Antimalarial Activity of Potential Inhibitors of Plasmodium falciparum Lactate Dehydrogenase Enzyme Selected by Docking Studies

The Plasmodium falciparum lactate dehydrogenase enzyme (PfLDH) has been considered as a potential molecular target for antimalarials due to this parasite's dependence on glycolysis for energy production. Because the LDH enzymes found in P. vivax, P. malariae and P. ovale (pLDH) all exhibit ∼90% identity to PfLDH, it would be desirable to have new anti-pLDH drugs, particularly ones that are effective against P. falciparum, the most virulent species of human malaria. Our present work used docking studies to select potential inhibitors of pLDH, which were then tested for antimalarial activity against P. falciparum in vitro and P. berghei malaria in mice. A virtual screening in DrugBank for analogs of NADH (an essential cofactor to pLDH) and computational studies were undertaken, and the potential binding of the selected compounds to the PfLDH active site was analyzed using Molegro Virtual Docker software. Fifty compounds were selected based on their similarity to NADH. The compounds with the best binding energies (itraconazole, atorvastatin and posaconazole) were tested against P. falciparum chloroquine-resistant blood parasites. All three compounds proved to be active in two immunoenzymatic assays performed in parallel using monoclonals specific to PfLDH or a histidine rich protein (HRP2). The IC50 values for each drug in both tests were similar, were lowest for posaconazole (<5 µM) and were 40- and 100-fold less active than chloroquine. The compounds reduced P. berghei parasitemia in treated mice, in comparison to untreated controls; itraconazole was the least active compound. The results of these activity trials confirmed that molecular docking studies are an important strategy for discovering new antimalarial drugs. This approach is more practical and less expensive than discovering novel compounds that require studies on human toxicology, since these compounds are already commercially available and thus approved for human use

In vivo antimalarial activity and mechanisms of action of 4-nerolidylcatechol derivatives

4-Nerolidylcatechol (1) is an abundant antiplasmodial metabolite that is isolated from Piper peltatum roots. O-Acylation or O-alkylation of compound 1 provides derivatives exhibiting improved stability and significant in vitro antiplasmodial activity. The aim of this work was to study the in vitro inhibition of hemozoin formation, inhibition of isoprenoid biosynthesis in Plasmodium falciparum cultures, and in vivo antimalarial activity of several 4-nerolidylcatechol derivatives. 1,2-O,O-Diacetyl-4-nerolidylcatechol (2) inhibited in vitro hemozoin formation by up to 50%. In metabolic labeling studies using [1-(n)-3H]geranylgeranyl pyrophosphate, diester 2 significantly inhibited the biosynthesis of isoprenoid metabolites ubiquinone 8, menaquinone 4, and dolichol 12 in cultures of P. falciparum 3D7. Similarly, 2-O-benzyl-4-nerolidylcatechol (3) significantly inhibited the biosynthesis of dolichol 12. P. falciparum in vitro protein synthesis was not affected by compounds 2 or 3. At oral doses of 50 mg per kg of body weight per day, compound 2 suppressed Plasmodium berghei NK65 in infected BALB/c mice by 44%. This in vivo result for derivative 2 represents marked improvement over that obtained previously for natural product 1. Compound 2 was not detected in mouse blood 1 h after oral ingestion or in mixtures with mouse blood/blood plasma in vitro. However, it was detected after in vitro contact with human blood or blood plasma. Derivatives of 4-nerolidylcatechol exhibit parasite-specific modes of action, such as inhibition of isoprenoid biosynthesis and inhibition of hemozoin formation, and they therefore merit further investigation for their antimalarial potential. Copyright © 2015, American Society for Microbiology. All Rights Reserved

Busca de novos fármacos para o tratamento da malária humana através de diferentes abordagens

