Discovery of an anti-malarial compound, burnettiene A, with a multidrug-sensitive Saccharomyces cerevisiae screening system based on mitochondrial function inhibitory activity.

In this paper, we describe our discovery of burnettiene A (1) as an anti-malarial compound from the culture broth of Lecanicillium primulinum (Current name: Flavocillium primulinum) FKI-6715 strain utilizing our original multidrug-sensitive yeast system. This polyene-decalin polyketide natural product was originally isolated as an anti-fungal active compound from Aspergillus burnettii. However, the anti-fungal activity of 1 has been revealed in only one fungal species for and the mechanism of action of 1 remains unknown. After the validation of mitochondrial function inhibitory of 1, we envisioned a new anti-malarial drug discovery platform based on mitochondrial function inhibitory activity. We evaluated anti-malarial activity and 1 showed anti-malarial activity against Plasmodium falciparum FCR3 (chloroquine sensitive) and K1 strain (chloroquine resistant). Our study revealed the utility of our original screening system based on a multidrug-sensitive yeast and mitochondrial function inhibitory activity for the discovery of new anti-malarial drug candidates.

Discovery of antimalarial drugs from secondary metabolites in actinomycetes culture library.

BACKGROUND: Natural products play a key role as potential sources of biologically active substances for the discovery of new drugs. This study aimed to identify secondary metabolites from actinomycete library extracts that are potent against the asexual stages of Plasmodium falciparum (P. falciparum). METHODS: Secondary metabolites from actinomycete library extracts were isolated from culture supernatants by ethyl acetate extraction. Comprehensive screening was performed to identify novel antimalarial compounds from the actinomycete library extracts (n = 28). The antimalarial activity was initially evaluated in vitro against chloroquine/mefloquine-sensitive (3D7) and-resistant (Dd2) lines of P. falciparum. The cytotoxicity was then evaluated in primary adult mouse brain (AMB) cells. RESULTS: Out of the 28 actinomycete extracts, 17 showed parasite growth inhibition > 50% at a concentration of 50 µg/mL, nine were identified with an IC50 value < 10 µg/mL, and seven suppressed the parasite significantly with an IC50 value < 5 µg/mL. The extracts from Streptomyces aureus strains HUT6003 (Extract ID number: 2), S. antibioticus HUT6035 (8), and Streptomyces sp. strains GK3 (26) and GK7 (27), were found to have the most potent antimalarial activity with IC50 values of 0.39, 0.09, 0.97, and 0.36 µg/mL (against 3D7), and 0.26, 0.22, 0.72, and 0.21 µg/mL (against Dd2), respectively. Among them, Streptomyces antibioticus strain HUT6035 (8) showed the highest antimalarial activity with an IC50 value of 0.09 µg/mL against 3D7 and 0.22 µg/mL against Dd2, and a selective index (SI) of 188 and 73.7, respectively. CONCLUSION: Secondary metabolites obtained from the actinomycete extracts showed promising antimalarial activity in vitro against 3D7 and Dd2 cell lines of P. falciparum with minimal toxicity. Therefore, secondary metabolites obtained from actinomycete extracts represent an excellent starting point for the development of antimalarial drug leads.

Natural Peroxides from Plants: Historical Discovery, Biosynthesis, and Biological Activities.

Naturally occurring peroxides received great interest and attention from scientific research groups worldwide due to their structural diversity, versatile biological activities, and pharmaceutical properties. In the present review, we describe the historical discovery of natural peroxides from plants systematically and update the researchers with recently explored ones justifying their structural caterogrization and biological/pharmaceutical properties intensively. Till the end of 2023, 192 peroxy natural products from plants were documented herein for the first time implying most categories of natural scaffolds (e.g. terpenes, polyketides, phenolics and alkaloids). Numerically, the reported plants' peroxides have been classified into seventy-four hydro-peroxides, hundred seven endo-peroxides and eleven acyl-peroxides.  Endo-peroxides (cyclic alkyl peroxides) are an important group due to their high variety of structural frameworks, and we have further divided them into "four-, five-, six and seven"-membered rings. Biosynthetically, a shedding light on the intricate mechanisms behind the formation of plant-derived peroxides are addressed as well.

