In vitro and in vivo antileishmanial efficacy of a combination therapy of diminazene and artesunate against Leishmania donovani in BALB/c mice

The in vitro and in vivo activity of diminazene (Dim), artesunate (Art) and combination of Dim and Art (Dim-Art) against Leishmania donovani was compared to reference drug; amphotericin B. IC50 of Dim-Art was found to be 2.28 ± 0.24 µg/mL while those of Dim and Art were 9.16 ± 0.3 µg/mL and 4.64 ± 0.48 µg/mL respectively. The IC50 for Amphot B was 0.16 ± 0.32 µg/mL against stationary-phase promastigotes. In vivo evaluation in the L. donovani BALB/c mice model indicated that treatments with the combined drug therapy at doses of 12.5 mg/kg for 28 consecutive days significantly (p < 0.001) reduced parasite burden in the spleen as compared to the single drug treatments given at the same dosages. Although parasite burden was slightly lower (p < 0.05) in the Amphot B group than in the Dim-Art treatment group, the present study demonstrates the positive advantage and the potential use of the combined therapy of Dim-Art over the constituent drugs, Dim or Art when used alone. Further evaluation is recommended to determine the most efficacious combination ratio of the two compounds.

Tratamento da malária com artesunate (retocaps®) em crianças da Amazônia brasileira

Avaliamos a resposta clínica e parasitológica à terapêutica com o artesunate retocaps®, em 32 crianças internadas na Fundação de Medicina Tropical do Amazonas, que apresentavam malária com quadro clínico moderado e grave. Destas, 29 tinham a doença por P. falciparum e três, P. vivax. A melhora clínica foi observada após 24 horas do início da terapêutica, com 33,3% de pacientes afebris e, 48 horas após o tratamento, 77,2% das crianças não apresentavam febre. O acompanhamento da parasitemia assexuada, mostrou que no D2 58,6% das crianças com malária falciparum estavam negativas; em D4 todas haviam negativado, tanto na malária pelo P. falciparum como pelo P. vivax. No seguimento prolongado, na malária P. falciparum, encontramos 66,6% de recrudescências. Os resultados nos permitem concluir pela eficácia e praticidade no uso do artesunate retocaps® com rápida redução da parasitemia e melhora clínica. Entretanto, na malária P. falciparum a taxa de recrudescência foi elevada. Não foi observado para-efeito que possa ser imputado ao uso da droga.

In vitro action of antiparasitic drugs, especially artesunate, against Toxoplasma gondii

INTRODUCTION: Toxoplasmosis is usually a benign infection, except in the event of ocular, central nervous system (CNS), or congenital disease and particularly when the patient is immunocompromised. Treatment consists of drugs that frequently cause adverse effects; thus, newer, more effective drugs are needed. In this study, the possible activity of artesunate, a drug successfully being used for the treatment of malaria, on Toxoplasma gondii growth in cell culture is evaluated and compared with the action of drugs that are already being used against this parasite. METHODS: LLC-MK2 cells were cultivated in RPMI medium, kept in disposable plastic bottles, and incubated at 36ºC with 5% CO2. Tachyzoites of the RH strain were used. The following drugs were tested: artesunate, cotrimoxazole, pentamidine, pyrimethamine, quinine, and trimethoprim. The effects of these drugs on tachyzoites and LLC-MK2 cells were analyzed using nonlinear regression analysis with Prism 3.0 software. RESULTS: Artesunate showed a mean tachyzoite inhibitory concentration (IC50) of 0.075µM and an LLC MK2 toxicity of 2.003µM. Pyrimethamine was effective at an IC50 of 0.482µM and a toxicity of 11.178µM. Trimethoprim alone was effective against the in vitro parasite. Cotrimoxazole also was effective against the parasite but at higher concentrations than those observed for artesunate and pyrimethamine. Pentamidine and quinine had no inhibitory effect over tachyzoites. CONCLUSIONS: Artesunate is proven in vitro to be a useful alternative for the treatment of toxoplasmosis, implying a subsequent in vivo effect and suggesting the mechanism of this drug against the parasite.

