Second-generation pterocarpanquinones: synthesis and antileishmanial activity

Despite the development of new therapies for leishmaniasis, among the 200 countries or territories reporting to the WHO, 87 were identified as endemic for Tegumentary Leishmaniasis and 75 as endemic for Visceral Leishmaniasis. The identification of antileishmanial drug candidates is essential to fill the drug discovery pipeline for leishmaniasis. In the hit molecule LQB-118 selected, the first generation of pterocarpanquinones was effective and safe against experimental visceral and cutaneous leishmaniasis via oral delivery. In this paper, we report the synthesis and antileishmanial activity of the second generation of pterocarpanoquinones. Methods: The second generation of pterocarpanquinones 2a-f was prepared through a palladium-catalyzed oxyarylation of dihydronaphtalen and chromens with iodolawsone, easily prepared by iodination of lawsone. The spectrum of antileishmanial activity was evaluated in promastigotes and intracellular amastigotes of L. amazonensis, L. braziliensis, and L. infantum. Toxicity was assessed in peritoneal macrophages and selective index calculated by CC50/IC50. Oxidative stress was measured by intracellular ROS levels and mitochondrial membrane potential in treated cells. Results: In this work, we answered two pertinent questions about the structure of the first-generation pterocarpanquinones: the configuration and positions of rings B (pyran) and C (furan) and the presence of oxygen in the B ring. When rings B and C are exchanged, we noted an improvement of the activity against promastigotes and amastigotes of L. amazonensis and promastigotes of L. infantum. As to the oxygen in ring B of the new generation, we observed that the oxygenated compound 2b is approximately twice as active against L. braziliensis promastigotes than its deoxy derivative 2a. Another modification that improved the activity was the addition of the methylenedioxy group. A variation in the susceptibility among species was evident in the clinically relevant form of the parasite, the intracellular amastigote. L. amazonensis was the species most susceptible to novel derivatives, whilst L. infantum was resistant to most of them. The pterocarpanoquinones (2b and 2c) that possess the oxygen atom in ring B showed induction of increased ROS production. Conclusions: The data presented indicate that the pterocarpanoquinones are promising compounds for the development of new leishmanicidal agents.(AU)

Estudos pré-clínicos da pterocarpanoquinona LQB-118: toxicidade subaguda, eficácia em leishmaniose visceral experimental e mecanismo de ação / Preclinical studies of pterocarpanquinone lqb-118: subacute toxicity, efficacy in experimental visceral leishmaniasis and mechanism of action

A leishmaniose visceral (LV) é a forma mais severa de leishmaniose e é a segunda maior causa de mortes por doenças parasitarias depois da malária. O arsenal terapêutico contra a leishmaniose é pequeno, e cada um dos medicamentos disponíveis apresenta ao menos uma das desvantagens: toxicidade, eficácia, preço ou regime de tratamento. Nós temos concentrado esforços em estudar novos candidatos a fármacos como alternativas aos tratamentos atuais. A pterocarpanquinona LQB-118 foi desenhada e sintetizada com base em hibridação molecular e apresentou atividade em protozoários e linhagens celulares de leucemia. Resultados prévios demonstraram que a LQB-118 foi eficaz no tratamento da leishmaniose cutânea experimental e que o mecanismo de ação envolve a indução do estresse oxidativo com eventos característicos da morte celular por apoptose em Leishmania amazonensis. Neste estudo, foi observado que o tratamento com 10 mg/kg/dia por via oral de LQB- 118 inibiu o desenvolvimento de hepatoesplenomegalia em camundongos infectados com L. infantum, com uma redução de 99 por cento na carga parasitária. A análise toxicológica in vivo não apresentou nenhuma mudança nos parâmetros clínicos, bioquímicos ou hematológicos. A análise histológica evidenciou que os órgãos não apresentaram anormalidades, com a exceção do fígado, no qual foi observado focos de necrose com infiltração leucocitária com uma dose cinco vezes maior do que a dose terapêutica. Entretando, estas alterações não foram acompanhadas por aumento das transaminases. Para avaliar os eventos iniciais do mecanismo de ação da LQB- 118, promastigotas de L. amazonensis foram incubados com LQB-118 e antioxidantes. As perdas da viabilidade celular e do potencial de membrana mitocondrial não foram revertidas com os antioxidantes, embora a produção de EROs tenha sido prevenida, sugerindo que a produção de EROs não é a causa primária de morte do parasito...

