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Avaliação in vitro e in vivo da atividade antimalárica dos trioxolanos NAC89, LDC67 e LC50 em estirpes sensíveis e resistentes à artemisinina e ao artesunato.
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A malária mantem-se como uma das doenças mais importantes do mundo, causando a morte de mais de 1 milhão de pessoas anualmente e elevada morbilidade. Face à propagação da resistência do Plasmodium falciparum à maioria dos medicamentos antimaláricos disponíveis, a Organização Mundial de Saúde (OMS), desde 2006, recomenda a utilização de terapias combinadas com artemisinina (ACTs) como tratamento de primeira linha para a malária não complicada. Em 2008, relatórios clínicos revelaram a falha terapêutica dos ACTs na fronteira Tailândia-Camboja e uma vez que não existem alternativas para o tratamento da malária é fundamental manter linhas de investigação sobre novos e eficazes fármacos. A partir da artemisinina (ART) surgiram novos peróxidos designados trioxolanos que apresentam como farmacóforo a função 1,2,4-trioxano. A acessibilidade, a preparação relativamente económica e a estabilidade da função 1,2,4-trioxano permite a síntese de derivados com estruturas diversas, alargando a possibilidade de desenvolvimento de novos fármacos. Foram realizados testes in vitro de triagem com o MARK III (OMS micro-ensaio), com controlos positivos (artemisinina e dihidroartemisinina) e controlo negativo (sem fármaco). Foram efetuados ensaios diversos com 3 compostos, aqui denominados NAC89, LCD67 e LC50 em culturas da estirpe de P. falciparum (Dd2) para avaliação da atividade antimalárica dos compostos, bem como ensaios utilizando o modelo de malária de murino, Plasmodium chabaudi, com 4 estirpes, denominadas AS-3CQ, AS-ATN, respectivamente sensível e resistente ao artesunato (ATN) e AS-30CQ e AS-ART respectivamente sensível e resistente à (ART). Também foi avaliada a citotoxicidade dos compostos, utilizando células HepG2 de hepatoma humano pelo ensaio com método colorimétrico metil-tiazol-tetrazólico (MTT). No modelo murino compararam-se também duas vias de administração dos novos compostos, sendo uma por via subcutânea nos 3 compostos e outra por via tópica apenas para LC50. A verificação de cura foi efetuada por observação microscópica de esfregaços sanguíneos corados pelo método de Giemsa e determinação da parasitemia. Os resultados observados foram: a) baixa citotoxicidade dos três compostos; b) o composto LC50 eliminou a parasitémia nos ensaios in vitro em cultivos de P. falciparum bem como eliminou P. chabaudi nos tratamentos por via subcutânea e tópica na dose de 50 mg/kg e na dose de 10 mg/Kg na via subcutânea; c) o NAC89 mostrou boa atividade no mesmo ensaio in vivo, na dose de 10 mg/Kg e 50 mg/Kg por via subcutânea; d) fraca atividade para LCD67 na dose de 50 mg/Kg. O LC50 e o NAC89 foram muito eficazes contra parasitas resistentes ao ATN e à ART sugerindo novos mecanismos de ação. Assim, este trabalho de investigação trouxe resultados promissores na àrea de potenciais novos antimaláricos.
