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. 2018 Jan 12;359(6372):191-199.
doi: 10.1126/science.aan4472.

Mapping the Malaria Parasite Druggable Genome by Using in Vitro Evolution and Chemogenomics

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Free PMC article

Mapping the Malaria Parasite Druggable Genome by Using in Vitro Evolution and Chemogenomics

Annie N Cowell et al. Science. .
Free PMC article

Abstract

Chemogenetic characterization through in vitro evolution combined with whole-genome analysis can identify antimalarial drug targets and drug-resistance genes. We performed a genome analysis of 262 Plasmodium falciparum parasites resistant to 37 diverse compounds. We found 159 gene amplifications and 148 nonsynonymous changes in 83 genes associated with drug-resistance acquisition, where gene amplifications contributed to one-third of resistance acquisition events. Beyond confirming previously identified multidrug-resistance mechanisms, we discovered hitherto unrecognized drug target-inhibitor pairs, including thymidylate synthase and a benzoquinazolinone, farnesyltransferase and a pyrimidinedione, and a dipeptidylpeptidase and an arylurea. This exploration of the P. falciparum resistome and druggable genome will likely guide drug discovery and structural biology efforts, while also advancing our understanding of resistance mechanisms available to the malaria parasite.

Figures

Fig. 1
Fig. 1
Genetic changes acquired by 235 compound-resistant P. falciparum clones. (A) Circos plot (68) summarizing single nucleotide variants (SNVs), insertions and deletions (indels), and copy number variants (CNVs) acquired by the 235 P. falciparum clones resistant to 37 diverse compounds with antimalarial activity, grouped by chromosome. Each bar on the outer three rings represents 30,000 bp, and the darkness of the bar indicates a greater number ofmutations.Orange ring,mutations lead to loss of function; blue ring, mutations lead to protein modification (nonsynonymous change or inframe deletion); green ring, no protein change (synonymous mutations or introns). The purple ring displays a histogram showing total counts of all three types of mutations, with orange bars where counts exceed 15.The gray rings display variants for resistant clones grouped by compound,with each ring representing one of 37 compounds. Red bars represent the location of CNVs, and blue bars represent the location of SNVs. (B) CNVs in the known drug-resistance gene pfmdr1.The heatmap (left) shows clones grouped by compound and is brighter yellowfor genes with higher coverage (>2.5-fold above the standard deviation). Resistance shifts (top right) relative to the sensitive parent (circles) were demonstrated in MMV665789-resistant clones (triangles). PCR products (bottom right) shows that resistant clones contain the pfmdr1 CNV junction sequence,whereas a 3D7 control lacks this sequence. (C) CNVs in pfabcI3 are associated with resistance to multiple compounds. Resistance shifts relative to the sensitive parent (circles) were demonstrated in MMV029272-resistant clones (triangles). Again, the CNV junction sequence was not found in the 3D7 control DNA, whereas it was identified in clones resistant to MMV029272.
Fig. 2
Fig. 2
The P. falciparum resistome. Compounds were clustered first on the basis of their target profile similarities, then by their chemical structural similarities. Each compound was assigned a color code, which is shared with Fig. 1. Genes that were detected as mutated (CNV, SNV, or indel) in independently created clones are listed.
Fig. 3
Fig. 3
A computational model of active antimalarial small molecules docked against their respective targets. Schematics of the biochemical pathway, the compound, and conserved pfam domains are shown in the right panel. (A) MMV027634 occupies the thymidylate synthase active site. The benzoquinazoline head group of MMV027634 is stabilized by hydrogen-bonding within the dUMP active site of dihydrofolate reductase– thymidylate synthase (DHFR-TS). Further stabilization occurs at the tail with multiple hydrogen bonds to Gly378, mutation of which confers resistance to this compound. Mutations are all found in the thymidylate synthase portion of the molecule (http://pfam.xfam.org/family/pf00303). (B) MMV019066 and farnesyl pyrophosphate (FPP) are shown concurrently docked within the binding pocket of farnesyltransferase. A model of the P. falciparum farnesyltransferase beta subunit was constructed using the rat homolog (Protein Data Bank ID, 2ZIR) as a template. Despite substantial interspecies protein sequence variance, the FPP binding pocket is largely conserved (69). The preferential binding states of FPP andMMV019066 are shown competing for similar hydrophobic space. Resistance mutations are found in the squalene-hopene cyclase domain (http://pfam.xfam.org/family/pf13249). The yellow circle represents the rest of the donor protein to which the farnesyl group is attached. dTMP, deoxythymidine monophosphate; NADP+, nicotinamide adenine dinucleotide phosphate; NADPH, reduced form of NADP+; Me, methyl; R, benzoyl-L-glutamic acid. Single-letter abbreviations for the amino acid residues are as follows: A, Ala; C, Cys; D, Asp; E,Glu; F, Phe; G,Gly; H, His; I, Ile; K, Lys; L, Leu; M, Met; N, Asn; P, Pro; Q, Gln; R, Arg; S, Ser; T, Thr; V, Val; W, Trp; and Y, Tyr.

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References

    1. Ashley E. A., et al. , N. Engl. J. Med. 371, 411–423 (2014). - PMC - PubMed
    1. Ariey F., et al. , Nature 505, 50–55 (2014). - PMC - PubMed
    1. Flannery E. L., Fidock D. A., Winzeler E. A., J. Med. Chem. 56, 7761–7771 (2013). - PMC - PubMed
    1. Spangenberg T., et al. , PLOS ONE 8, e62906 (2013). - PMC - PubMed
    1. Gamo F.-J., et al. , Nature 465, 305–310 (2010). - PubMed

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