Optimization of Plasmodium vivax infection of colonized Amazonian Anopheles darlingi.

Obtaining Plasmodium vivax sporozoites is essential for in vitro culture of liver stage parasites, not only to understand fundamental aspects of parasite biology, but also for drug and vaccine development. A major impediment to establish high-throughput in vitro P. vivax liver stage assays for drug development is obtaining sufficient numbers of sporozoites. To do so, female anopheline mosquitoes have to be fed on blood from P. vivax-infected patients through an artificial membrane-feeding system, which in turns requires a well-established Anopheles colony. In this study we established conditions to provide a robust supply of P. vivax sporozoites. Adding a combination of serum replacement and antibiotics to the membrane-feeding protocol was found to best improve sporozoite production. A simple centrifugation method appears to be a possible tool for rapidly obtaining purified sporozoites with a minimal loss of yield. However, this method needs to be better defined since sporozoite viability and hepatocyte infection were not evaluated.

Terpenes extracted from marine sponges with antioxidant activity: a systematic review.

Marine biodiversity has emerged as a very promising resource of bioactive compounds and secondary metabolites from different sea organisms. The sponge's secondary metabolites demonstrated various bioactivities and potential pharmacological properties. This systematic review of the literature focuses on the advances achieved in the antioxidant potential of marine sponges in vitro. The review was performed in accordance with PRISMA guidelines. The main inclusion criterion for analysis was articles with identification of compounds from terpene classes that demonstrate antioxidant activity in vitro. Searching in three different databases, two hundred articles were selected. After screening abstracts, titles and evaluating for eligibility of manuscripts 14 articles were included. The most performed analyzes to detect antioxidant activity were scavenging activity 2,2-diphenyl-1-picrylhydrazyl (DPPH) and measurement of intracellular reactive oxygen species (ROS). It was possible to identify 17 compounds of the terpene class with pronounced antioxidant activity in vitro. Scientific evidence of the studies included in this review was accessed by the GRADE analysis. Terpenes play an important ecological role, moreover these molecules have a pharmaceutical and industrial application.

Transmission-blocking activity of antimalarials for Plasmodium vivax malaria in Anopheles darlingi.

Malaria is caused by parasite of the genus Plasmodium and is still one of the most important infectious diseases in the world. Several biological characteristics of Plasmodium vivax contribute to the resilience of this species, including early gametocyte production, both of which lead to efficient malaria transmission to mosquitoes. This study evaluated the impact of currently used drugs on the transmission of P. vivax. Participants received one of the following treatments for malaria: i) chloroquine [10 mg/kg on day 1 and 7.5 mg/kg on day 2 and 3] co-administered with Primaquine [0.5 mg/kg/day for 7 days]; ii) Chloroquine [10 mg/kg on day 1 and 7.5 mg/kg on day 2 and 3] co-administered with one-dose of Tafenoquine [300 mg on day 1]; and iii) Artesunate and Mefloquine [100 mg and 200 mg on day 1, 2 and 3] co-administered with Primaquine [0.5 mg/kg/day for 14 days]. Patient blood was collected before treatment and 4 h, 24 h, 48 h and 72 h after treatment. The blood was used to perform a direct membrane feeding assay (DMFA) using Anopheles darlingi mosquitoes. The results showed 100% inhibition of the mosquito infection after 4 h using ASMQ+PQ, after 24 h for the combination of CQ+PQ and 48 h using CQ+TQ. The density of gametocytes declined over time in all treatment groups, although the decline was more rapid in the ASMQ+PQ group. In conclusion, it was possible to demonstrate the transmission-blocking efficacy of the malaria vivax treatment and that ASMQ+PQ acts faster than the two other treatments.

Antiplasmodial, Trypanocidal, and Genotoxicity In Vitro Assessment of New Hybrid α,α-Difluorophenylacetamide-statin Derivatives.

