In vivo and in vitro antimalarial effect and toxicological evaluation of the chloroquine analogue PQUI08001/06.

Antimalarial interventions mostly rely upon drugs, as chloroquine. However, plasmodial strains resistant to many drugs are constantly reported, leading to an expansion of malaria cases. Novel approaches are required to circumvent the drug resistance issue. Here, we describe the antimalarial potential of the chloroquine analogue 2-[[2-[(7-chloro-4-quinolinyl)amino]ethyl]amino] ethanol (PQUI08001/06). We observed that PQUI08001/06 treatment reduces parasitemia of both chloroquine-resistant and -sensitive strains of Plasmodium falciparum in vitro and P. berghei in vivo. Our data suggests that PQUI08001/06 is a potential antimalarial therapeutic alternative approach that could also target chloroquine-resistant plasmodial strains.

Design, Synthesis and Evaluation of New Fluoroamodiaquine Analogues.

Malaria is one of the most important tropical diseases; the use of amodiaquine as a current chemotherapy in the treatment of malaria has shown some problems such as hepatotoxicity and agranulocytosis. In this work we present the rational design, synthesis, and biological evaluation (antimalarial activity, cytotoxicity and genotoxicity) of four new fluoroamodiaquine analogues. The results showed significant correlation between MolDock score and IC50 values. The molecules 7b and c were the most active of the planned compounds, with lower IC50 against Plasmodium falciparum W2 strain (0.9 and 0.8 µM, respectively) and an excellent cytotoxicity profile. The present study revealed no mutagenicity or genotoxicity for the analogues. Confirming our docking results, the molecular dynamics showed that compound 7b remains stably bound to the heme group by means of π-stacking interactions between quinoline and the porphyrin ring. Based on these findings, this study may prove to be an efficient approach for the rational design of hemozoin inhibiting compounds to treat malaria.

Terpenic fraction of Pterodon pubescens inhibits nuclear factor kappa B and extracellular signal-regulated protein kinase 1/2 activation and deregulates gene expression in leukemia cells.

BACKGROUND: Plant derived compounds have been shown to be important sources of several anti-cancer agents. As cell cycle deregulation and tumor growth are intimately linked, the discovery of new substances targeting events in this biochemical pathway would be of great value. The anti-leukemic effect of an ethanolic extract of Pterodon pubescens seeds (EEPp) has been previously demonstrated and now we show that a terpenic subfraction (SF5) of EEPp containing farnesol, geranylgeraniol and vouacapan derivatives induces apoptosis in the human chronic myelogenous leukemia cell line K562. This work addresses SF5's antiproliferative mechanisms in these cells since they are still unclear. METHODS: DNA synthesis in K562 cells was assessed by [3H]-methyl-thymidine incorporation and cell cycle status by flow cytometry. The expression of cyclins D1 and E2, of the cell cycle inhibitor p21 and of the proto-oncogene c-myc was evaluated by semi-quantitative RT-PCR. Extracellular-signal-regulated kinases (ERK) 1/2 and nuclear factor kappa B (NF-κB) activation was evaluated by western blotting. RESULTS: In K562 cells, SF5 treatment induced a higher inhibition of DNA synthesis and cell growth than the original EEPp hexanic fraction from which SF5 originated, and also arrested the cell cycle in G1. Exposure of these cells to SF5 led to a decrease in cyclin E2 and c-myc expression while p21 mRNA levels were increased. Furthermore, SF5 inhibited the activation of mitogen-activated protein kinase (MAPK) ERK 1/2 and NF-κB. CONCLUSIONS: This work suggests that the anti-leukemic action of SF5 is linked to the inhibition of ERKs, NF-κB and c-myc signaling pathways resulting in reduced cyclin E2 mRNA expression and cell cycle arrest in the G1 phase.

Terpenic subfraction of Pterodon pubescens induces apoptosis of K562 leukemic cells by modulating gene expression.

Deregulation of cell proliferation and apoptosis is linked to malignant cell development. Leukemia is the most frequent cancer in children, and plants are important sources for new potential anti-cancer agents. Although anti-tumoral effects have been shown for Pterodon pubescens extracts, the mechanisms are still obscure. This study describes in Pterodon pubescens a furane diterpene only reported in Pterodon polygalaeflorus, the methyl-6α-acetoxy-7ß-hydroxyvouacapan-17ß-oate, indicated by HRMS and 13C-NMR analysis, and demonstrates some mechanisms of the anti-leukemia action of its terpene subfraction SF5. SF5 induced cytotoxic and anti-proliferative effects on K562 cells. Increased sub-G1 nuclei and Annexin V+-FITC cells confirmed apoptosis of leukemic cells by treatment of these cells with SF5. Down-regulation of DNMT1 gene transcription and over-expression of Apaf-1 mRNA suggested that SF5 may be inducing apoptosis of K562 cells by epigenetic up-regulation of pro-apoptotic proteins involved in the mitochondrial intrinsic pathway.

Pterodon pubescens seed extract induces the cell cycle arrest of leukemic cells by deregulating cyclin D1 and E2 mRNA levels.