Visceral leishmaniasis (VL) is the most severe form of leishmaniasis and is the second major cause of deaths byparasites after malaria. The arsenal of drugs against leishmaniasis is small, and each has a disadvantage in termsof toxicity, efficacy, price or treatment regimen. Our group has focused on studying new drug candidates asalternatives to current treatments. The pterocarpanquinone LQB-118 was designed and synthesized based onmolecular hybridization and exhibited antiprotozoal and anti-leukemic cell line activity. Our previous workdemonstrated that LQB-118 was an effective treatment of experimental cutaneous leishmaniasis and that themechanism of action involves induction of oxidative stress with characteristic events of cell death via apoptosisin Leishmania amazonensis. In this study, we observed that treatment with 10 mg/kg/day LQB-118 orallyinhibited the development of hepatosplenomegaly in L. infantum-infected mice, with a 99 percent reduction in parasiteload. The in vivo toxicological analysis showed no change in clinical, biochemical or hematological parameters.Histologically, all of the analyzed organs were normal with the exception of liver, in which focal points ofnecrosis with leukocyte infiltration were observed with a dose five times higher than the therapeutic dose.However, these changes were not accompanied by increase in transaminases. To assess the early effects of themechanism of action of LQB-118, promastigotes of L. amazonensis were incubated with LQB-118 andantioxidants. Cell viability and mitochondrial membrane potential were not reversed by the antioxidants,although the ROS production was, suggesting that ROS production is not the primary cause of parasite death...

Antileishmanial activity of amides from Piper amalago and synthetic analogs

Two natural amides isolated from the chloroform extract of Piper amalago L., Piperaceae, leaves, a hydrogenated derivative and seven synthetic analogs were tested against the promastigote and intracellular amastigote forms of Leishmania amazonensis. The antileishmanial activity was evaluated in terms of growth inhibitory concentration for 50% of protozoa (IC50). The cytotoxicity toward the J774A1 macrophages was evaluated in terms of the cytotoxic concentrations for 50% of macrophages (CC50). The ability to induce nitric oxide production was also investigated for all compounds. The saturated amide 7-(1,3-benzodioxol-5-yl)-1-(1-pyrrolidinyl)-1-heptanone was obtained by hydrogenation of the natural compound N-[7-(3',4'-methylenedioxyphenyl)-2(Z),4(Z)-heptadienoyl]pyrrolidine. Synthetic amides were prepared by addition of the appropriate amine to the corresponding acyl chloride. The natural compound, N-[7-(3',4'-methylenedioxyphenyl)-2(E),4(E)-heptadienoyl]pyrrolidine, was the most active of all tested compounds against the promastigote and intracellular amastigote forms with IC50 values of 15 µM and 14.5 µM, respectively. None of the compounds modulated the production of nitric oxide.

Avaliação da atividade e mecanismo de ação da LQB-118 em leishmania amazonensis / Evaluation of activity and mechanism of action of LQB-118 in leishmania amazonensis

A leishmaniose é, ainda hoje, uma doença negligenciada, estando entre os agravos prioritários do programe de pesquisa sobre doenças da pobreza da Organização Mundial da Saúde (TDR/OMS). Além de não haver vacinas disponíveis, a terapia é baseada em medicamentos injetáveis que causam sérios efeitos colaterais, tornando o tratamento inviável para muitos países endêmicos. Fica evidente, então, a necessidade e urgência do desenvolvimento de novos medicamentos eficazes, seletivos, de baixo custo e, de preferência, de administração oral. A LQB-118 e seus análogos surgiram da união de dois grupos de moléculas bioativas, os pterocarpanos e as quinonas, formando as naftopterocarpanoquinonas, que vem demonstrando atividade leishmanicida. Neste trabalho avaliamos a atividade in vitro e in vivo do protótipo mais promissor. A LQB-118 demonstrou atividade leishmanicida em promastigotas de diferentes espécies de Leishmania, tanto do novo mundo como do velho mundo. No tratamento da leishmaniose cutânea em modelo murino, mostrou-se eficaz, tanto no tratamento subcutâneo por vias de administração sistêmica, oral e intraperitoneal. Ensaios bioquímicos indicaram que a LQB-118 induz um aumento expressivo da atividade de redutases mitocondriais nas primeiras horas, com concomitante aumento da produção de espécies reativas de oxigênio (ERO) pelos parasitos. Juntos, esses resultados indicaram que a LQB-118 é um protótipo seletivo, atuando primariamente sobre a mitocôndria dos parasitos, com atividade in vivo por via oral, o que a credencia como uma potencial candidata a fármaco.