BACKGROUND: Statins present a plethora of pleiotropic effects including anti-inflammatory and antimicrobial responses. A,α-difluorophenylacetamides, analogs of diclofenac, are potent pre-clinical anti-inflammatory non-steroidal drugs. Molecular hybridization based on the combination of pharmacophoric moieties has emerged as a strategy for the development of new candidates aiming to obtain multitarget ligands. METHODS: Considering the anti-inflammatory activity of phenylacetamides and the potential microbicidal action of statins against obligate intracellular parasites, the objective of this work was to synthesize eight new hybrid compounds of α,α-difluorophenylacetamides with the moiety of statins and assess their phenotypic activity against in vitro models of Plasmodium falciparum and Trypanosoma cruzi infection besides exploring their genotoxicity safety profile. RESULTS: None of the sodium salt compounds presented antiparasitic activity and two acetated compounds displayed mild anti-P. falciparum effect. Against T. cruzi, the acetate halogenated hybrids showed moderate effect against both parasite forms relevant for human infection. Despite the considerable trypanosomicidal activity, the brominated compound revealed a genotoxic profile impairing future in vivo testing. CONCLUSIONS: However, the chlorinated derivative was the most promising compound with chemical and biological profitable characteristics, without presenting genotoxicity in vitro, being eligible for further in vivo experiments.

Population-based genomic study of Plasmodium vivax malaria in seven Brazilian states and across South America.

Background: Brazil is a unique and understudied setting for malaria, with complex foci of transmission associated with human and environmental conditions. An understanding of the population genomic diversity of P. vivax parasites across Brazil can support malaria control strategies. Methods: Through whole genome sequencing of P. vivax isolates across 7 Brazilian states, we use population genomic approaches to compare genetic diversity within country (n = 123), continent (6 countries, n = 315) and globally (26 countries, n = 885). Findings: We confirm that South American isolates are distinct, have more ancestral populations than the other global regions, with differentiating mutations in genes under selective pressure linked to antimalarial drugs (pvmdr1, pvdhfr-ts) and mosquito vectors (pvcrmp3, pvP45/48, pvP47). We demonstrate Brazil as a distinct parasite population, with signals of selection including ABC transporter (PvABCI3) and PHIST exported proteins. Interpretation: Brazil has a complex population structure, with evidence of P. simium infections and Amazonian parasites separating into multiple clusters. Overall, our work provides the first Brazil-wide analysis of P. vivax population structure and identifies important mutations, which can inform future research and control measures. Funding: AI is funded by an MRC LiD PhD studentship. TGC is funded by the Medical Research Council (Grant no. MR/M01360X/1, MR/N010469/1, MR/R025576/1, MR/R020973/1 and MR/X005895/1). SC is funded by Medical Research Council UK grants (MR/M01360X/1, MR/R025576/1, MR/R020973/1 and MR/X005895/1) and Bloomsbury SET (ref. CCF17-7779). FN is funded by The Shloklo Malaria Research Unit - part of the Mahidol Oxford Research Unit, supported by the Wellcome Trust (Grant no. 220211). ARSB is funded by São Paulo Research Foundation - FAPESP (Grant no. 2002/09546-1). RLDM is funded by Brazilian National Council for Scientific and Technological Development - CNPq (Grant no. 302353/2003-8 and 471605/2011-5); CRFM is funded by FAPESP (Grant no. 2020/06747-4) and CNPq (Grant no. 302917/2019-5 and 408636/2018-1); JGD is funded by FAPESP fellowships (2016/13465-0 and 2019/12068-5) and CNPq (Grant no. 409216/2018-6).

New Strategies for Novel Drugs: Antimicrobial Peptides Containing Ferrocene with Improved Antifungal and Antiplasmodial Biological Activity.

