Plasmodium falciparum causes the most lethal and widespread form of malaria. Eradication of malaria remains a priority due to the increasing number of cases of drug resistance. The heat shock protein 90 of P. falciparum (PfHsp90) is a validated drug target essential for parasite survival. Most PfHsp90 inhibitors bind at the ATP binding pocket found in its N-terminal domain, abolishing the chaperone's activities, which leads to parasite death. The challenge is that the NTD of PfHsp90 is highly conserved, and its disruption requires selective inhibitors that can act without causing off-target human Hsp90 activities. We endeavored to discover selective inhibitors of PfHsp90 using pharmacophore modeling, virtual screening protocols, induced fit docking (IFD), and cell-based and biochemical assays. The pharmacophore model (DHHRR), composed of one hydrogen bond donor, two hydrophobic groups, and two aromatic rings, was used to mine commercial databases for initial hits, which were rescored to 20 potential hits using IFD. Eight of these compounds displayed moderate to high activity toward P. falciparum NF54 (i.e., IC₅₀s ranging from 6.0 to 0.14 μM) and averaged >10 in terms of selectivity indices toward CHO and HepG2 cells. Additionally, four compounds inhibited PfHsp90 with greater selectivity than a known inhibitor, harmine, and bound to PfHsp90 with weak to moderate affinity. Our findings support the use of a pharmacophore model to discover diverse chemical scaffolds such as FM2, FM6, F10, and F11 exhibiting anti-Plasmodium activities and serving as valuable new PfHsp90 inhibitors. Optimization of these hits may enable their development into potent leads for future antimalarial drugs.

Group A rotaviruses (RVA) represent the most common cause of pediatric gastroenteritis in children <5 years, worldwide. There has been an increase in global detection and reported cases of acute gastroenteritis caused by RVA genotype G12 strains, particularly in Africa. This study sought to characterize the genomic relationship between African G12 strains and determine the possible origin of these strains. Whole genome sequencing of 34 RVA G12P[6] and G12P[8] strains detected from the continent including southern (South Africa, Zambia, Zimbabwe), eastern (Ethiopia, Uganda), central (Cameroon), and western (Togo) African regions, were sequenced using the Ion Torrent PGM method. The majority of the strains possessed a Wa-like backbone with consensus genotype constellation of G12-P[6]/P[8]-I1-R1-C1-M1-A1-N1-T1-E1-H1, while a single strain from Ethiopia displayed a DS-1-like genetic constellation of G12-P[6]-I2-R2-C2-M2-A2-N2-T2-E2-H2. In addition, three Ethiopian and one South African strains exhibited a genotype 2 reassortment of the NSP3 gene, with genetic constellation of G12-P[8]-I1-R1-C1-M1-A1-N1-T2-E1-H1. Overall, 10 gene segments (VP1–VP4, VP6, and NSP1–NSP5) of African G12 strains were determined to be genetically related to cognate gene sequences from globally circulating human Wa-like G12, G9, and G1 strains with nucleotide (amino acid) identities in the range of 94.1–99.9% (96.5–100%), 88.5–98.5% (93–99.1%), and 89.8–99.0% (88.7–100%), respectively. Phylogenetic analysis showed that the Ethiopian G12P[6] possessing a DS-1-like backbone consistently clustered with G2P[4] strains from Senegal and G3P[6] from Ethiopia with the VP1, VP2, VP6, and NSP1–NSP4 genes. Notably, the NSP2, NSP3, and NSP4 of most of the study strains exhibited the closest relationship with porcine strains suggesting the occurrence of reassortment between human and porcine strains. Our results add to the understanding of potential roles that interspecies transmission play in generating human rotavirus diversity through reassortment events and provide insights into the evolutionary dynamics of G12 strains spreading across selected sub-Saharan Africa regions.

Plasmodium falciparum is the most lethal malaria parasite. Increasing incidences of drug resistance of P. falciparum have prompted the need for discovering new and effective antimalarial compounds with an alternative mode of action. Heat shock protein 90 (PfHsp90) facilitates protein folding and is a promising antimalarial drug target. We have previously reported that iso-mukaadial acetate (IMA) and ursolic acid acetate (UAA) exhibit antimalarial activity. We investigated the abilities of IMA and UAA to bind PfHsp90 by molecular docking and dynamics simulations. The in silico predictions were validated by biochemical assays conducted on recombinant PfHsp90. The interaction between the ligands and PfHsp90 was evaluated using ultraviolet-visible spectroscopy (UV-vis), Fourier transform infrared (FTIR), and surface plasmon resonance (SPR) analysis. The results obtained by docking calculations and MD dynamics simulation predicted that UAA and IMA preferentially bound to PfHsp90 via the N-terminal domain, with UAA binding more stable than IMA. UV-vis-based data suggest that PfHsp90 harbors buried aromatic amino acids, which were exposed in the presence of either IMA or UAA. In addition, data obtained using FTIR suggested that IMA and UAA destabilized the secondary structure of PfHsp90. Of the two compounds, UAA bound to PfHsp90 within the micromolar range based on surface plasmon resonance (SPR)-based binding assay. Furthermore, both compounds disrupted the holdase chaperone function of PfHsp90 as the chaperone failed to suppress heat-induced aggregation of the model proteins, malate dehydrogenase (MDH), luciferase, and citrate synthase in vitro. In addition, both compounds lowered the ATPase activity of PfHsp90. The molecular dynamics simulation analysis indicated that the docked complexes were mostly stable for 100 ns, validating the data obtained through the biochemical assays. Altogether, this study expands the repository of antiplasmodial compounds that have PfHsp90 among their possible targets.

