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  • Recently naphthoquinone derivatives have shown promising res

    2019-12-03

    Recently, naphthoquinone derivatives have shown promising results as antiparasitic lead compounds [28]. Conjugated hybrid compounds could be an effective path to discovery of new drugs by associating two different pharmacophore groups with different mechanisms of action in a single molecule [29,30], which could yield a drug that kills the plasmodial parasites in two different ways. The chloroquine pharmacophoric group, responsible to inhibit the polymerization of the harmful heme group into the hemozoin non toxic pigment, was also planned to be introduced in a final hybrid kasugamycin to have two possible mechanisms of action, as shown in Scheme 1 [29]. A synthetic approach to produce a hybrid molecule is via click chemistry, introduced by Sharpless, and currently widely used [31], including for natural product activity enhancement [[32], [33], [34], [35], [36], [37], [38], [39]]. Hybrid molecules can be easily produced by combining a terminal alkyne of a naphthoquinone intermediate (6) with different organic azides. Our strategy was to modify a known active natural product lapachol (1), first by alkylating at the hydroxyl position. This ether naphthoquinolyl intermediate was then reacted with different organic azides supporting sulfonamide or 7-chloroquinoline moieties besides miscellaneous aryl groups using a copper cycloaddition click reaction and to evaluate the antiplasmodial activity of the new compounds.
    Results and discussion Lapachol is an alkyl naphthoquinone natural product occurring in Handronthus species (Syn. Tabebuia) disclosing a large spectrum of biological activities. During the Second World War, lapachol analogues including hydrolapachol and lapidone were studied in a search for alternative antimalarial compounds. Later, a lapachol modified structure led to development of the antimalarial atovaquone, a synthetic chloride naphthoquinone [40]. Structural modification of lapachol, which is abundant in Brazilian Handroanthus spp. (Bignoniaceae), was the strategy aimed to prepare potentially more potent antimalarial naphthoquinones. The possibility of linking different moieties to the lapachol hydroxyl group was the inspiration for the present work, once its easy alkylation by a Williamson reaction would afford the alkynyl moiety necessary for the click reactions to be carried out. From the propargyl ether 2, a total of 17 new naphthoquinonolyl-1,2,3-triazole hybrid compounds were synthesized and spectroscopically characterized. Natural product lapachol (1) was isolated by basic/acid work up and further purified by silica column chromatography and the synthesized compounds were evaluated for their in vitro antimalarial activity against P. falciparum W2 that is a chloroquine resistant and mefloquine-sensitive strain. Table 1 shows the yields of the click reactions, the IC50 values for the in vitro antimalarial assay (pLDH method), the CC50 values for the cytotoxicity to HepG2 cells (MTT method) and the SI for each compound. In general, low cytotoxicity (CC50 > 100 μM) was observed except for the hybrid lapachol-AZT derivative (18) (CC50 < 100 μM). All the compounds disclosed IC50 < 100 μM and were thus more active than lapachol (IC50 = 123.5 ± 11.5 μM), except possibly for 21, for which the exact IC50 was not determined (IC50 > 104.6 μM). Eleven compounds showed IC50 < 10 μM, with the most active being 13, 14 and 25 with IC50 of 4.0 ± 1.6, 4.6 ± 1.9, and 4.1 ± 1.0 μM, respectively. The most promising compounds are 17 and 25 that disclosed the highest SI values (197.7 and 171.0, respectively). Compound 7 also has a favorable SI (128.8). Its effect might be because of a second mechanism of action, that is, the inhibition of heme polymerization, similarly to chloroquine, due to the presence of a chloroquinolyl pharmacophore group. Molecular modeling can provide important information about the interaction of the obtained compounds and potential parasite targets, such as DHODH. The inhibitor-binding site of PfDHODH, located in proximity to the cofactor-binding site, is characterized by the presence of two regions: the H-bond pocket, comprising H185, Y528 and R265, and the hydrophobic pocket. It has been crystallographically proven that the 1,4-naphthoquinone substructure of atovaquone binds into the H-bond pocket, while its large hydrophobic substituent resides in the hydrophobic pocket. Interestingly, the compounds reported in this paper also possess a 1,4-naphthoquinone moiety, similarly to atovaquone, as well as a large hydrophobic substituent attached at position 3. On the basis of this information, molecular modeling studies were designed aiming to explore the putative binding modes of these new compounds at the inhibitor-binding site of PfDHODH. In general, several available structures of the DHODH enzyme with bound inhibitors of various sizes show that there is an overlap of the portions of the structures that fit into the H-bond pocket and exhibit polar interactions with H185, Y528 and R265. On the other hand, the size of the hydrophobic pocket is variable, depending on the conformations of the side chains of F171 and F188. In addition, we observed that M536 and Y168 are two additional residues with a high degree of conformational flexibility in the same hydrophobic pocket. Therefore, in order to study the binding mode of the new ligands of variable size, we used the Induced Fit docking protocol (Schrӧdinger package), in which the ligand-binding site flexibility was taken into account to explain the binding modes of ligands varying in size. A summary of docking scores predicted for the interaction of the obtained compounds with PfDHODH (PDB-Id: 1TV5) is given in Table 2, together with their IC50 values.