BACKGROUND: Fungal and parasitic diseases are global health problems, and the available treatments are becoming ineffective, mainly due to the emergence of resistant strains of pathogens. Furthermore, the drugs currently in use exhibit high toxicity and side effects. The scarcity of efficient treatments for fungal and parasitic diseases has motivated the search for new drug candidates, including antimicrobial peptides. The chemokine class RP1 peptide shows inhibitory activity against bacteria, viruses, cancer cells and parasites. In addition, the organometallic compound ferrocene showed antiparasitic activity. OBJECTIVE: Study aimed to assess the effect of conjugation of the RP1 peptide with ferrocene in terms of its structure, biological activity against fungi and parasites and toxicity. METHODS: Peptides and conjugates were synthesized using solid phase peptide synthesis (SPPS). The Fc-RP1 peptide showed antifungal and antimalarial activities with low toxicity in the U87 and HepG2 cell lines. RESULTS: The mechanism of action of these peptides, analyzed by flow cytometry in the fungus Cryptococcus neoformans, was through membrane permeabilization, with an emphasis on the Fc-RP1 peptide that presented the highest rate of PI-positive cell marking. CONCLUSION: In conclusion, ferrocene conjugated to antimicrobial peptide RP1 is an attractive biomolecule for drug discovery against fungal and parasitic diseases.

Improving the Molecular Diagnosis of Malaria: Droplet Digital PCR-Based Method Using Saliva as a DNA Source.

Malaria is an acute febrile disease caused by a protozoan of the genus Plasmodium. Light microscopy (LM) is the gold standard for the diagnosis of malaria. Despite this method being rapid and inexpensive, it has a low limit of detection, which hampers the identification of low parasitemia infections. By using multicopy targets and highly sensitive molecular techniques, it is possible to change this scenario. In this study, we evaluated the performance of droplet digital PCR (ddPCR) to detect Plasmodium DNA obtained from saliva samples (whole saliva and buccal swab) of 157 individuals exposed to malaria transmission from the Brazilian Amazon region. We used the highly sensitive ddPCR method with non-ribosomal multicopy targets for Plasmodium vivax (Pvr47) and Plasmodium falciparum (Pfr364). There was good concordance between the quantitative real-time PCR (qPCR) results from the saliva and blood, except for mixed-species infections. The sensitivity of qPCR was 93% for blood, 77% for saliva, and 47% for swabs. Parasite DNA was not detected in saliva samples in low-density infections compared with the detection in blood samples. ddPCR showed increased sensitivity for detecting Plasmodium in the blood and swabs (99% in blood, 73% in saliva, and 59% in swabs). Notably, ddPCR detected more mixed infections in the blood (15%), saliva (9%), and swabs (18%) than qPCR. Our data showed that the differences between ddPCR and qPCR were the result of a higher number of P. falciparum infections detected by ddPCR. Overall, there was a moderate correlation between parasite densities estimated by the different methods in the blood. Our findings highlight the possibility of using non-invasive sample collection methods for malaria diagnosis by targeting multicopy sequences combined with highly sensitive molecular methods.

Discovery and Preclinical Pharmacology of INE963, a Potent and Fast-Acting Blood-Stage Antimalarial with a High Barrier to Resistance and Potential for Single-Dose Cures in Uncomplicated Malaria.

A series of 5-aryl-2-amino-imidazothiadiazole (ITD) derivatives were identified by a phenotype-based high-throughput screening using a blood stage Plasmodium falciparum (Pf) growth inhibition assay. A lead optimization program focused on improving antiplasmodium potency, selectivity against human kinases, and absorption, distribution, metabolism, excretion, and toxicity properties and extended pharmacological profiles culminated in the identification of INE963 (1), which demonstrates potent cellular activity against Pf 3D7 (EC50 = 0.006 µM) and achieves "artemisinin-like" kill kinetics in vitro with a parasite clearance time of <24 h. A single dose of 30 mg/kg is fully curative in the Pf-humanized severe combined immunodeficient mouse model. INE963 (1) also exhibits a high barrier to resistance in drug selection studies and a long half-life (T1/2) across species. These properties suggest the significant potential for INE963 (1) to provide a curative therapy for uncomplicated malaria with short dosing regimens. For these reasons, INE963 (1) was progressed through GLP toxicology studies and is now undergoing Ph1 clinical trials.

Synthesis and antiplasmodial assessment of nitazoxanide and analogs as new antimalarial candidates.