Antimalarial and antioxidant activities of novel artesunate-ellagic acid hybrid compound in vitro and in vivo.

Introduction: Emergence of drug resistant strains of Plasmodium species has necessitated the search for novel antimalarials with unique mechanisms of action. Synthesis of hybrid compounds has been one approach to tackling this challenge. In this study, the synthesis of artesunate-ellagic acid hybrid compound (EA31) from ellagic acid and artesunate and its evaluation for antimalarial and antioxidant activities using in vitro and in vivo models were carried out. Method: EA31 was synthesized from artesunate and ellagic acid. The activities of the hybrid compound against Plasmodium falciparum W2 and P. berghei NK65 were evaluated, and its antioxidant activities were also determined. Results: The results revealed that EA31 was more active against P. falciparum W2 (chloroquine resistant) clone and less cytotoxic to buffalo green monkey kidney cell line compared to artesunate. EA31 was also active against Plasmodium berghei NK65 in vivo. The results revealed inhibition of ß-hematin formation as one of the mechanisms of action of EA31. EA31 also exhibited antioxidant activities. Conclusion: The results revealed that EA31 may exert dual action of killing malaria parasite and mopping the reactive oxygen species that mediate the secondary complications of malaria.

[Biological threats to global malaria elimination I Antimalarial drug resistance].

Malaria is an infectious disease that seriously threatens human health. Currently, malaria control mainly depends on antimalarial chemotherapy. However, antimalarial drug resistance is becoming increasingly severe, which poses a great challenge to malaria control, notably treatment of Plasmodium falciparum malaria. To address this challenge, there is a need to facilitate development of novel antimalarial drugs and innovation of treatment strategies, as well as reinforce surveillance and research on antimalarial drug resistance. This article reviews the main categories and use guidelines of current antimalarial agents, summarizes the current status and monitoring methods of antimalarial drug resistance, and proposes the response to antimalarial drug resistance, so as to provide insights into the use of antimalarial drugs and response to antimalarial drug resistance, and contribute to global malaria elimination.

Expression and biochemical characterization of the putative insulinase enzyme PF11_0189 found in the Plasmodium falciparum genome.

PF11_0189 is a putative insulin degrading enzyme present in Plasmodium falciparum genome. The catalytic domain of PF11_0189 is about 27 kDa. Substrate specificity study shows PF11_0189 acts upon different types of proteins. The substrate specificity is found to be highest when insulin is used as a substrate. Metal dependency study shows highest dependency of PF11_0189 towards zinc metal for its proteolytic activity. Chelation of zinc metal with EDTA shows complete absence of PF11_0189 activity. Peptide inhibitors, P-70 and P-121 from combinatorial peptide library prepared against PF11_0189 show inhibition with an IC50 value of 4.8 µM and 7.5 µM respectively. A proven natural anti-malarial peptide cyclosporin A shows complete inhibition against PF11_0189 with an IC50 value of 0.75 µM suggesting PF11_0189 as a potential target for peptide inhibitors. The study implicates that PF11_0189 is a zinc metalloprotease involved in catalysis of insulin. The study gives a preliminary insight into the mechanism of complications arising from glucose abnormalities during severe malaria.

Photoaffinity probe-based antimalarial target identification of artemisinin in the intraerythrocytic developmental cycle of Plasmodium falciparum.

