axis). For each approach, the two plain lines represent an upper and lower FDR estimate (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0095275#s4" target="_blank">Methods</a> for details). Dashed lines represent the standard error associated with these two estimates. For the sake of clarity, only the standard error above (resp. below) the upper (resp. lower) FDR estimate are represented here.</p

Cross-validation experiments on <i>P. falciparum</i> and <i>L. major</i>.

<p>The test was done on the 561 and 913 proteins of <i>P. falciparum</i> and <i>L. major</i> that have at least two known Pfam domains, respectively. The table reports the number of domains identified by HMMER and HHPRED that are certified by CODD at 3% FDR. Columns “# total certif.” and “# recovered domains” reports the total number of certified domains and the number of discarded domains that are recovered, respectively. Column “# overlaps” reports the number of newly certified domains that overlap a discarded domain.</p

Metabolomic and chemogenomic profiling.

<p>(A) Metabolic profiling: Heat map showing metabolic fingerprints of 80 Malaria Box compounds and atovaquone control. Parasite extracts were analyzed by LC-MS, and changes in metabolite pools were calculated for drug-treated parasites as compared to untreated controls. Hierarchical clustering was performed on ²log-fold changes in metabolites (data in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005763#ppat.1005763.s003" target="_blank">S2 Table</a>), scaled from -3 to +3. Six of seven compounds (indicated in red) reported to target <i>Pf</i>ATP4 [<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005763#ppat.1005763.ref025" target="_blank">25</a>] showed a distinct metabolic response characterized by the accumulation of dNTPs and a decrease in hemoglobin-derived peptides. A large cluster of compounds (indicated in blue) clustered with the atovaquone control (indicated in orange), and exhibit an atovaquone-like signature characterized by dysregulation of pyrimidine biosynthesis, and showed a distinct metabolic response characterized by the accumulation of dNTPs and a decrease in hemoglobin-derived peptides. (B) Chemogenomic profiling: A collection of 35 <i>P</i>. <i>falciparum</i> single insertion <i>piggyBac</i> mutants were profiled with 53 MMV compounds and 3 artemisinin (ART) compounds [Artesunate (AS), Artelinic acid (AL) and Artemether (AM)] for changes in IC₅₀ relative to the wild-type parent NF54 (data in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005763#ppat.1005763.s004" target="_blank">S3 Table</a>, genes queried in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005763#ppat.1005763.s005" target="_blank">S4 Table</a>). Clone PB58 carried a <i>piggyBac</i> insertion in the promoter region of the K13 gene and has an increased sensitivity to ART compounds as do PB54 and PB55 [<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005763#ppat.1005763.ref033" target="_blank">33</a>]. Drug-drug relationships based on similarities in IC₅₀ deviations of compounds generated with <i>piggyBac</i> mutants created chemogenomic profiles used to define drug-drug relationships. The significance of similarity in MoA between Malaria Box compounds and ART was evaluated by Pearson’s correlation calculations from pairwise comparisons. The X axis shows the chemogenomic profile correlation between a Malaria Box compound and AS, the Y axis with AM; the color gradient indicates the average correlation with all ART derivatives tested. Five Malaria Box compounds (MMV006087, MMV006427, MMV020492, MMV665876, MMV396797) were identified as having similar drug-drug chemogenomic profiles to the ART sensitivity cluster.</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

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