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Despite advances in breast cancer treatment, there remains a need for local management of noninvasive, low-grade ductal carcinoma in situ (DCIS). These focal lesions are well suited for local intraductal treatment. Intraductal administration supported target site drug retention, improved efficacy, and reduced systemic exposure. Here, we used a poly(N-isopropyl acrylamide, pNIPAM) nanoparticle delivery system loaded with cytotoxic piplartine and an MAPKAP Kinase 2 inhibitor (YARA) for this purpose. For tumor environment targeting, a collagen-binding peptide SILY (RRANAALKAGELYKSILYGSG-hydrazide) was attached to pNIPAM nanoparticles, and the nanoparticle diameter, zeta potential, drug loading, and release were assessed. The system was evaluated for cytotoxicity in a 2D cell culture and 3D spheroids. In vivo efficacy was evaluated using a chemical carcinogenesis model in female Sprague-Dawley rats. Nanoparticle delivery significantly reduced the IC50 of piplartine (4.9 times) compared to the drug in solution. The combination of piplartine and YARA in nanoparticles further reduced the piplartine IC50 (~15 times). Treatment with these nanoparticles decreased the in vivo tumor incidence (5.2 times). Notably, the concentration of piplartine in mammary glands treated with nanoparticles (35.3 ± 22.4 µg/mL) was substantially higher than in plasma (0.7 ± 0.05 µg/mL), demonstrating targeted drug retention. These results indicate that our nanocarrier system effectively reduced tumor development with low systemic exposure.
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Chagas disease (CD) is one of the main neglected tropical diseases that promote relevant socioeconomic impacts in several countries. The therapeutic options for the treatment of CD are limited, and parasite resistance has been reported. Piplartine is a phenylpropanoid imide that has diverse biological activities, including trypanocidal action. Thus, the objective of the present work was to prepare a collection of thirteen esters analogous to piplartine (1-13) and evaluate their trypanocidal activity against Trypanosoma cruzi. Of the tested analogues, compound 11 ((E)-furan-2-ylmethyl 3-(3,4,5-trimethoxyphenyl)acrylate) showed good activity with IC50 values = 28.21 ± 5.34 µM and 47.02 ± 8.70 µM, against the epimastigote and trypomastigote forms, respectively. In addition, it showed a high rate of selectivity to the parasite. The trypanocidal mechanism of action occurs through the induction of oxidative stress and mitochondrial damage. In addition, scanning electron microscopy showed the formation of pores and leakage of cytoplasmic content. Molecular docking indicated that 11 probably produces a trypanocidal effect through a multi-target mechanism, including affinity with proteins CRK1, MPK13, GSK3B, AKR, UCE-1, and UCE-2, which are important for the survival of the parasite. Therefore, the results suggest chemical characteristics that can serve for the development of new trypanocidal prototypes for researching drugs against Chagas disease.
Assuntos
Doença de Chagas , Tripanossomicidas , Trypanosoma cruzi , Humanos , Tripanossomicidas/química , Simulação de Acoplamento Molecular , Doença de Chagas/tratamento farmacológico , Doença de Chagas/parasitologia , Estresse OxidativoRESUMO
Cancer is a principal cause of death in the world, and providing a better quality of life and reducing mortality through effective pharmacological treatment remains a challenge. Among malignant tumor types, squamous cell carcinoma-esophageal cancer (EC) is usually located in the mouth, with approximately 90% located mainly on the tongue and floor of the mouth. Piplartine is an alkamide found in certain species of the genus Piper and presents many pharmacological properties including antitumor activity. In the present study, the cytotoxic potential of a collection of piplartine analogs against human oral SCC9 carcinoma cells was evaluated. The analogs were prepared via Fischer esterification reactions, alkyl and aryl halide esterification, and a coupling reaction with PyBOP using the natural compound 3,4,5-trimethoxybenzoic acid as a starting material. The products were structurally characterized using 1H and 13C nuclear magnetic resonance, infrared spectroscopy, and high-resolution mass spectrometry for the unpublished compounds. The compound 4-methoxy-benzyl 3,4,5-trimethoxybenzoate (9) presented an IC50 of 46.21 µM, high selectively (SI > 16), and caused apoptosis in SCC9 cancer cells. The molecular modeling study suggested a multi-target mechanism of action for the antitumor activity of compound 9 with CRM1 as the main target receptor.