Malaria continues to pose a serious global health threat, and artemisinin remains the core drug for global malaria control. However, the situation of malaria resistance has become increasingly severe due to the emergence and spread of artemisinin resistance. In recent years, significant progress has been made in understanding the mechanism of action (MoA) of artemisinin. Prior research on the MoA of artemisinin mainly focused on covalently bound targets that are alkylated by artemisinin-free radicals. However, less attention has been given to the reversible noncovalent binding targets, and there is a paucity of information regarding artemisinin targets at different life cycle stages of the parasite. In this study, we identified the protein targets of artemisinin at different stages of the parasite's intraerythrocytic developmental cycle using a photoaffinity probe. Our findings demonstrate that artemisinin interacts with parasite proteins in vivo through both covalent and noncovalent modes. Extensive mechanistic studies were then conducted by integrating target validation, phenotypic studies, and untargeted metabolomics. The results suggest that protein synthesis, glycolysis, and oxidative homeostasis are critically involved in the antimalarial activities of artemisinin. In summary, this study provides fresh insights into the mechanisms underlying artemisinin's antimalarial effects and its protein targets.

Exploring antimalarial potential: Conjugating organometallic moieties with organic fragments for enhanced efficacy.

In the search for novel ligands with efficacy against various diseases, particularly parasitic diseases, molecular hybridization of organometallic units into biologically active scaffolds has been hailed as an appealing strategy in medicinal chemistry. The conjugation to organometallic fragments can be achieved by an appropriate linker or by directly coordinating the existing drugs to a metal. The success of Ferroquine (FQ, SR97193), an effective chloroquine-ferrocene conjugate currently undergoing the patient-exploratory phase as a combination therapy with the novel triaminopyrimidine ZY-19489 for malaria, has sparked intense interest in organometallic compound drug discovery. We present the evolution of organometallic antimalarial agents over the last decade, focusing on the parent moiety's class and the type of organometallics involved. Four main organometallic antimalarial compounds have been chosen based on conjugated organic moieties: existing antimalarial drugs, other clinical drugs, hybrid drugs, and promising scaffolds of thiosemicarbazones, benzimidazoles, and chalcones, in particular. The presented insights contribute to the ongoing discourse on organometallic compound drug development for malaria diseases.

Indoloquinoline Alkaloids as Antimalarials: Advances, Challenges, and Opportunities.

Malaria infections affect almost half of the world's population, with over 200 million cases reported annually. Cryptolepis sanguinolenta, a plant native to West Africa, has long been used across various regions of Africa for malaria treatment. Chemical analysis has revealed that the plant is abundant in indoloquinolines, which have been shown to possess antimalarial properties. Cryptolepine, neocryptolepine, and isocryptolepine are well-studied indoloquinoline alkaloids known for their potent antimalarial activity. However, their structural rigidity and associated cellular toxicity are major drawbacks for preclinical development. This review focuses on the potential of indoloquinoline alkaloids (cryptolepine, neocryptolepine, and isocryptolepine) as scaffolds in drug discovery. The article delves into their antimalarial effects in vitro and in vivo, as well as their proposed mechanisms of action and structure-activity relationship studies. Several studies aim to improve these leads by reducing cytotoxicity while preserving or enhancing antimalarial activity and gaining insights into their mechanisms of action. These investigations highlight the potential of indoloquinolines as a scaffold for developing new antimalarial drugs.

Experimental and model-based approach to evaluate solvent effects on the solubility of the pharmaceutical artemisinin.

The separation and purification of plant-based Active Pharmaceutical Ingredients (API) from extracts is a crucial part in pharmaceutical process development. For the purification of the antimalarial drug component artemisinin (ARTE) from an Artemisia anna L. toluene extract, antisolvent crystallization is considered. Solubilities of ARTE in binary solvent mixtures of toluene and two potential antisolvents, n-heptane and ethanol, were determined at temperatures from 278.15 K to 313.15 K. The experimental work was supported by the application of various models, utilizing varying amounts of experimental input data. The goal was the identification of models that are able to predict solubilities in binary solvent mixtures sufficiently accurate and, thus, can help to reduce the experimental effort for future solvent screenings. In this study, we applied the PC-SAFT model both with and without fitting the binary interaction parameter kij between ARTE and the respective solvent, as well as the empirical Jouyban-Acree model. From the experiments, n-heptane demonstrated to be a promising antisolvent, while ethanol acted more as a cosolvent. All models tested were capable of distinguishing between effective and ineffective antisolvents. The purely predictive PC-SAFT model applied with kij = 0 exhibited the largest deviation from the experimental data. This was followed by the PC-SAFT model including fitted kij values, based on at least four experimental data points. The Jouyban-Acree model fitted the data most accurately. Its parametrization required a minimum of ten experimental data points.