During the last years, the progression to control malaria disease seems to be slowed and WHO (World Health Organization) reported a modeling analysis with the prediction of the increase in malaria morbidity and mortality in sub-Saharan Africa during the COVID-19 pandemic. A rapid way to the discovery of new drugs could be carried out by performing investigations to identify drugs based on repurposing of "old" drugs. The 5-nitrothiazole drug, Nitazoxanide was shown to be active against intestinal protozoa, human helminths, anaerobic bacteria, viruses, etc. In this work, Nitazoxanide and analogs were prepared using two methodologies and evaluated against P. falciparum 3D7. A bithiazole analog, showed attractive inhibitory activity with an EC50 value of 5.9 µM, low propensity to show toxic effect against HepG2 cells at 25 µM, and no cross-resistance with standard antimalarials.

New Isoquinoline Alkaloids from Paraphaeosphaeria sporulosa F03, a Fungal Endophyte Isolated from Paepalanthus planifolius.

As part of our continuing efforts to discover new bioactive compounds from endophytic fungal sources, we have investigated the extract of the Paraphaeosphaeria sporulosa F03 strain. The study led to the isolation of four new 3-methyl-isoquinoline alkaloids (1:  - 4: ) and four known polyketides (5:  - 8: ). The structures of compounds 1:  - 4: were elucidated by 1D and 2D NMR experiments and HRMS analysis. The absolute configuration of 4: was determined by comparison of its experimental electronic circular dichroism spectrum with calculated data. Compounds 1:  - 4: exhibited antifungal activity with minimal inhibitory concentration values ranging from 6.25 - 50 µg/mL against six Candida species but they did not present any cytotoxic activity against the human tumor cell lines A549 (lung), MCF-7 (breast), and HepG2 (hepatocellular). In addition, compound 4: exhibited antiplasmodial activity in the low micromolar range (IC50 = 4 µM).

In vitro assessment for cytotoxicity screening of new antimalarial candidates

Abstract In antimalarial research there are no standard procedures to determine the toxicity of a drug candidate. Among the alternatives available, in vitro cytotoxicity assays are the most widely used to predict toxic effects of future therapeutic products. They have the advantage over the in vivo assays, in that they offer the possibility to restrain the number of experimental variables. The objective of the present study was to compare in vitro cytotoxic methods by testing various compounds currently used to treat malaria against different cell lines. Neutral red (NR) uptake and methylthiazoletetrazolium (MTT) colorimetric in vitro assays were used to determine preliminary toxicity of commercially available antimalarial drugs against tumor and non-tumor cells lines. Toxicity through brine shrimp lethality bioassay and hemolytic activity were also evaluated. Significant differences were observed in the tests measured by NR uptake. The tumor cell lines TOV-21G and HepG2 and non-tumor WI-26VA4 cells showed relatively uniform toxicity results, with TOV-21G being the most sensitive cell tested, presenting the lowest concentration to cause death to 50% of viable cells (CC50) values. The results of this study support the use of TOV-21G, HepG2 and WI-26VA4 cells lines as the choice for cytotoxicity tests to evaluate potential bioactive compounds.

MEFAS, a hybrid of artesunate-mefloquine active against asexual stages of Plasmodium vivax in field isolates, inhibits malaria transmission.

Human malaria continues to be a public health problem and an important cause of morbidity and mortality in the world. Malaria control is achieved through both individual protection against mosquito bites and drug treatment, which is hampered by the spread of Plasmodium falciparum resistance to most antimalarials, including artemisinin derivatives. One of the key pharmacological strategies for controlling malaria is to block transmission of the parasites to their mosquito vectors. Following this rational, MEFAS, a synthetic hybrid salt derived from artesunate (AS) and mefloquine has been previously reported for its activity against asexual P. falciparum parasites in vitro, in addition to a pronounced reduction in the viability of mature gametocytes. Herein, MEFAS was tested against asexual forms of Plasmodium vivax and for its ability to block malaria transmission in Anopheles darlingi mosquitoes in a membrane feeding assay using P. vivax field isolates. MEFAS demonstrated high potency, with a IC50 of 6.5 nM against asexual forms of P. vivax. At 50 µM, MEFAS completely blocked oocyst formation in mosquitoes, regardless of the oocyst number in the control group. At lower doses, MEFAS reduced oocyst prevalence by greater than 20%. At equivalent doses, AS irregularly reduced oocyst formation and caused only slight inhibition of mosquito infections. These results highlight the potential of MEFAS as a novel transmission-blocking molecule, as well as its high blood schizonticidal activity against P. vivax and P. falciparum field isolates, representing a starting point for further development of a new drug with dual antimalarial activity.