Assuntos
Neoplasias Bucais , Carcinoma de Células Escamosas de Cabeça e Pescoço , Humanos , Linhagem Celular Tumoral , Neoplasias Bucais/tratamento farmacológico , Qualidade de Vida , Carcinoma de Células Escamosas de Cabeça e Pescoço/tratamento farmacológico , Piperidonas/farmacologiaRESUMO
Nanoemulsions modified with chitosan (NE-Q) or hyaluronic acid (NE-HA), developed for intraductal administration of piplartine (piperlongumine) and local breast cancer treatment, were evaluated for cytotoxic effects in vitro in 2D and 3D breast cancer models and in vivo in a chemically induced carcinogenesis model. Droplet size was lower than 100 nm, and zeta potential varied from +17.9 to -25.5 mV for NE-Q and NE-HA, respectively. Piplartine nanoencapsulation reduced its IC50 up to 3.6-fold in T-47D and MCF-7 monolayers without differences between NE-Q and NE-HA, and up to 6.6-fold in cancer spheroids. Cytotoxicity improvement may result from a more efficient NE-mediated delivery, as suggested by stronger fluorescent staining of cells and spheroids. In 1-methyl-1-nitrosourea -induced breast cancer models, intraductal administration of piplartine-loaded NE-HA inhibited breast tumor development and histological alterations. These results support the potential applicability of piplartine-loaded NE-HA for intraductal treatment of breast cancer.
Assuntos
Neoplasias da Mama , Quitosana , Nanopartículas , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Quitosana/farmacologia , Feminino , Humanos , Ácido Hialurônico/farmacologia , PiperidonasRESUMO
A library of nine hybrids of 4-hydroxygoniothalamin (2), 4-hydroxypiplartine (4), monastrol (5) and oxo-monastrol (6) was prepared via a modular synthetic route with a diester or a 1,2,3-triazole as linkers. The compounds were assayed against a panel of human cancer cell lines, including MCF-7 (breast adenocarcinoma), HeLa (cervical adenocarcinoma), Caco-2 (colorectal adenocarcinoma) and PC3 (prostate adenocarcinoma), as well as against normal breast (MCF10A) and prostate (PNT2) cells. In general, hybrids with an ester linker containing 4-hydroxypiplartine (4) were more potent than the corresponding hybrids with 4-hydroxygoniothalamin (2). On the other hand, compounds presenting the 1,2,3-triazole linker displayed enhanced cytotoxicity and selectivity when compared to their corresponding hybrids with the diester linker. The 4-hydroxypiplartine-based hybrids 12 and 22 displayed high cytotoxicity (IC50 values below 10 µM) against all cancer cells studied, especially in MCF-7 cells with IC50 values of 1.7 ± 0.1 and 1.6 ± 0.9 µM, respectively. Furthermore, the 4-hydroxygoniothalamin-monastrol hybrid (compound 21) and the 4-hydroxypiplartine-oxo-monastrol hybrid (compound 25), both bearing a 1,2,3-triazole linker, displayed high selectivity and potency towards breast cancer cell line (MCF-7 vs. MCF10 cells, selectivity index = 15.8 and 7.1, respectively), while the 4-hydroxypiplartine -4-hydroxymethylgoniothalamin hybrid with a diester linker (compound 33) showed high selectivity towards melanoma cancer cells (selectivity index = 9.6). Antiproliferative and pro-apoptotic potential of compounds 12 and 22 against MCF-7 cancer cells were further investigated. Cell cycle studies revealed increased G2/M population in MCF-7 cultures as well as reduced G0/G1 population compared to the control groups indicating cell cycle arrest in G2/M phase. In addition, the frequency of positive cells for annexin V was higher in treated samples suggesting that compounds 12 and 22 induce apoptosis in estrogen-positive MCF-7 cells.