Artesunate + amodiaquine versus artemether-lumefantrine for the treatment of uncomplicated Plasmodium falciparum malaria in the Colombian Pacific region: a noninferiority trial

INTRODUCTION: In Colombia, there are no published studies for the treatment of uncomplicated Plasmodium falciparum malaria comparing artemisinin combination therapies. Hence, it is intended to demonstrate the non-inferior efficacy/safety profiles of artesunate + amodiaquine versus artemether-lumefantrine treatments. METHODS: A randomized, controlled, open-label, noninferiority (Δ≤5%) clinical trial was performed in adults with uncomplicated P. falciparum malaria using the 28‑day World Health Organization validated design/definitions. Patients were randomized 1:1 to either oral artesunate + amodiaquine or artemether-lumefantrine. The primary efficacy endpoint: adequate clinical and parasitological response; secondary endpoints: - treatment failures defined per the World Health Organization. Safety: assessed through adverse events. RESULTS: A total of 105 patients was included in each group: zero censored observations. Mean (95%CI - Confidence interval) adequate clinical and parasitological response rates: 100% for artesunate + amodiaquine and 99% for artemether-lumefantrine; the noninferiority criteria was met (Δ=1.7%). There was one late parasitological therapeutic failure (1%; artemether-lumefantrine group), typified by polymerase chain reaction as the MAD20 MSP1 allele. The fever clearance time (artesunate + amodiaquine group) was significantly shorter (p=0.002). Respectively, abdominal pain for artesunate + amodiaquine and artemether-lumefantrine was 1.9% and 3.8% at baseline (p=0.68) and 1% and 13.3% after treatment (p<0.001). CONCLUSIONS: Uncomplicated P. falciparum malaria treatment with artesunate + amodiaquine is noninferior to the artemether-lumefantrine standard treatment. The efficacy/safety profiles grant further studies in this and similar populations.

Pharmacokinetic parameters of artesunate and dihydroartemisinin in rats infected with Fasciola hepatica

OBJECTIVES: The pharmacokinetic (PK) parameters of artesunate, recently discovered to possess promising trematocidal activity, and its main metabolite dihydroartemisinin (DHA) were determined in rats infected with hepatic and biliary stages of Fasciola hepatica and compared with uninfected rats after single intragastric and intravenous (iv) doses. METHODS: Rats infected with F. hepatica for 25 and 83 days and uninfected rats were cannulated in the right jugular vein and blood samples were withdrawn at selected timepoints following 10 mg/kg of iv and a single 100 mg/kg oral dose of artesunate. Plasma was analysed for artesunate and DHA by liquid chromatography coupled to tandem mass spectrometry. RESULTS: Rats harbouring juvenile and adult F. hepatica infections revealed considerable changes in PK parameters of artesunate and DHA. Following oral administration, maximum plasma concentrations (C(max)) of artesunate and DHA were 1.8-2.3-fold higher in infected rats [artesunate: 1334 +/- 1404 ng/mL (no infection) versus 2454 +/- 1494 ng/mL (acute infection) and 2768 +/- 538 ng/mL (chronic infection); DHA: 3802 +/- 2149 ng/mL (no infection) versus 6507 +/- 3283 ng/mL (acute infection) and 9093 +/- 884 ng/mL (chronic infection)]. The AUCs of artesunate and DHA were 2.1-4.4-fold greater in infected rats. An opposite trend was observed after iv injection. C(max) and AUC of artesunate and DHA following iv dosing were 5784 +/- 3718 and 140 938 +/- 128 783 ng.min/mL and 3849 +/- 3060 and 86 107 +/- 41 863 ng.min/mL, respectively, in uninfected rats versus 2623 +/- 1554 and 21 617 +/- 12 230 ng.min/mL and 2835 +/- 980 and 64 290 +/- 29 057 ng.min/mL, respectively, in rats harbouring a chronic infection. The elimination half-lives (t(1/2)) of artesunate and DHA were considerably altered in infected rats following oral and iv administration of artesunate. CONCLUSIONS: F. hepatica infections strongly influence the disposition kinetics of artesunate and its metabolite in rats. The clinical implications of this finding need to be carefully studied.

Argemone mexicana decoction versus artesunate-amodiaquine for the management of malaria in Mali: policy and public-health implications.