A Comprehensive Analysis of the Genetic Diversity of Plasmodium falciparum Histidine-Rich Protein 2 (PfHRP2) in the Brazilian Amazon.

Early diagnosis and treatment are fundamental to the control and elimination of malaria. In many endemic areas, routine diagnosis is primarily performed microscopically, although rapid diagnostic tests (RDTs) provide a useful point-of-care tool. Most of the commercially available RDTs detect histidine-rich protein 2 (HRP2) of Plasmodium falciparum in the blood of infected individuals. Nonetheless, parasite isolates lacking the pfhrp2 gene are relatively frequent in some endemic regions, thereby hampering the diagnosis of malaria using HRP2-based RDTs. To track the efficacy of RDTs in areas of the Brazilian Amazon, we assessed pfhrp2 deletions in 132 P. falciparum samples collected from four malaria-endemic states in Brazil. Our findings show low to moderate levels of pfhrp2 deletion in different regions of the Brazilian Amazon. Overall, during the period covered by this study (2002-2020), we found that 10% of the P. falciparum isolates were characterized by a pfhrp2 deletion. Notably, however, the presence of pfhrp2-negative isolates has not been translated into a reduction in RDT efficacy, which in part may be explained by the presence of polyclonal infections. A further important finding was the discrepancy in the proportion of pfhrp2 deletions detected using two assessed protocols (conventional PCR versus nested PCR), which reinforces the need to perform a carefully planned laboratory workflow to assess gene deletion. This is the first study to perform a comprehensive analysis of PfHRP2 sequence diversity in Brazilian isolates of P. falciparum. We identified 10 PfHRP2 sequence patterns, which were found to be exclusive of each of the assessed regions. Despite the small number of PfHRP2 sequences available from South America, we found that the PfHRP2 sequences identified in Brazil and neighboring French Guiana show similar sequence patterns. Our findings highlight the importance of continuously monitoring the occurrence and spread of parasites with pfrhp2 deletions, while also taking into account the limitations of PCR-based testing methods associated with accuracy and the complexity of infections.

Design of proteasome inhibitors with oral efficacy in vivo against Plasmodium falciparum and selectivity over the human proteasome.

The Plasmodium falciparum proteasome is a potential antimalarial drug target. We have identified a series of amino-amide boronates that are potent and specific inhibitors of the P. falciparum 20S proteasome (Pf20S) ß5 active site and that exhibit fast-acting antimalarial activity. They selectively inhibit the growth of P. falciparum compared with a human cell line and exhibit high potency against field isolates of P. falciparum and Plasmodium vivax They have a low propensity for development of resistance and possess liver stage and transmission-blocking activity. Exemplar compounds, MPI-5 and MPI-13, show potent activity against P. falciparum infections in a SCID mouse model with an oral dosing regimen that is well tolerated. We show that MPI-5 binds more strongly to Pf20S than to human constitutive 20S (Hs20Sc). Comparison of the cryo-electron microscopy (EM) structures of Pf20S and Hs20Sc in complex with MPI-5 and Pf20S in complex with the clinically used anti-cancer agent, bortezomib, reveal differences in binding modes that help to explain the selectivity. Together, this work provides insights into the 20S proteasome in P. falciparum, underpinning the design of potent and selective antimalarial proteasome inhibitors.

Antiprotozoal and Antibacterial Activity of Ravenelin, a Xanthone Isolated from the Endophytic Fungus Exserohilum rostratum.