Assuntos
Antineoplásicos/farmacologia , Piperidonas/farmacologia , Pironas/farmacologia , Triazóis/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Estrutura Molecular , Piperidonas/química , Pironas/química , Relação Estrutura-Atividade , Triazóis/químicaRESUMO
Due to the limited options for topical management of skin cancer, this study aimed at developing and evaluating nanoemulsions (NE) for topical delivery of the cytotoxic agent piplartine (piperlongumine). NEs were modified with chitosan or sodium alginate, and the effects on the physicochemical properties, piplartine delivery and formulation efficacy were evaluated. The nanoemulsion droplets displayed similar size (96-112 nm), but opposite charge; the polysaccharides improved piplartine penetration into and across the skin (1.3-1.9-fold) in a similar manner, increasing the ratio "drug in the skin/receptor phase" by 1.4-1.5-fold compared to the plain NE and highlighting their relevance for cutaneous localization. Oleic acid addition to the chitosan-containing NE further increased drug penetration (~1.9-2.0-fold), as did increases in drug content from 0.5 to 1%. The cytotoxicity of piplartine was ~2.8-fold higher when the drug was incorporated in the chitosan-containing NE compared to its solution (IC50 = 14.6 µM) against melanoma cells. The effects of this nanocarrier on 3D melanoma tissues were concentration-related; at 1%, piplartine elicited marked epidermis destruction. These results support the potential applicability of the chitosan-modified nanoemulsion containing piplartine as a new strategy for local management of skin cancer.
Assuntos
Emulsões/química , Melanoma/tratamento farmacológico , Nanopartículas/química , Neoplasias Cutâneas/tratamento farmacológico , Alginatos/química , Proliferação de Células/efeitos dos fármacos , Quitosana/química , Citotoxinas/química , Emulsões/farmacologia , Humanos , Melanócitos/efeitos dos fármacos , Melanócitos/patologia , Piperidonas/química , Piperidonas/farmacologia , Neoplasias Cutâneas/patologiaRESUMO
Schistosomiasis is one of the most important parasitic infections in terms of its negative effects on public health and economics. Since praziquantel is currently the only drug available to treat schistosomiasis, there is an urgent need to identify new anthelmintic agents. Piplartine, also known as piperlongumine, is a biologically active alkaloid/amide from peppers that can be detected in high amounts in the roots of Piper tuberculatum. Previously, it has been shown to have in vitro schistosomicidal effects. However, its anthelmintic activity in an animal host has not been reported. In the present work, in vivo antischistosomal properties of isolated piplartine were evaluated in a mouse model of schistosomiasis infected with either adult (patent infection) or juvenile (pre-patent infection) stages of Schistosoma mansoni. A single dose of piplartine (100, 200 or 400 mg/kg) or daily doses for five consecutive days (100 mg/kg/day) administered orally to mice infected with schistosomes resulted in a reduction in worm burden and egg production. Treatment with the highest piplartine dose (400 mg/kg) caused a significant reduction in a total worm burden of 60.4% (P < 0.001) in mice harbouring adult parasites. S. mansoni egg production, a process responsible for pathology in schistosomiasis, was also significantly inhibited by piplartine. Studies using scanning electron microscopy revealed substantial tegumental alterations in parasites recovered from mice. Since piplartine has well-characterized mechanisms of toxicity, is easily available, and is cost-effective, our results indicate that this bioactive molecule derived from medicinal plants could be a potential lead compound for novel antischistosomal agents.