Malaria which is mainly caused by Plasmodium falciparum parasite remains a devastating public health concern, necessitating the need to develop new antimalarial agents. P. falciparum heat shock protein 90 (Hsp90), is indispensable for parasite survival and a promising drug target. Inhibitors targeting the ATP-binding pocket of the N-terminal domain have anti-Plasmodium effects. We proposed a de novo active learning (AL) driven method in tandem with docking to predict inhibitors with unique scaffolds and preferential selectivity towards PfHsp90. Reference compounds, predicted to bind PfHsp90 at the ATP-binding pocket and possessing anti-Plasmodium activities, were used to generate 10,000 unique derivatives and to build the Auto-quantitative structures activity relationships (QSAR) models. Glide docking was performed to predict the docking scores of the derivatives and > 15,000 compounds obtained from the ChEMBL database. Re-iterative training and testing of the models was performed until the optimum Kennel-based Partial Least Square (KPLS) regression model with a regression coefficient R2 = 0.75 for the training set and squared correlation prediction Q2 = 0.62 for the test set reached convergence. Rescoring using induced fit docking and molecular dynamics simulations enabled us to prioritize 15 ATP/ADP-like design ideas for purchase. The compounds exerted moderate activity towards P. falciparum NF54 strain with IC₅₀ values of ≤ 6μM and displayed moderate to weak affinity towards PfHsp90 (K_D range: 13.5–19.9μM) comparable to the reported affinity of ADP. The most potent compound was FTN-T5 (PfN54 IC₅₀:1.44μM; HepG2/CHO cells SI≥ 29) which bound to PfHsp90 with moderate affinity (K_D:7.7μM), providing a starting point for optimization efforts. Our work demonstrates the great utility of AL for the rapid identification of novel molecules for drug discovery (i.e., hit identification). The potency of FTN-T5 will be critical for designing species-selective inhibitors towards developing more efficient agents against malaria.

The causative agent of malaria, Plasmodium falciparum, encodes four heat shock protein 90 isoforms in the cytosol, endoplasmic reticulum, mitochondria and apicoplast. PfHsp90s are considered potential targets for developing antimalarial drugs. However, the similarity between the druggable ATP binding pocket of these isoforms and their human counterparts has hindered efforts in discovering Hsp90-based antimalarial drugs. There is widespread concern that the chemotypes targeting PfHsp90 isoforms may not possess the selectivity required for translatability. Most studies have focused on the cytosolic (canonical Hsp90) and have used various inhibitors of Hsp90 to conduct anti-Plasmodium and mammalian safety studies without considering the on-target enzymatic activity and binding affinity. The extent to which the cytosolic Hsp90 shares common mechanisms with the other isoforms remains elusive. As such, detailed structural comparisons of the Hsp90 isoforms may reveal exploitable differences that could favour preferential binding. It is essential to consider whether molecules that act as pan-inhibitors would put more pressure on the parasite or have detrimental effects. Studies should go beyond molecular docking and whole-cell activity to advance the development of Hsp90 inhibitors as future antimalarials. There is a need also to assess the physicochemical properties, drug metabolism, pharmacokinetics, and In vivo proof-of-concept. We argue that the potential benefits of PfHsp90 isoform inhibitors outweigh the off-target and selectivity risks. Additionally, opportunities for using machine learning and computer-aided drug discovery efforts to design inhibitors that preferentially bind to each isoform, particularly the cytosolic PfHsp90, are highlighted as a means of overcoming the resistance challenge.

Malaria which is mainly caused by Plasmodium falciparum parasite remains a devastating public health concern, necessitating the need to develop new antimalarial agents. P. falciparum heat shock protein 90 (Hsp90), is indispensable for parasite survival and a promising drug target. Inhibitors targeting the ATP-binding pocket of the N-terminal domain have anti- Plasmodium effects. We proposed a de novo active learning (AL) driven method in tandem with docking to predict inhibitors with unique scaffolds and preferential selectivity towards PfHsp90. Reference compounds, predicted to bind PfHsp90 at the ATP-binding pocket and possessing anti- Plasmodium activities, were used to generate 10,000 unique derivatives and to build the Auto-quantitative structures activity relationships (QSAR) models. Glide docking was performed to predict the docking scores of the derivatives and > 15,000 compounds obtained from the ChEMBL database. Re-iterative training and testing of the models was performed until the optimum Kennel-based Partial Least Square (KPLS) regression model with a regression coefficient R2 = 0.75 for the training set and squared correlation prediction Q2 = 0.62 for the test set reached convergence. Rescoring using induced fit docking and molecular dynamics simulations enabled us to prioritize 15 ATP/ADP-like design ideas for purchase. The compounds exerted moderate activity towards P. falciparum NF54 strain with IC ₅₀ values of ≤ 6μM and displayed moderate to weak affinity towards PfHsp90 (K _D range: 13.5-19.9μM) comparable to the reported affinity of ADP. The most potent compound was FTN-T5 (PfN54 IC ₅₀ :1.44μM; HepG2/CHO cells SI≥ 29) which bound to PfHsp90 with moderate affinity (K _D :7.7μM), providing a starting point for optimization efforts. Our work demonstrates the great utility of AL for the rapid identification of novel molecules for drug discovery (i.e., hit identification). The potency of FTN-T5 will be critical for designing species-selective inhibitors towards developing more efficient agents against malaria.