The natural compound ravenelin was isolated from the biomass extracts of Exserohilum rostratum fungus, and its antimicrobial, antiplasmodial, and trypanocidal activities were evaluated. Ravenelin was isolated by column chromatography and HPLC and identified by NMR and MS. The susceptibility of Gram-positive and Gram-negative bacteria strains to ravenelin was determined by microbroth dilution assay. Cytotoxicity was evaluated in hepatocarcinoma cells (HepG2) and BALB/c peritoneal macrophages by using MTT. SYBR Green I-based assay was used in the asexual stages of Plasmodium falciparum. Trypanocidal activity was tested against the epimastigote and intracellular amastigote forms of Trypanosoma cruzi. Ravenelin was active against Gram-positive bacteria strains, with emphasis on Bacillus subtilis (MIC value of 7.5 µM). Ravenelin's antiparasitic activities were assessed against both the epimastigote (IC50 value of 5 ± 1 µM) and the intracellular amastigote forms of T. cruzi (IC50 value of 9 ± 2 µM), as well as against P. falciparum (IC50 value of 3.4 ± 0.4 µM). Ravenelin showed low cytotoxic effects on both HepG2 (CC50 > 50 µM) and peritoneal macrophage (CC50 = 185 ± 1 µM) cells with attractive selectivity for the parasites (SI values > 15). These findings indicate that ravenelin is a natural compound with both antibacterial and antiparasitic activities, and considerable selectivity indexes. Therefore, ravenelin is an attractive candidate for hit-to-lead development.

Distinctive genetic structure and selection patterns in Plasmodium vivax from South Asia and East Africa.

Despite the high burden of Plasmodium vivax malaria in South Asian countries, the genetic diversity of circulating parasite populations is not well described. Determinants of antimalarial drug susceptibility for P. vivax in the region have not been characterised. Our genomic analysis of global P. vivax (n = 558) establishes South Asian isolates (n = 92) as a distinct subpopulation, which shares ancestry with some East African and South East Asian parasites. Signals of positive selection are linked to drug resistance-associated loci including pvkelch10, pvmrp1, pvdhfr and pvdhps, and two loci linked to P. vivax invasion of reticulocytes, pvrbp1a and pvrbp1b. Significant identity-by-descent was found in extended chromosome regions common to P. vivax from India and Ethiopia, including the pvdbp gene associated with Duffy blood group binding. Our investigation provides new understanding of global P. vivax population structure and genomic diversity, and genetic evidence of recent directional selection in this important human pathogen.

Potent Antimalarials with Development Potential Identified by Structure-Guided Computational Optimization of a Pyrrole-Based Dihydroorotate Dehydrogenase Inhibitor Series.

Dihydroorotate dehydrogenase (DHODH) has been clinically validated as a target for the development of new antimalarials. Experience with clinical candidate triazolopyrimidine DSM265 (1) suggested that DHODH inhibitors have great potential for use in prophylaxis, which represents an unmet need in the malaria drug discovery portfolio for endemic countries, particularly in areas of high transmission in Africa. We describe a structure-based computationally driven lead optimization program of a pyrrole-based series of DHODH inhibitors, leading to the discovery of two candidates for potential advancement to preclinical development. These compounds have improved physicochemical properties over prior series frontrunners and they show no time-dependent CYP inhibition, characteristic of earlier compounds. Frontrunners have potent antimalarial activity in vitro against blood and liver schizont stages and show good efficacy in Plasmodium falciparum SCID mouse models. They are equally active against P. falciparum and Plasmodium vivax field isolates and are selective for Plasmodium DHODHs versus mammalian enzymes.

Repositioning and Characterization of 1-(Pyridin-4-yl)pyrrolidin-2-one Derivatives as Plasmodium Cytoplasmic Prolyl-tRNA Synthetase Inhibitors.

