Emily K. Bremers

Due to the emergence of resistance to both artemisinin derivatives, and their partner drugs, new antimalarials are urgently needed. Ideally, new orally active antimalarials would not only engage new targets but also demonstrate a high barrier to resistance selection, and the ability to kill both proliferating rings and growth-arrested rings resulting from artemisinin exposure. In this report we disclose a novel antimalarial chemotype, the imidazo[4,5-c]pyridine-6-carboxamides, and a representative compound 10b that possesses all these qualities. Orally dosed 10b (4 × 60 mg/kg/day or 1 × 160 mg/kg) cures Plasmodium yoelii-infected mice out to 28 days. This compound is unaffected by over 40 distinct target- and efflux-based resistance mutations in the AReBaR resistome screen, suggesting a novel mode of action. Furthermore, at a minimum inoculum of resistance of 10⁹ parasites, 10b proved refractory to resistance selection. Lastly, with a 6 h exposure, 250 nM 10b kills both proliferating rings and dihydroartemisinin-induced dormant parasites.

Half the world's population is at risk of developing a malaria infection, which is caused by parasites of the genus Plasmodium. Currently, resistance has been identified to all clinically available antimalarials, highlighting an urgent need to develop novel compounds and better understand common mechanisms of resistance. We previously identified a novel tetrahydro-beta-carboline compound, PRC1590, which potently kills the malaria parasite. To better understand its mechanism of action, we selected for and characterized resistance to PRC1590 in Plasmodium falciparum. Through in vitro selection of resistance to PRC1590, we have identified that a single-nucleotide polymorphism on the parasite's multidrug resistance protein 1 (PfMDR1 G293V) mediates resistance to PRC1590. This mutation results in stereospecific resistance and sensitizes parasites to other antimalarials, such as mefloquine, quinine, and MMV019017. Intraerythrocytic asexual stage specificity assays have revealed that PRC1590 is most potent during the trophozoite stage when the parasite forms a single digestive vacuole (DV) and actively digests hemoglobin. Moreover, fluorescence microscopy revealed that PRC1590 disrupts the function of the DV, indicating a potential molecular target associated with this organelle. Our findings mark a significant step in understanding the mechanism of resistance and the mode of action of this emerging class of antimalarials. In addition, our results suggest a potential link between resistance mediated by PfMDR1 and PRC1590's molecular target. This research underscores the pressing need for future research aimed at investigating the intricate relationship between a compound's chemical scaffold, molecular target, and resistance mutations associated with PfMDR1.

The development of parasite resistance to both artemisinin derivatives and their partner drugs jeopardizes the effectiveness of the artemisinin combination therapy. Thus, the discovery of new antimalarial drugs, with new mechanisms of action, is urgently needed. We recently disclosed that β-carboline 1a was orally efficacious in Plasmodium berghei-infected mice and that it showed low cross-resistance between susceptible Plasmodium falciparum and four different drug-resistant strains. In this report, we describe the synthesis and in vitro antimalarial evaluation of 91 new derivatives of 1a. The asexual blood stage growth inhibition data show a clear preference for a 3,4-dihalogenated, 3,5-dihalogenated, 3,4,5-trichloro-, or 4-trifluoromethyphenyl ring at the C1-position. The most potent compound, 3,4,5-trichlorophenyl-substituted 42a, is twice as potent as 1a. Six potent analogues were assessed for their drug-like properties, and four of these were subjected to in vitro barcoded cross-resistance profiling. Compounds 1a, 1m, 42a, and 42m showed no cross-resistance to 32 resistance mutations on the Dd2 genetic background and 10 resistance mutations on the 3D7 genetic background. These data suggest that compounds in this scaffold possess a novel mechanism of antimalarial action.

Cripowellins from Crinum erubescens are known pesticidal and have potent antiplasmodial activity. To gain mechanistic insights to this class of natural products, studies to determine the timing of action of cripowellins within the asexual intraerythrocytic cycle of Plasmodium falciparum were performed and led to the observation that this class of natural products induced reversible cytostasis in the ring stage within the first 24 h of treatment. The transcriptional program necessary for P. falciparum to progress through the asexual intraerythrocytic life cycle is well characterized. Whole transcriptome abundance analysis showed that cripowellin B “pauses” the transcriptional program necessary to progress through the intraerythrocytic life cycle coinciding with the lack of morphological progression of drug treated parasites. In addition, cripowellin B-treated parasites re-enter transcriptional progression after treatment was removed. This study highlights the use of cripowellins as chemical probes to reveal new aspects of cell cycle progression of the asexual ring stage of P. falciparum which could be leveraged for the generation of future antimalarial therapeutics.

The tetrahydro-β-carboline scaffold has proven fertile ground for the discovery of antimalarial agents (e.g., MMV008138 (1) and cipargamin (2)). Similarity searching of a publicly disclosed collection of antimalarial hits for molecules resembling 1 drew our attention to N2-acyl tetrahydro-β-carboline GNF-Pf-5009 ((±)-3b). Compound purchase, “analog by catalog”, and independent synthesis of hits indicated the benzofuran-2-yl amide portion was required for in vitro efficacy against P. falciparum. Preparation of pure enantiomers demonstrated the pharmacological superiority of (R)-3b. Synthesis and evaluation of D- and F-ring substitution variants and benzofuran isosteres indicated a clear structure–activity relationship. Ultimately (R)-3b was tested in Plasmodium berghei-infected mice; unfavorable physicochemical properties may be responsible for the lack of oral efficacy.

The Scientific Teaching (ST) pedagogical framework encompasses many of the best practices recommended in the literature and highlighted in national reports. Understanding the growth and impact of ST requires instruments to accurately measure the extent to which practitioners implement ST in their courses. Researchers have typically relied on students, instructors, or observers to document course teaching practices, but it remains unclear whether and how these perspectives differ from each other. To address this issue, we modified our previously published instrument to generate the Measurement Instrument for Scientific Teaching-Observable (MISTO), which can be completed by students, instructors, and observers, and we investigated the degree of similarity between these three perspectives across 70 undergraduate science courses at seven different institutions in the USA. We found that the full MISTO and Active Learning subcategory scores showed the highest correlations among the three perspectives, but the degree of correlation between perspectives varied for the other subcategories. Match scores between students and instructors were significantly higher than observer matches for the full MISTO and for the Active Learning, Inclusivity, and Responsiveness subcategories. We find that the level and type of agreement between perspectives varies across MISTO subcategories and that this variation likely stems from intrinsic differences in the course access and scoring decisions of the three perspectives. Building on this data, we recommend MISTO users consider their research goals, available resources, and potential artifacts that may arise when deciding which perspective best fits their needs in measuring classroom teaching practices.