Plant-derived compounds are important sources of effective anti-cancer agents. Pterodon pubescens is a native Brazilian plant popularly known for its anti-inflammatory and anti-arthritic effects. The ethanolic extract of its seeds (EEPp) is a viscous, brown and fragrant oil containing geranylgeraniol, farnesol, naphthalene, dimethyldodecatrienol and vouacapan diterpene derivatives, in addition to other compounds. This study investigated the in vitro anti-leukemic properties of EEPp using the resistant human leukemia cell line K562. The EEPp anti-proliferative effect was demonstrated by the inhibition of DNA synthesis and cell growth, and the induction of cell cycle arrest in the G(1) phase. Furthermore, cyclin E2 mRNA levels were down-regulated, while those of cyclin D1 were up-regulated. An EEPp anti-leukemic effect may have also triggered apoptosis, as it increased the number of shrunken cells and phosphatidylserine cell membrane exposure. These observations suggest that EEPp deregulates cyclin D1 and E2 expression, inducing cell cycle arrest and apoptosis of leukemic cells.

Avaliação dos mecanismos envolvidos na inibição da proliferação e na indução da apoptose de linhagem leucêmica pelo óleo de sementes de Pterodon pubescens e suas frações/subfrações / Assessment of the mechanisms involved in inhibiting proliferation and inducing apoptosis of lineage leukemia by oil seed Pterodon pubescens and its fractions/subfractions

Alterações no ciclo celular e desenvolvimento de tumor encontram-se intimamente ligados. Expressão inadequada e/ou mutações de quinases dependentes de ciclinas, ciclinas e inibidores de quinases dependentes de ciclinas têm sido descritas em vários tipos de cânceres. Portanto, a busca por novas molédulas que têm como alvo eventos em tal via bioquímica tem aumentado ao longo dos anos. Como as plantas são importantes fontes de novas potenciais drogas quimioterápicas, o estudo da fitoquímica, aliado às pesquisas farmacológicas, permite desvendar novas substâncias com potencial anti câncer. Embora efeitos antitumorais tenham sido demonstrados para extratos de Pterodon pubescens, os mecanismos que induzem estes efeitos ainda não estão esclarecidos. Portanto, o presente trabalho teve como objetivos subfracionar a fração hexano do extrato bruto de frutos de Pterodon pubescens, avaliar alguns mecanismos pelos quais a subfração mais ativa exerce seu efeito antiproliferativo sobre linhagem leucêmica (K562) e tentar identificar compostos envolvidos em tais efeitos. O extrato bruto (OPp) foi obtido por maceração em etanol e posteriormente foi feita uma partição líquido-liquido obtendo-se a fração Hexano (HEX). O subfracionamento por coluna de sílica da HEX resultou em oito subfrações, sendo que a subfração 5 (SF5) a que apresentou menor complexidade por cromatografia gasosa e maior efeito biológico. A citotoxicidade foi avaliada pelo método de redução do sal tetrazol (MTT) e a proliferação celular pela síntese de DNA e curva de crescimento. As fases do ciclo celular e a apoptose foram analisados por citometria de fluxo e a expressão do RNA mensageiro por RT-PCR. A expressão de proteína foi avaliada por Western blotting e a identificação de compostos por CG-EM e 13C-RMN. A SF5 exerceu efeito citotóxico maior do que o extrato bruto, fração HEX e as outras subfrações. Na síntese de DNA e curva de crescimento, a SF5 exerceu efeito inibitório tempo e concentração...

Alterations in the cell cycle and tumor development are closely connected. Inapropriated expression and/or mutations of cyclin dependet kinases, cyclins and cyclin kinase inhibitors have been described in many kinds of cancer. So, the search of new molecules that have such biochemical pathway as target has increased over the years. Since plants are important sources of new potential chemotherapic drugs, phytochemical studies combined to pharmacological investigations permit the discover of new substances with potential anti-cancer properties. Although anti tumor effects have been demonstrated for Pterodon pubescens extracts, its mechanisms are still obscure. Therefore, the aims of present work were to subfractionate the HEX fraction of Pterodon pubescens, evaluate some mechanisms by which the most active subfraction exerts its antiproliferative effect on leukemic cell line (K562) and try to elucidate compounds involved in such effects. The crude extract (PpO) was obtained by maceration in ethanol with further liquid-liquid partition yielding the Hexan fraction (HEX). The HEX subfractioning by silica column yielded 8 subfractions, being the subfraction 5 (SF5) that with minor complexity by gas chromatography and greater biological effect. The citotoxicity was evaluated by tetrazolium salt reduction (MTT) and cellular proliferation by DNA synthesis and growth curve. The cell cycle phases and apoptosis were analyzed by flow cytometry and messenger RNA expression by RT-PCR. The protein expression was evaluated by Western blotting and the identification of compounds by GC-MS and 13C-NMR. SF5 induced greater citotoxic effects when compared to crude extract, HEX fraction or the others subfractions. On DNA synthesis and growth curve, SF5 induced time and concentration dependent inhibiory effect, also greater than HEX fraction. The cell cycle analysis showed an increase of cells on G1 phase with consequently reduction of S and G2/M phases. SF5 inhibited the MAP...