A classic way of delaying drug resistance is to use an alternative when possible. We tested the malaria treatment Argemone mexicana decoction (AM), a validated self-prepared traditional medicine made with one widely available plant and safe across wide dose variations. In an attempt to reflect the real situation in the home-based management of malaria in a remote Malian village, 301 patients with presumed uncomplicated malaria (median age 5 years) were randomly assigned to receive AM or artesunate-amodiaquine [artemisinin combination therapy (ACT)] as first-line treatment. Both treatments were well tolerated. Over 28 days, second-line treatment was not required for 89% (95% CI 84.1-93.2) of patients on AM, versus 95% (95% CI 88.8-98.3) on ACT. Deterioration to severe malaria was 1.9% in both groups in children aged </=5 years (there were no cases in patients aged >5 years) and 0% had coma/convulsions. AM, now government-approved in Mali, could be tested as a first-line complement to standard modern drugs in high-transmission areas, in order to reduce the drug pressure for development of resistance to ACT, in the management of malaria. In view of the low rate of severe malaria and good tolerability, AM may also constitute a first-aid treatment when access to other antimalarials is delayed.

Genotoxicity assessment of the antimalarial compound artesunate in somatic cells of mice

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Genotoxic evaluation of the antimalarial drugs artemisinin and artesunate in human HepG2 cells and effects on CASP3 and SOD1 gene expressions

The malaria treatment recommended by the World Health Organization involves medicines derived from artemisinin, an active compound extracted from the plant Artemisia annua, and some of its derivatives, such as artesunate. Considering the lack of data regarding the genotoxic effects of these compounds in human cells, the objective of this study was to evaluate the cytotoxicity and genotoxicity, and expressions of the CASP3 and SOD1 genes in a cultured human hepatocellular liver carcinoma cell line (HepG2 cells) treated with artemisinin and artesunate. We tested concentrations of 2.5, 5, 7.5, 10, and 20 μg/mL of both substances with a resazurin cytotoxicity assay, and the concentrations used in the genotoxicity experiments (2.5, 5, and 10 μg/mL) and gene expression analysis (5 mg/mL) were determined. The results of the comet assay in cells treated with artemisinin and artesunate showed a significant dosedependent increase (P < 0.001) in the number of cells with DNA damage at all concentrations tested. However, the gene expression analysis revealed no significant change in expression of CASP3 or SOD1. Our data showed that although artemisinin and artesunate exhibited genotoxic effects in cultured HepG2 cells, they did not significantly alter expression of the CASP3 and SOD1 genes at the doses tested. ©FUNPEC-RP.

In vitro action of antiparasitic drugs, especially artesunate, against Toxoplasma gondii

Introduction: Toxoplasmosis is usually a benign infection, except in the event of ocular, central nervous system (CNS), or congenital disease and particularly when the patient is immunocompromised. Treatment consists of drugs that frequently cause adverse effects; thus, newer, more effective drugs are needed. In this study, the possible activity of artesunate, a drug successfully being used for the treatment of malaria, on Toxoplasma gondii growth in cell culture is evaluated and compared with the action of drugs that are already being used against this parasite. Methods: LLC-MK2 cells were cultivated in RPMI medium, kept in disposable plastic bottles, and incubated at 36 degrees C with 5% CO2. Tachyzoites of the RH strain were used. The following drugs were tested: artesunate, cotrimoxazole, pentamidine, pyrimethamine, quinine, and trimethoprim. The effects of these drugs on tachyzoites and LLC-MK2 cells were analyzed using nonlinear regression analysis with Prism 3.0 software. Results: Artesunate showed a mean tachyzoite inhibitory concentration (IC50) of 0.075 mu M and an LLC MK2 toxicity of 2.003 mu M. Pyrimethamine was effective at an IC50 of 0.482 mu M and a toxicity of 11.178 mu M. Trimethoprim alone was effective against the in vitro parasite. Cotrimoxazole also was effective against the parasite but at higher concentrations than those observed for artesunate and pyrimethamine. Pentamidine and quinine had no inhibitory effect over tachyzoites. Conclusions: Artesunate is proven in vitro to be a useful alternative for the treatment of toxoplasmosis, implying a subsequent in vivo effect and suggesting the mechanism of this drug against the parasite.