Assuntos
Piperidonas/uso terapêutico , Esquistossomose mansoni/tratamento farmacológico , Esquistossomicidas/uso terapêutico , Animais , Modelos Animais de Doenças , Feminino , Camundongos , Piper/químicaRESUMO
Ruthenium complexes with piplartine, [Ru(piplartine)(dppf)(bipy)](PF6)2 (1) and [Ru(piplartine)(dppb)(bipy)](PF6)2 (2) (dppf = 1,1-bis(diphenylphosphino) ferrocene; dppb = 1,4-bis(diphenylphosphino)butane and bipy = 2,2'-bipyridine), were recently synthesized and displayed more potent cytotoxicity than piplartine in different cancer cells, regulated RNA transcripts of several apoptosis-related genes, and induced reactive oxygen species (ROS)-mediated apoptosis in human colon carcinoma HCT116 cells. The present work aimed to explore the underlying mechanisms through which these ruthenium complexes induce cell death in HCT116 cells in vitro, as well as their in vivo action in a xenograft model. Both complexes significantly increased the percentage of apoptotic HCT116 cells, and co-treatment with inhibitors of JNK/SAPK, p38 MAPK, and MEK, which inhibits the activation of ERK1/2, significantly reduced the apoptosis rate induced by these complexes. Moreover, significant increase in phospho-JNK2 (T183/Y185), phospho-p38α (T180/Y182), and phospho-ERK1 (T202/Y204) expressions were observed in cells treated with these complexes, indicating MAPK-mediated apoptosis. In addition, co-treatment with a p53 inhibitor (cyclic pifithrin-α) and the ruthenium complexes significantly reduced the apoptosis rate in HCT116 cells, and increased phospho-p53 (S15) and phospho-histone H2AX (S139) expressions, indicating induction of DNA damage and p53-dependent apoptosis. Both complexes also reduced HCT116 cell growth in a xenograft model. Tumor mass inhibition rates were 35.06, 29.71, and 32.03% for the complex 1 (15 µmol/kg/day), complex 2 (15 µmol/kg/day), and piplartine (60 µmol/kg/day), respectively. These data indicate these ruthenium complexes as new anti-colon cancer drugs candidates.
RESUMO
As a new strategy for treatment of ductal carcinoma in situ, biocompatible and bioadhesive nanoemulsions for intraductal administration of the cytotoxic agent piplartine (piperlongumine) were optimized in this study. To confer bioadhesive properties, the nanoemulsion was modified with chitosan or hyaluronic acid. Tricaprylin was selected as the nanoemulsion non-polar phase due to its ability to dissolve larger drug amounts compared to isopropyl myristate and monocaprylin. Use of phosphatidylcholine as sole surfactant did not result in a homogeneous nanoemulsion, while its association with polysorbate 80 and glycerol (in a surfactant blend) led to the formation of nanoemulsions with droplet size of 76.5⯱â¯1.2â¯nm. Heating the aqueous phase to 50⯰C enabled sonication time reduction from 20 to 10â¯min. Inclusion of either chitosan or hyaluronic acid resulted in nanoemulsions with similar in vitro bioadhesive potential, and comparable ability to prolong mammary tissue retention (to 120â¯h) in vivo without causing undesirable histological alterations. Piplartine was stable in both nanoemulsions for 60â¯days; however, the size of loaded NE-HA was maintained at a similar range for longer periods of time, suggesting that this nanoemulsion may be a stronger candidate for intraductal delivery.
Assuntos
Antineoplásicos Fitogênicos/administração & dosagem , Dioxolanos/administração & dosagem , Glândulas Mamárias Animais/metabolismo , Nanopartículas/administração & dosagem , Piperidonas/administração & dosagem , Adesividade , Animais , Antineoplásicos Fitogênicos/química , Galinhas , Quitosana/administração & dosagem , Quitosana/química , Membrana Corioalantoide/efeitos dos fármacos , Dioxolanos/química , Vias de Administração de Medicamentos , Emulsões , Feminino , Glicerol/administração & dosagem , Glicerol/química , Ácido Hialurônico/administração & dosagem , Ácido Hialurônico/química , Nanopartículas/química , Fosfatidilcolinas/administração & dosagem , Fosfatidilcolinas/química , Piperidonas/química , Polissorbatos/administração & dosagem , Polissorbatos/química , Ratos Wistar , Pele/química , SuínosRESUMO
Piplartines are alkaloid amides present in the roots and stems of different pepper species which have promising pharmacological properties including cancer prevention. Some recent studies have determined pharmacokinetic parameters of piplartine in rat blood plasma but without pointing to any molecular target or describing the physicochemical forces of the interaction. The present study investigated the interaction between piplartine and human serum albumin (HSA) the predominant protein in blood plasma. Fluorescence spectroscopy was utilized to observe the complex HSA-piplartine formation. Thermodynamic parameter analysis indicates that the process occurs spontaneously and is enthalpically driven; the affinity constant suggests that this interaction is reversible. This was reinforced by the binding density function method and by the displacement analysis that the piplartine binds on HSA at a single site, which was determined to be the IIA sub-domain. In silico analysis (molecular docking) identified the main residues involved in binding and the corresponding forces, which corroborates well with the experimental results.