Establishment and evaluation of a qPCR method for the detection of pfmdr1 mutations in Plasmodium falciparum, the causal agent of fatal malaria.

Drug resistance surveillance is a major integral part of malaria control programs. Molecular methods play a pivotal role in drug resistance detection and related molecular research. This study aimed to develop a rapid and accurate detection method for drug resistance of Plasmodium falciparum (P. falciparum). A quantitative real-time PCR (qPCR) assay has been developed that identifies the mutation at locus A256T in the P.falciparum multi-drug resistance(pfmdr1) gene producing amino acid change at position 86. The results of 198 samples detected by qPCR were consistent with nested PCR and sequencing, giving an accuracy of 94.3%. The sensitivity, specificity, positive and negative predictive value of qPCR were 85.7%, 97.6%, 90.0% and 96.4%, respectively. The results of qPCR are basically consistent with the nested PCR, which is expected to replace the nested PCR as a new molecular biological method for drug resistance detection, providing reliable technical support for global malaria prevention and control.

Optimization of diastereomeric dihydropyridines as antimalarials.

The increase in research funding for the development of antimalarials since 2000 has led to a surge of new chemotypes with potent antimalarial activity. High-throughput screens have delivered several thousand new active compounds in several hundred series, including the 4,7-diphenyl-1,4,5,6,7,8-hexahydroquinolines, hereafter termed dihydropyridines (DHPs). We optimized the DHPs for antimalarial activity. Structure-activity relationship studies focusing on the 2-, 3-, 4-, 6-, and 7-positions of the DHP core led to the identification of compounds potent (EC50 < 10 nM) against all strains of P. falciparum tested, including the drug-resistant parasite strains K1, W2, and TM90-C2B. Evaluation of efficacy of several compounds in vivo identified two compounds that reduced parasitemia by >75 % in mice 6 days post-exposure following a single 50 mg/kg oral dose. Resistance acquisition experiments with a selected dihydropyridine led to the identification of a single mutation conveying resistance in the gene encoding for Plasmodium falciparum multi-drug resistance protein 1 (PfMDR1). The same dihydropyridine possessed transmission blocking activity. The DHPs have the potential for the development of novel antimalarial drug candidates.

Varied prevalence of antimalarial drug resistance markers in different populations of newly arrived refugees in Uganda.

Newly arrived refugees offer insights into malaria epidemiology in their countries of origin. We evaluated asymptomatic refugee children within 7 days of arrival in Uganda from South Sudan and the Democratic Republic of Congo (DRC) in 2022 for parasitemia, parasite species, and Plasmodium falciparum drug resistance markers. Asymptomatic P. falciparum infections were common in both populations. Co-infection with P. malariae was more common in DRC refugees. Prevalences of markers of aminoquinoline resistance (PfCRT K76T, PfMDR1 N86Y) were much higher in South Sudan refugees, of antifolate resistance (PfDHFR C59R and I164L, PfDHPS A437G and K540E) much higher in DRC refugees, and of artemisinin partial resistance (ART-R; PfK13 C469Y and A675V) moderate in both populations. Prevalences of most mutations differed from those seen in Ugandans attending health centers near the refugee centers. Refugee evaluations yielded insights into varied malaria epidemiology and identified markers of ART-R in two previously little-studied countries.

Ganopyrone A, a highly rearranged lanostane triterpenoid with antimalarial activity from artificially cultivated fruiting bodies of Ganoderma colossus.