Effect of artesunate-mefloquine fixed-dose combination in malaria transmission in amazon basin communities

Background: Studies in South-East Asia have suggested that early diagnosis and treatment with artesunate (AS) and mefloquine (MQ) combination therapy may reduce the transmission of Plasmodium falciparum malaria and the progression of MQ resistance. Methods: The effectiveness of a fixed-dose combination of AS and MQ (ASMQ) in reducing malaria transmission was tested in isolated communities of the Jurua valley in the Amazon region. Priority municipalities within the Brazilian Legal Amazon area were selected according to pre-specified criteria. Routine national malaria control programmatic procedures were followed. Existing health structures were reinforced and health care workers were trained to treat with ASMQ all confirmed falciparum malaria cases that match inclusion criteria. A local pharmacovigilance structure was implemented. Incidence of malaria and hospitalizations were recorded two years before, during, and after the fixed-dose ASMQ intervention. In total, between July 2006 and December 2008, 23,845 patients received ASMQ. Two statistical modelling approaches were applied to monthly time series of P. falciparum malaria incidence rates, P. falciparum/Plasmodium vivax infection ratio, and malaria hospital admissions rates. All the time series ranged from January 2004 to December 2008, whilst the intervention period span from July 2006 to December 2008. Results: The ASMQ intervention had a highly significant impact on the mean level of each time series, adjusted for trend and season, of 0.34 (95% CI 0.20 - 0.58) for the P. falciparum malaria incidence rates, 0.67 (95% CI 0.50 - 0.89) for the P. falciparum/P. vivax infection ratio, and 0.53 (95% CI 0.41 - 0.69) for the hospital admission rates. There was also a significant change in the seasonal (or monthly) pattern of the time series before and after intervention, with the elimination of the malaria seasonal peak in the rainy months of the years following the introduction of ASMQ. No serious adverse events relating to the use of fixed-dose ASMQ were reported. Conclusions: In the remote region of the Jurua valley, the early detection of malaria by health care workers and treatment with fixed-dose ASMQ was feasible and efficacious, and significantly reduced the incidence and morbidity of P. falciparum malaria.

Comparison of bioassay and high performance liquid chromatographic assay of artesunate and dihydroartemisinin in plasma

The study was a comparison of bioassay and HPLC analysis of artesunate (ARTS) and dihydroartemisinin (DHA) in plasma. ARTS and DHA in plasma samples from patients treated with ARTS were quantified by HPLC and expressed as DHA. DHA-equivalents in the same plasma samples were measured using a standardised parasite culture technique. DHA concentrations estimated by both methods were highly correlated (bioassay = 0.96 x HPLC + 11.0; r(2) = 0.92). At high concentrations ( > 12 000 nmol/l) bioassay sometimes overestimated DHA. Bioassay of active drug in plasma correlates well with specific chemical analysis by HPLC. ARTS and DHA appear to account for the total antimalarial activity in plasma after ARTS administration. (C) 2003 Elsevier Science B.V. All rights reserved.

Plasmodium vivax resistance to chloroquine (R2) and mefloquine (R3) in Brazilian Amazon region

We report for the first time a patient with malaria due to Plasmodium vivax who showed R2 resistance to chloroquine and R3 resistance to mefloquine in the Brazilian Amazon region based on WHO clinical criteria for diagnosis of malaria resistance. Failure was observed with unsupervised oral chloroquine, chloroquine under rigorous supervision and mefloquine in the same scheme. Finally, the patient was cured with oral artesunate.

Tratamento de crianças com malária pelo Plasmodium falciparum com derivados da artemisinina

No período compreendido entre janeiro de 1996 e dezembro de 1998, administramos derivados da artemisinina em 108 crianças com malária por Plasmodium falciparum, para avaliar a resposta clínica e terapêutica. Foram incluídas apenas crianças com clínica de malária moderada ou grave. No Grupo I, incluímos 62 pacientes e administramos artesunate por via endovenosa. Clinicamente, 50,8% tinham malária moderada e 49,2% malária grave; a parasitemia foi baixa em 53,2%, média em 22,6% e alta em 24,2%; no D2 a parasitemia estava negativa em 58,1%. No Grupo II,incluímos 46 pacientes que receberam artemeter (Paluter®) intramuscular. Clinicamente, 67,4% apresentavam malária moderada e 32,6% malária grave; a parasitemia foi baixa em 52,2%, média em 36,2% e alta em 15,2%; em D2, 56,5% apresentaram negativação da parasitemia. Nos dois grupos, a melhora clínica e evolução da parasitemia não mostraram diferença estatística; no D7 havia clareada a parasitemia em todos os pacientes. Para evitar recrudescência usamos mefloquina ou clindamicina.