Exportado OPUSMade available in DSpace on 2019-08-11T15:02:33Z (GMT). No. of bitstreams: 1 tese_julia_penna_coutinho.pdf: 13940140 bytes, checksum: 0702718c7d07da31d77951b82707b2b8 (MD5) Previous issue date: 27A malária humana permanece como uma das principais doenças parasitárias do mundo, afetando milhões de pessoas ao ano e causando intensa morbidade e mortalidade, sobretudo em crianças da África. A doença causa ainda pobreza em países com alta prevalência, incluindo o Brasil, na Região Amazônica. Devido à resistência disseminada do Plasmodium falciparum aos antimaláricos disponíveis e do P. vivax à cloroquina, inclusive no Brasil, novas terapias são necessárias, já que o tratamento medicamentoso permanece como a principal estratégia no controle da doença. A busca de novos fármacos tem como alvo principal os estágios eritrocíticos do Plasmodium, mas o interesse em reduzir a transmissão e erradicar a malária inclui combater as formas sexuadas, tornando necessário o desenvolvimento de fármacos contra os gametócitos. No presente trabalho a proposta principal foi a busca por novos compostos com ação dupla, sendo avaliada a atividade esquizonticida sanguínea, in vitro e in vivo, e bloqueadora da transmissão, in vitro. Os compostos foram selecionados por diferentes abordagens: (i) estudos computacionais de modelagem molecular, tendo como alvos específicos as enzimas lactato desidrogenase (atorvastatina, itraconazol e posaconazol) e topoisomerase I (pterocarpanquinona e derivados) do P. falciparum; (ii) o conhecimento popular do uso de infusões da planta medicinal Aspidosperma nitidum contra a malária na Amazônia Brasileira, e, (iii) o desenvolvimento de compostos híbridos, derivados da ligação do artesunato com mefloquina (MEFAS) ou com primaquina (PRIMAS). Os resultados demonstraram que: (i) atorvastatina, itraconazol e posaconazol, comercialmente disponíveis, e extratos e frações da casca do caule de A. nitidum, foram ativos in vitro contra P. falciparum; (ii) atorvastatina, posaconazol e compostos isolados de A. nitidum, inibiram ativamente a parasitemia de camundongos experimentalmente infectados com P. berghei, e, (iii) MEFAS e PRIMAS apresentaram atividade dupla, sendo ativos contra as formas assexuadas e contra os gametócitos do P. falciparum. Conclui-se que o conhecimento científico gerado neste trabalho poderá contribuir para a formulação de novos fármacos com ação antimalárica dupla e seletiva.The human malaria remains one of the major parasitic diseases in the world, affecting millions of people each year and causing severe morbidity and mortality, especially in children in Africa. The disease causes poverty in countries with high prevalence, including Brazil, in the Amazon Region. Due to the spread of Plasmodium falciparum resistance to available antimalarial drugs and of P. vivax to chloroquine, including in Brazil, new therapies are needed, as drug treatment remains the main strategy to control the disease. The search for new drugs has the erythrocytic stages of Plasmodium as the main target, but the interest in reducing malaria transmission and eradicating the disease includes eliminating sexual forms, which requires the development of drugs against gametocytes. In this study the main proposal was to search for new compounds with dual activity of blood schizonticidal activity evaluated in vitro and in vivo, as well as transmission blocking activity, in vitro. The compound selection for screening was performed by different approaches: (i) docking studies having the enzymes as specific targets lactate dehydrogenase (atorvastatin, itraconazole, posaconazole) and topoisomerase I (pterocarpanquinones and derivatives) of P. falciparum; (ii) the popular knowledge about the use of infusions of the medicinal plant Aspidosperma nitidum against malaria in the Brazilian Amazon, and, (iii) the development of hybrid compounds, derived from artesunate with mefloquine (MEFAS) or primaquine (PRIMAS). The results showed that: (i) the commercially available drugs atorvastatin, itraconazole, posaconazole, and the extracts and fractions of A. nitidum stem bark were active in vitro against P. falciparum; (ii) atorvastatin, posaconazole and the compounds isolated from A. nitidum, also inhibited the parasitemia of mice experimentally infected with P. berghei, and, (iii) MEFAS and PRIMAS showed a dual activity against P. falciparum asexual forms and gametocytes. It can be concluded that scientific knowledge generated in this study may contribute to the development of new drugs with dual and selective antimalarial activity

Commercial drugs containing flavonoids are active in mice with malaria and in vitro against chloroquine-resistant Plasmodium falciparum