Three previously undescribed highly modified lanostane triterpenoids, ganopyrone A, ganocolossusin I, and ganodermalactone Y, were isolated from the artificially cultivated fruiting bodies of the basidiomycete Ganoderma colossus TBRC-BCC 17711. Ganopyrone A possesses an unprecedented polycyclic carbon skeleton with an α-pyrone ring and C-18/C-23 bond. It showed antimalarial activity against Plasmodium falciparum K1 (multidrug-resistant strain) with an IC50 value of 7.8 µM (positive control: dihydroartemisinin, IC50 1.4 nM), while its cytotoxicity (Vero cells) was much weaker (IC50 103 µM).

Anti-malarial and Multi-bioactive Co (II), Cu (II) and Ni (II) Salen Complexes: Synthesis, Characterization and Computational Studies.

Herein, we report anti-malarial, anti-bacterial and anti-inflammatory activities of theN2O2 donor tetradentate salen ligand and its CoL, NiL, and CuL metal complexes. All the compoundswere synthesized and characterized by various spectroscopic analytical methods.Thein-vitro antimalarial investigations revealed that the complex CuL exhibited equipotency with quinine drug having IC50 value 0.25 µg/mL.The compound L shows significant inhibition of bacterial spp.viz.E. Coli, P. Aeruginosa, and S. Aureus (MIC=12.5-50µg/mL), while the compound CoL (MIC=12.5µg/mL) exhibited potency against gram-positive bacteria. In the in-vitro anti-inflammatory study, the compound CuL has moderate activity than other tested compounds. The compound CuL showedthe highest anti-malarial docking score with enzyme pLDH at -8.12 Kcal/mol. The DFT study also gives authentication of higher antimalarial activity of CuL due to high dipole moment. None of the potent compound was found cytotoxic towards verocell lines.

Cu-catalyzed click reaction in synthesis of eugenol derivatives as potent antimalarial agents.

Eugenol(1), a terpenoid found in Ocimum, has various biological activities. The present study aims at extraction, isolation of the plant secondary metabolite eugenol (1), it's derivatisation and structure identification as bioactive molecules. Synthesis and antiplasmodial activity (in-vitro and in-vivo), of a series of fourteen novel eugenol-based 1,2,3-triazole derivatives was done in the present study. Derivatives 5a-5n showed good antimalarial activity against the strain Plasmodium falciparum NF54. Derivative 5 m, IC50 at 2.85 µM was found to be several times better than its precursor 1 (106.82 µM) whereas the derivative 5n showed three fold better activity than compound 1, in vitro. The structure-activity relationship of the synthesised compounds indicated that the presence of triazole ring in eugenol analogues is responsible for their good activity. Compound 5m, was further evaluated for in-vivo antimalarial activity which showed about 79% parasitemia suppression. It is the first report on antimalarial activity of triazole eugenol derivatives.

Protein target similarity is positive predictor of in vitro antipathogenic activity: a drug repurposing strategy for Plasmodium falciparum.

Drug discovery is an intricate and costly process. Repurposing existing drugs and active compounds offers a viable pathway to develop new therapies for various diseases. By leveraging publicly available biomedical information, it is possible to predict compounds' activity and identify their potential targets across diverse organisms. In this study, we aimed to assess the antiplasmodial activity of compounds from the Repurposing, Focused Rescue, and Accelerated Medchem (ReFRAME) library using in vitro and bioinformatics approaches. We assessed the in vitro antiplasmodial activity of the compounds using blood-stage and liver-stage drug susceptibility assays. We used protein sequences of known targets of the ReFRAME compounds with high antiplasmodial activity (EC50 < 10 uM) to conduct a protein-pairwise search to identify similar Plasmodium falciparum 3D7 proteins (from PlasmoDB) using NCBI protein BLAST. We further assessed the association between the compounds' in vitro antiplasmodial activity and level of similarity between their known and predicted P. falciparum target proteins using simple linear regression analyses. BLAST analyses revealed 735 P. falciparum proteins that were similar to the 226 known protein targets associated with the ReFRAME compounds. Antiplasmodial activity of the compounds was positively associated with the degree of similarity between the compounds' known targets and predicted P. falciparum protein targets (percentage identity, E value, and bit score), the number of the predicted P. falciparum targets, and their respective mutagenesis index and fitness scores (R2 between 0.066 and 0.92, P < 0.05). Compounds predicted to target essential P. falciparum proteins or those with a druggability index of 1 showed the highest antiplasmodial activity.