Assuntos
Piperidonas/química , Piperidonas/metabolismo , Albumina Sérica Humana/química , Albumina Sérica Humana/metabolismo , Sítios de Ligação , Simulação por Computador , Humanos , Simulação de Acoplamento Molecular , Espectrometria de Fluorescência , Espectrofotometria Ultravioleta , TermodinâmicaRESUMO
This study reports the development of a simple and reproducible method, with high rates of recovery, to extract the cytotoxic agent piplartine from skin layers, and a sensitive and rapid UV-HPLC method for its quantification. Considering the potential of piplartine for topical treatment of skin cancer, this method may find application for formulation development and pharmacokinetics studies to assess cutaneous bioavailability. Porcine skin was employed as a model for human tissue. Piplartine was extracted from the stratum corneum (SC) and remaining viable skin layers (VS) using methanol, vortex homogenization and bath sonication, and subsequently assayed by HPLC using a C18 column, and 1:1 (v/v) acetonitrile-water (adjusted to pH 4.0 with acetic acid 0.1%) as mobile phase. The quantification limit of piplartine was 0.2 µg/mL (0.6 µm), and the assay was linear up to 5 µg/mL (15.8 µm), with within-day and between-days assay coefficients of variation and relative errors <15%. Piplartine recovery from SC and VS varied from 86 to 96%. The method was suitable to assay samples from skin penetration studies, enabling detection of differences in cutaneous delivery in different skin compartments resulting from treatment with various formulations and time periods.
Assuntos
Antineoplásicos Fitogênicos/análise , Dioxolanos/análise , Piperidonas/análise , Pele/química , Animais , Antineoplásicos Fitogênicos/farmacocinética , Disponibilidade Biológica , Cromatografia Líquida de Alta Pressão , Dioxolanos/farmacocinética , Modelos Lineares , Piperidonas/farmacocinética , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Pele/metabolismo , Absorção Cutânea , SuínosRESUMO
Piplartine (piperlongumine) is a plant-derived compound found in some Piper species that became a novel potential antineoplastic agent. In the present study, we synthesized a novel platinum complex containing a piplartine derivative cis-[PtCl(PIP-OH)(PPh3)2]PF6 (where, PIP-OH = piplartine demethylated derivative; and PPh3 = triphenylphosphine) with enhanced cytotoxicity in different cancer cells, and investigated its apoptotic action in human promyelocytic leukemia HL-60 cells. The structure of PIP-OH ligand was characterized by X-ray crystallographic analysis and the resulting platinum complex was characterized by infrared, molar conductance measurements, elemental analysis and NMR experiments. We found that the complex is more potent than piplartine in a panel of cancer cell lines. Apoptotic cell morphology, increased internucleosomal DNA fragmentation, without cell membrane permeability, loss of the mitochondrial transmembrane potential, increased phosphatidylserine externalization and caspase-3 activation were observed in complex-treated HL-60 cells. Treatment with the complex also caused a marked increase in the production of reactive oxygen species (ROS), and the pretreatment with N-acetyl-L-cysteine, an antioxidant, reduced the complex-induced apoptosis, indicating activation of ROS-mediated apoptosis pathway. Important, pretreatment with a p38 MAPK inhibitor (PD 169316) and MEK inhibitor (U-0126), known to inhibit ERK1/2 activation, also prevented the complex-induced apoptosis. The complex did not induce DNA intercalation in cell-free DNA assays. In conclusion, the complex exhibits more potent cytotoxicity than piplartine in a panel of different cancer cells and triggers ROS/ERK/p38-mediated apoptosis in HL-60 cells.