Malaria remains one of the most important diseases in the world, causing the deaths of 1 million people annually and high morbidity. Given the spread of Plasmodium falciparum resistance to most antimalarial drugs available, the World Health Organization (WHO) recommended the use of artemisinin based combination therapies (ACTs) as first-line treatment since 2006 for uncomplicated malaria. In 2008, clinical reports have revealed the therapeutic failure of ACTs in Thailand-Cambodia border and since there is no alternative for the treatment of malaria it is essential to keep lines of research into new and effective drugs. From the discovery of artemisinin (ART) new peroxides emerged designated trioxolanes presenting the function 1,2,4-trioxane as pharmacophore. The accessibility, relatively inexpensive preparation and stability of the function 1,2,4-trioxane allow the synthesis of derivatives with different structures, extending the possibility of developing new drugs. Tests were performed in vitro screening with the MARK III (WHO micro-test) with the positive controls (artemisinin and dihidroartemisinin) and negative control (no drug). Assays were performed with three different compounds, here called NAC89, LCD67 and LC50 in cultures of P. falciparum (strain Dd2) as well as assays using the murine model of malaria, Plasmodium chabaudi, with 4 strains, denoted AS-3CQ, AS-ATN, respectively sensitive and resistant to artesunato (ATN) and AS-30CQ and AS-ART respectively sensitive and resistant to artemisinin. We also evaluated the cytotoxicity of the compounds using HepG2 human hepatoma cells, by using the colorimetric method metil-tiazol-tetrazolic (MTT) assay. In the murine model a comparison of two routes of administration of the new compounds was carried out for the 3 compounds NAC89, LCD67 and LC50 - subcutaneous inoculation while for LC50 we also assayed a topic administration. Cure was assessed by determining parasitaemia by microscopic observation of Giema stained blood smears. Results were: a) low cytotoxicity of the three compounds, b) compound LC50 eliminated parasites in in vitro assays in cultured P. falciparum and in the P. chabaudi model subcutaneously and topically at a dose of 50 mg/Kg and a 10 mg/Kg in subcutaneously, c) the NAC89 showed good activity in vivo in the same assay at a dose of 10 mg/Kg and 50 mg/Kg subcutaneously route; d) LCD67 showed weak activity at a dose of 50 mg/Kg. Those two compounds were very effective against parasites resistant to (ATN) and (ART) suggesting novel mechanisms of action. Thus, this research work gave new information of interest in the area potential new antimalarials.
Malaria remains one of the most important diseases in the world, causing the deaths of 1 million people annually and high morbidity. Given the spread of Plasmodium falciparum resistance to most antimalarial drugs available, the World Health Organization (WHO) recommended the use of artemisinin based combination therapies (ACTs) as first-line treatment since 2006 for uncomplicated malaria. In 2008, clinical reports have revealed the therapeutic failure of ACTs in Thailand-Cambodia border and since there is no alternative for the treatment of malaria it is essential to keep lines of research into new and effective drugs. From the discovery of artemisinin (ART) new peroxides emerged designated trioxolanes presenting the function 1,2,4-trioxane as pharmacophore. The accessibility, relatively inexpensive preparation and stability of the function 1,2,4-trioxane allow the synthesis of derivatives with different structures, extending the possibility of developing new drugs. Tests were performed in vitro screening with the MARK III (WHO micro-test) with the positive controls (artemisinin and dihidroartemisinin) and negative control (no drug). Assays were performed with three different compounds, here called NAC89, LCD67 and LC50 in cultures of P. falciparum (strain Dd2) as well as assays using the murine model of malaria, Plasmodium chabaudi, with 4 strains, denoted AS-3CQ, AS-ATN, respectively sensitive and resistant to artesunato (ATN) and AS-30CQ and AS-ART respectively sensitive and resistant to artemisinin. We also evaluated the cytotoxicity of the compounds using HepG2 human hepatoma cells, by using the colorimetric method metil-tiazol-tetrazolic (MTT) assay. In the murine model a comparison of two routes of administration of the new compounds was carried out for the 3 compounds NAC89, LCD67 and LC50 - subcutaneous inoculation while for LC50 we also assayed a topic administration. Cure was assessed by determining parasitaemia by microscopic observation of Giema stained blood smears. Results were: a) low cytotoxicity of the three compounds, b) compound LC50 eliminated parasites in in vitro assays in cultured P. falciparum and in the P. chabaudi model subcutaneously and topically at a dose of 50 mg/Kg and a 10 mg/Kg in subcutaneously, c) the NAC89 showed good activity in vivo in the same assay at a dose of 10 mg/Kg and 50 mg/Kg subcutaneously route; d) LCD67 showed weak activity at a dose of 50 mg/Kg. Those two compounds were very effective against parasites resistant to (ATN) and (ART) suggesting novel mechanisms of action. Thus, this research work gave new information of interest in the area potential new antimalarials.
Descrição
Palavras-chave
Parasitologia médica Malária Trioxolanos Modelo murino Plasmodium chabaudi Actividade in vitro e in vivo
Contexto Educativo
Citação
Editora
Instituto de Higiene e Medicina Tropical