BACKGROUND The main strategy to control human malaria still relies on specific drug treatment, limited now by Plasmodium falciparum-resistant parasites, including that against artemisinin derivatives. Despite the large number of active compounds described in the literature, few of them reached full development against human malaria. Drug repositioning is a fast and less expensive strategy for antimalarial drug discovery, because these compounds are already approved for human use. OBJECTIVES To identify new antimalarial drugs from compounds commercially available and used for other indications. METHODS Accuvit®, Ginkgo® and Soyfit®, rich in flavonoids, and also the standard flavonoids, hesperidin, quercetin, and genistein were tested against blood cultures of chloroquine-resistant P. falciparum, as well as chloroquine, a reference antimalarial. Inhibition of parasite growth was measured in immunoenzymatic assay with monoclonal anti-P. falciparum antibodies, specific to the histidine-rich protein II. Tests in mice with P. berghei malaria were based on percent of parasitaemia reduction. These compounds were also evaluated for in vitro cytotoxicity. FINDINGS The inhibition of parasite growth in vitro showed that Accuvit® was the most active drug (IC50 5 ± 3.9 μg/mL). Soyfit® was partially active (IC50 13.6 ± 7.7 μg/mL), and Ginkgo® (IC50 38.4 ± 14 μg/mL) was inactive. All such compounds were active in vivo at a dose of 50 mg/kg body weight. Accuvit® and quercetin induced the highest reduction of P. berghei parasitaemia (63% and 53%, respectively) on day 5 after parasite inoculation. As expected, the compounds tested were not toxic. MAIN CONCLUSIONS The antimalarial activity of Accuvit® was not related to flavonoids only, and it possibly results from synergisms with other compounds present in this drug product, such as multivitamins. Multivitamins in Accuvit® may explain its effect against the malaria parasites. This work demonstrated for the first time the activity of these drugs, which are already marketed

Theoretical and experimental studies of new modified isoflavonoids as potential inhibitors of topoisomerase I from Plasmodium falciparum.

DNA topoisomerase I from Plasmodium falciparum (PfTopoI), a potential selective target for chemotherapy and drug development against malaria, is used here, together with human Topo I (HssTopoI), for docking, molecular dynamics (MD) studies and experimental assays. Six synthetic isoflavonoid derivatives and the known PfTopoI inhibitors camptothecin and topotecan were evaluated in parallel. Theoretical results suggest that these compounds dock in the binding site of camptothecin and topotecan inside both enzymes and that LQB223 binds selectively in PfTopoI. In vitro tests against P. falciparum blood parasites corroborated the theoretical findings. The selectivity index (SI) of LQB223 ≥ 98 suggests that this molecule is the most promising in the group of compounds tested. In vivo experiments in mice infected with P. berghei showed that LQB223 has an antimalarial activity similar to that of chloroquine

Aspidosperma (Apocynaceae) plant cytotoxicity and activity towards malaria parasites. Part I: Aspidosperma nitidum (Benth) used as a remedy to treat fever and malaria in the Amazon

Infusions of Aspidosperma nitidum (Apocynaceae) wood bark are used to treat fever and malaria in the Amazon Region. Several species of this family are known to possess indole alkaloids and other classes of secondary metabolites, whereas terpenoids, an inositol and the indole alkaloids harmane-3 acid and braznitidumine have been described in A. nitidum . In the present study, extracts from the wood bark, leaves and branches of this species were prepared for assays against malaria parasites and cytotoxicity testing using human hepatoma and normal monkey kidney cells. The wood bark extracts were active against Plasmodium falciparum and showed a low cytotoxicity in vitro, whereas the leaf and branch extracts and the pure alkaloid braznitidumine were inactive. A crude methanol extract was subjected to acid-base fractionation aimed at obtaining alkaloid-rich fractions, which were active at low concentrations against P. falciparum and in mice infected with and sensitive Plasmodium berghei parasites. Our data validate the antimalarial usefulness of A. nitidum wood bark, a remedy that can most likely help to control malaria. However, the molecules responsible for this antimalarial activity have not yet been identified. Considering their high selectivity index, the alkaloid-rich fractions from the plant bark might be useful in the development of new antimalarials

Superposition of topotecan (yellow) to its crystallographic structure (green) after re-docking calculations.

<p>Superposition of topotecan (yellow) to its crystallographic structure (green) after re-docking calculations.</p