Assuntos
Apoptose/efeitos dos fármacos , Leucemia Promielocítica Aguda/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Platina/farmacologia , Caspases/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Células HL-60 , Humanos , Leucemia Promielocítica Aguda/tratamento farmacológico , Leucemia Promielocítica Aguda/patologia , Ligantes , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Modelos Moleculares , Estrutura Molecular , Platina/química , Espécies Reativas de Oxigênio/metabolismoRESUMO
The hemoflagellate protozoan, Trypanosoma cruzi, mainly transmitted by triatomine insects through blood transfusion or from mother-to-child, causes Chagas' disease. This is a serious parasitic disease that occurs in Latin America, with considerable social and economic impact. Nifurtimox and benznidazole, drugs indicated for treating infected persons, are effective in the acute phase, but poorly effective during the chronic phase. Therefore, it is extremely urgent to find innovative chemotherapeutic agents and/or effective vaccines. Since piplartine has several biological activities, including trypanocidal activity, the present study aimed to evaluate it on two T. cruzi strains proteome. Considerable changes in the expression of some important enzymes involved in parasite protection against oxidative stress, such as tryparedoxin peroxidase (TXNPx) and methionine sulfoxide reductase (MSR) was observed in both strains. These findings suggest that blocking the expression of the two enzymes could be potential targets for therapeutic studies.
Assuntos
Piperidonas/farmacologia , Extratos Vegetais/farmacologia , Proteínas/análise , Tripanossomicidas/farmacologia , Trypanosoma cruzi/química , Trypanosoma cruzi/efeitos dos fármacos , Eletroforese em Gel Bidimensional , Espectrometria de Massas , Estresse Oxidativo , Proteômica , Valores de Referência , Reprodutibilidade dos Testes , Trypanosoma cruzi/metabolismoRESUMO
ABSTRACT The hemoflagellate protozoan, Trypanosoma cruzi, mainly transmitted by triatomine insects through blood transfusion or from mother-to-child, causes Chagas' disease. This is a serious parasitic disease that occurs in Latin America, with considerable social and economic impact. Nifurtimox and benznidazole, drugs indicated for treating infected persons, are effective in the acute phase, but poorly effective during the chronic phase. Therefore, it is extremely urgent to find innovative chemotherapeutic agents and/or effective vaccines. Since piplartine has several biological activities, including trypanocidal activity, the present study aimed to evaluate it on two T. cruzi strains proteome. Considerable changes in the expression of some important enzymes involved in parasite protection against oxidative stress, such as tryparedoxin peroxidase (TXNPx) and methionine sulfoxide reductase (MSR) was observed in both strains. These findings suggest that blocking the expression of the two enzymes could be potential targets for therapeutic studies.
Assuntos
Piperidonas/farmacologia , Tripanossomicidas/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Trypanosoma cruzi/química , Extratos Vegetais/farmacologia , Proteínas/análise , Valores de Referência , Espectrometria de Massas , Trypanosoma cruzi/metabolismo , Eletroforese em Gel Bidimensional , Reprodutibilidade dos Testes , Estresse Oxidativo , ProteômicaRESUMO
Piplartine (piperlongumine) is a plant-derived molecule that has been receiving intense interest due to its anticancer characteristics that target the oxidative stress. In the present paper, two novel piplartine-containing ruthenium complexes [Ru(piplartine)(dppf)(bipy)](PF6)2 (1) and [Ru(piplartine)(dppb)(bipy)](PF6)2 (2) were synthesized and investigated for their cellular and molecular responses on cancer cell lines. We found that both complexes are more potent than metal-free piplartine in a panel of cancer cell lines on monolayer cultures, as well in 3D model of cancer multicellular spheroids formed from human colon carcinoma HCT116 cells. Mechanistic studies uncovered that the complexes reduced the cell growth and caused phosphatidylserine externalization, internucleosomal DNA fragmentation, caspase-3 activation and loss of the mitochondrial transmembrane potential on HCT116 cells. Moreover, the pre-treatment with Z-VAD(OMe)-FMK, a pan-caspase inhibitor, reduced the complexes-induced apoptosis, indicating cell death by apoptosis through caspase-dependent and mitochondrial intrinsic pathways. Treatment with the complexes also caused a marked increase in the production of reactive oxygen species (ROS), including hydrogen peroxide, superoxide anion and nitric oxide, and decreased reduced glutathione levels. Application of N-acetyl-cysteine, an antioxidant, reduced the ROS levels and apoptosis induced by the complexes, indicating activation of ROS-mediated apoptosis pathway. RNA transcripts of several genes, including gene related to the cell cycle, apoptosis and oxidative stress, were regulated under treatment. However, the complexes failed to induce DNA intercalation. In conclusion, the complexes are more potent than piplartine against different cancer cell lines and are able to induce caspase-dependent and mitochondrial intrinsic apoptosis on HCT116 cells by ROS-mediated pathway.