Antimalarial activity of cecropin antimicrobial peptides derived from Anopheles mosquitoes.

The emergence of clinically drug-resistant malaria parasites requires the urgent development of new drugs. Mosquitoes are vectors of multiple pathogens and have developed resistance mechanisms against them, which often involve antimicrobial peptides (AMPs). An-cecB is an AMP of the malaria-transmitting mosquito genus Anopheles, and we herein report its antimalarial activity against Plasmodium falciparum 3D7, the artemisinin-resistant strain 803, and the chloroquine-resistant strain Dd2 in vitro. We also demonstrate its anti-parasite activity in vivo, using the rodent malaria parasite Plasmodium berghei (ANKA). We show that An-cecB displays potent antimalarial activity and that its mechanism of action may occur through direct killing of the parasite or through interaction with infected red blood cell membranes. Unfortunately, An-cecB was found to be cytotoxic to mammalian cells and had poor antimalarial activity in vivo. However, its truncated peptide An-cecB-1 retained most of its antimalarial activity and avoided its cytotoxicity in vitro. An-cecB-1 also showed better antimalarial activity in vivo. Mosquito-derived AMPs may provide new ideas for the development of antimalarial drugs against drug-resistant parasites, and An-cecB has potential use as a template for antimalarial peptides.

Unlocking nitrogen compounds' promise against malaria: A comprehensive review.

Plasmodium parasites are the primary cause of malaria, leading to high mortality rates, which require clinical attention. Many of the medications used in the treatment have resulted in resistance over time. Artemisinin combination therapy (ACT) has shown significant results for the treatment. However, mutations in the parasite have resulted in resistance, leading to decreased efficiency of the medications that are currently being used. Therefore, there is a critical need to find novel scaffolds that are safe, effective, and of economic advantage. Literature has reported several potent molecules with diverse scaffolds designed, synthesized, and evaluated against different strains of Plasmodium. With this growing list of compounds, it is essential to collect the data in one place to gain a concise overview of the emerging scaffolds in recent years. For this purpose, nitrogen-containing heterocycles such as ß-carboline, imidazole, quinazoline, quinoline, thiazole, and thiophene have been highly explored due to their wide biological applications. Besides these, another scaffold, benzodiazepine, which is majorly used as a central nervous system depressant, is emerging as an anti-malarial agent. Hence, this review centers on the latest medication advancements designed to combat malaria, emphasizing special attention to 1,4-benzodiazepines as a novel scaffold for antimalarial drug discovery.

3-Position Biaryl Endochin-like Quinolones with Enhanced Antimalarial Performance.

ELQ-300 is a potent antimalarial drug with activity against blood, liver, and vector stages of the disease. A prodrug, ELQ-331, exhibits reduced crystallinity and improved in vivo efficacy in preclinical testing, and currently, it is in the developmental pipeline for once-a-week dosing for oral prophylaxis against malaria. Because of the high cost of developing a new drug for human use and the high risk of drug failure, it is prudent to have a back-up plan in place. Here we describe ELQ-596, a member of a new subseries of 3-biaryl-ELQs, with enhanced potency in vitro against multidrug-resistant Plasmodium falciparum parasites. ELQ-598, a prodrug of ELQ-596 with diminished crystallinity, is more effective vs murine malaria than its progenitor ELQ-331 by 4- to 10-fold, suggesting that correspondingly lower doses could be used to protect and cure humans of malaria. With a longer bloodstream half-life in mice compared to its progenitor, ELQ-596 highlights a novel series of next-generation ELQs with the potential for once-monthly dosing for protection against malaria infection. Advances in the preparation of 3-biaryl-ELQs are presented along with preliminary results from experiments to explore key structure-activity relationships for drug potency, selectivity, pharmacokinetics, and safety.