Prolyl-tRNA synthetase (PRS) is a clinically validated antimalarial target. Screening of a set of PRS ATP-site binders, initially designed for human indications, led to identification of 1-(pyridin-4-yl)pyrrolidin-2-one derivatives representing a novel antimalarial scaffold. Evidence designates cytoplasmic PRS as the drug target. The frontrunner 1 and its active enantiomer 1-S exhibited low-double-digit nanomolar activity against resistant Plasmodium falciparum (Pf) laboratory strains and development of liver schizonts. No cross-resistance with strains resistant to other known antimalarials was noted. In addition, a similar level of growth inhibition was observed against clinical field isolates of Pf and P. vivax. The slow killing profile and the relative high propensity to develop resistance in vitro (minimum inoculum resistance of 8 × 105 parasites at a selection pressure of 3 × IC50) constitute unfavorable features for treatment of malaria. However, potent blood stage and antischizontal activity are compelling for causal prophylaxis which does not require fast onset of action. Achieving sufficient on-target selectivity appears to be particularly challenging and should be the primary focus during the next steps of optimization of this chemical series. Encouraging preliminary off-target profile and oral efficacy in a humanized murine model of Pf malaria allowed us to conclude that 1-(pyridin-4-yl)pyrrolidin-2-one derivatives represent a promising starting point for the identification of novel antimalarial prophylactic agents that selectively target Plasmodium PRS.

Metabolites from Marine Sponges and Their Potential to Treat Malarial Protozoan Parasites Infection: A Systematic Review.

Malaria is an infectious disease caused by protozoan parasites of the Plasmodium genus through the bite of female Anopheles mosquitoes, affecting 228 million people and causing 415 thousand deaths in 2018. Artemisinin-based combination therapies (ACTs) are the most recommended treatment for malaria; however, the emergence of multidrug resistance has unfortunately limited their effects and challenged the field. In this context, the ocean and its rich biodiversity have emerged as a very promising resource of bioactive compounds and secondary metabolites from different marine organisms. This systematic review of the literature focuses on the advances achieved in the search for new antimalarials from marine sponges, which are ancient organisms that developed defense mechanisms in a hostile environment. The principal inclusion criterion for analysis was articles with compounds with IC50 below 10 µM or 10 µg/mL against P. falciparum culture. The secondary metabolites identified include alkaloids, terpenoids, polyketides endoperoxides and glycosphingolipids. The structural features of active compounds selected in this review may be an interesting scaffold to inspire synthetic development of new antimalarials for selectively targeting parasite cell metabolism.

A Multistage Formulation Based on Full-Length CSP and AMA-1 Ectodomain of Plasmodium vivax Induces High Antibody Titers and T-cells and Partially Protects Mice Challenged with a Transgenic Plasmodium berghei Parasite.

Infections with Plasmodium vivax are predominant in the Americas, representing 75% of malaria cases. Previously perceived as benign, malaria vivax is, in fact, a highly debilitating and economically important disease. Considering the high complexity of the malaria parasite life cycle, it has been hypothesized that an effective vaccine formulation against Plasmodium should contain multiple antigens expressed in different parasite stages. Based on that, we analyzed a recombinant P. vivax vaccine formulation mixing the apical membrane antigen 1 ectodomain (PvAMA-1) and a full-length circumsporozoite protein (PvCSP-AllFL) previously studied by our group, which elicits a potent antibody response in mice. Genetically distinct strains of mice (C57BL/6 and BALB/c) were immunized with the proteins, alone or in combination, in the presence of poly(I:C) adjuvant, a TLR3 agonist. In C57BL/6, high-antibody titers were induced against PvAMA-1 and the three PvCSP variants (VK210, VK247, and P. vivax-like). Meanwhile, mixing PvAMA-1 with PvCSP-AllFL had no impact on total IgG antibody titers, which were long-lasting. Moreover, antibodies from immunized mice recognized VK210 sporozoites and blood-stage parasites by immunofluorescence assay. However, in the BALB/c model, the antibody response against PvCSP-AllFL was relatively low. PvAMA-1-specific CD3+CD4+ and CD3+CD8+ T-cell responses were observed in C57BL/6 mice, and the cellular response was impaired by PvCSP-AllFL combination. More relevant, the multistage vaccine formulation provided partial protection in mice challenged with a transgenic Plasmodium berghei sporozoite expressing the homologous PvCSP protein.