RESUMO
Piplartine, an alkaloid produced by plants in the genus Piper, displays promising anticancer activity. Understanding the gas-phase fragmentation of piplartine by electrospray ionization tandem mass spectrometry can be a useful tool to characterize biotransformed compounds produced by in vitro and in vivo metabolism studies. As part of our efforts to understand natural product fragmentation in electrospray ionization tandem mass spectrometry, the gas-phase fragmentation of piplartine and its two metabolites 3,4-dihydropiplartine and 8,9-dihydropiplartine, produced by the endophytic fungus Penicillium crustosum VR4 biotransformation, were systematically investigated. Proposed fragmentation reactions were supported by ESI-MS/MS data and computational thermochemistry. Cleavage of the C-7 and N-amide bond, followed by the formation of an acylium ion, were characteristic fragmentation reactions of piplartine and its analogs. The production of the acylium ion was followed by three consecutive and competitive reactions that involved methyl and methoxyl radical eliminations and neutral CO elimination, followed by the formation of a four-member ring with a stabilized tertiary carbocation. The absence of a double bond between carbons C-8 and C-9 in 8,9-dihydropiplartine destabilized the acylium ion and resulted in a fragmentation pathway not observed for piplartine and 3,4-dihydropiplartine. These results contribute to the further understanding of alkaloid gas-phase fragmentation and the future identification of piplartine metabolites and analogs using tandem mass spectrometry techniques. Copyright © 2017 John Wiley & Sons, Ltd.
Assuntos
Antineoplásicos Fitogênicos/metabolismo , Ascomicetos/metabolismo , Piperidonas/metabolismo , Biotransformação , Gases , Hidrogenação , Metabolômica , Simulação de Dinâmica Molecular , Estrutura Molecular , Espectrometria de Massas em TandemRESUMO
Schistosomiasis is a world health problem, and praziquantel is the only drug currently used for the treatment. There is some evidence that extensive monotherapy of praziquantel may be leading to drug resistance in the parasite. In order to find alternative treatments, the effects of the combination of epiisopiloturine (EPI), piplartine (PPT) and praziquantel (PZQ) were evaluated. Similarity analysis of these compounds was performed using optimized molecular structures to compare the shape and the charge modeling of combinations between PZQ and EPI or PPT. Supported by this data, in vitro association of PZQ-PPT, PZQ-EPI, and EPI-PPT was carried out, and the activity of these combinations against Schistosoma mansoni was assessed. The results showed synergistic activity with a combination index (CI) of 0.42 for the treatment with PZQ-PPT. Both PZQ-EPI and EPI-PPT combinations also showed synergistic effects, with CI values of 0.86 and 0.61, respectively. Surface alterations in the tegument of adult schistosomes after the treatments were observed using laser confocal microscopy and scanning electron microscopy. Additionally, the association of EPI-PPT decreased the cytotoxicity when compared with both isolated compounds in three different lines of mammalian cells. Thus, synergistic combinations of PZQ-PPT, PZQ-EPI, and EPI-PPT create the possibility of reduced doses to be used against Schistosoma mansoni.
Assuntos
4-Butirolactona/análogos & derivados , Imidazóis/farmacologia , Piperidonas/farmacologia , Praziquantel/farmacologia , Schistosoma mansoni/efeitos dos fármacos , 4-Butirolactona/química , 4-Butirolactona/farmacologia , Animais , Antiprotozoários/farmacologia , Forma Celular/efeitos dos fármacos , Chlorocebus aethiops , Cricetinae , Cães , Sinergismo Farmacológico , Quimioterapia Combinada , Imidazóis/química , Células Madin Darby de Rim Canino , Masculino , Camundongos , Microscopia Confocal , Piperidonas/química , Praziquantel/química , Schistosoma mansoni/ultraestrutura , Células VeroRESUMO
Because piplartine (PPT) has demonstrated biological activities, such as cytotoxic, anxiolytic, antidepressant, antifungal and antiplatelet activities, this molecule is a relevant drug candidate. The metabolic fate of drug candidates is an essential requirement in assessing their safety and efficacy. Based on this requirement, the biotransformation of PPT by cytochrome P450 enzymes (CYP) was investigated for the first time. To determine the in vitro enzymatic kinetic parameters, an HPLC method was developed and validated to quantify PPT. All samples were separated on a reversed-phase C18 column using a mobile phase of acetonitrile:water (40:60, v/v). The method exhibited a linear range of 2.4-157.7 µmol/L, with the following calibration curve: y=0.0934 (±0.0010)x+0.0027, r=0.9975. The lower limit of quantitation was verified to be 2.4 µmol/L, with an RSD below 7%. The precision and accuracy were assessed for both within-day and between-day determinations; neither relative standard (RSD%) deviations nor relative errors (RER) exceeded a value of 15%. The mean absolute recovery was 85%, with an RSD value below 6%. The enzymatic kinetic parameters revealed a sigmoidal profile, with V(max)=4.7±0.3 µmol/mg mL⻹/min, h=2.5±0.4, S50=44.7±0.3 µmol/L and CL(max)=0.054 µL/min/mg protein, indicating cooperativity behavior. Employing a mammalian model, PPT metabolism yielded two unreported monohydroxylated products (m/z 334). The identification and structural elucidation of the metabolites were performed by comparing their mass spectra with those spectra of the parent drug. For the first time, the in vitro metabolism studies employing microsomes were demonstrated to be a suitable tool for data regarding enzymatic kinetics and for the metabolites formed in the PPT mammalian metabolism.
Assuntos
Microssomos Hepáticos/metabolismo , Piperidonas/metabolismo , Animais , Cromatografia Líquida de Alta Pressão , Estabilidade de Medicamentos , Masculino , Ratos , Ratos Wistar , Espectrometria de Massas por Ionização por Electrospray , Espectrometria de Massas em TandemRESUMO
Piper is a notable genus among Piperaceae due to their secondary metabolites such as lignans, amides, esters and long chain fatty acids used as anti-herbivore defenses with comparable effects of pyrethroids, that holds a promise in insect control, including malaria vectors such as Anopheles darlingi, the main vector in the North of Brazil. Methanolic extracts of Piper tuberculatum Jacq., Piperaceae, and P. alatabaccum Trel. & Yunck., Piperaceae, and some isolated compounds, i.e, 3,4,5-trimetoxy-dihydrocinamic acid, dihydropiplartine; piplartine, piplartine-dihydropiplartine and 5,5',7-trimetoxy-3',4'-metilenodioxiflavone were tested as larvicides against A. darlingi. The Lethal Concentrations (LC50 and LC90) of methanolic extracts were 194 and 333 ppm for P. tuberculatum and 235 and 401 ppm for P. alatabacum, respectively. Isolated compounds had lower LC values, e.g. the LC50 and LC90 of the piplartine-dihidropiplartine isolated from both plant species was 40 and 79 ppm, respectively.
RESUMO
Schistosomiasis is one of the worlds greatly neglected tropical diseases, and its control is largely dependenton a single drug, praziquantel. Here, we report the in vitro effect of piplartine, an amide isolated from Piper tuberculatum (Piperaceae), on Schistosoma mansoni adult worms. A piplartine concentrationof 15.8 lM reduced the motor activity of worms and caused their death within 24 h in a RPMI 1640 medium.Similarly, the highest sub-lethal concentration of piplartine (6.3 lM) caused a 75% reduction in eggproduction in spite of coupling. Additionally, piplartine induced morphological changes on the tegument,and a quantitative analysis carried out by confocal microscopy revealed an extensive tegumental destructionand damage in the tubercles. This damage was dose-dependent in the range of 15.8630.2 lM. At doses higher than 157.6 lM, piplartine induced morphological changes in the oral and ventral sucker regions of the worms. It is the first time that the schistosomicidal activity has been reported forpiplartine.