Renal protective effect of ellipticine against streptozotocin induced diabetic nephropathy in rats via suppression of oxidative stress and inflammatory mediator.

PURPOSE: Diabetes mellitus is a serious health problem worldwide, and diabetic nephropathy is the complication. The diabetic nephropathy considerably enhances the oxidative stress, glycation, lipid parameters and inflammatory reaction. Ellipticine has potent free radical scavenging and anti-inflammatory effect. METHODS: In the current study, our objectives were to thoroughly examine the renal protective effects of ellipticine in a rat model of streptozotocin (STZ)-induced diabetic nephropathy (DN) and to elucidate the underlying mechanisms involved. For the induction of diabetic nephropathy, streptozotocin (50 mg/kg) was used, and rats were separated into groups and given varying doses of ellipticine (2.5, 5 and 7.5 mg/kg). The body weight, and renal weight were estimated. The inflammatory cytokines, renal biomarkers, inflammatory antioxidant, and urine parameters were estimated. RESULTS: Result showed that ellipticine considerably enhanced the body weight and reduced the renal tissue weight. Ellipticine treatment significantly (P < 0.001) repressed the level of blood urea nitrogen, serum creatinine, uric acid, blood glucose and altered the lipid parameters. Ellipticine significantly (P < 0.001) repressed the level of malonaldehyde and boosted the glutathione, catalase, superoxide dismutase, and glutathione peroxidase. Ellipticine treatment significantly (P < 0.001) reduced the inflammatory cytokines and inflammatory mediators. CONCLUSIONS: Ellipticine could be a renal protective drug via attenuating the inflammatory reaction, fibrosis and oxidative stress in streptozotocin induced rats.

Renal protective effect of ellipticine against streptozotocin induced diabetic nephropathy in rats via suppression of oxidative stress and inflammatory mediator

Purpose: Diabetes mellitus is a serious health problem worldwide, and diabetic nephropathy is the complication. The diabetic nephropathy considerably enhances the oxidative stress, glycation, lipid parameters and inflammatory reaction. Ellipticine has potent free radical scavenging and anti-inflammatory effect. Methods: In the current study, our objectives were to thoroughly examine the renal protective effects of ellipticine in a rat model of streptozotocin (STZ)-induced diabetic nephropathy (DN) and to elucidate the underlying mechanisms involved. For the induction of diabetic nephropathy, streptozotocin (50 mg/kg) was used, and rats were separated into groups and given varying doses of ellipticine (2.5, 5 and 7.5 mg/kg). The body weight, and renal weight were estimated. The inflammatory cytokines, renal biomarkers, inflammatory antioxidant, and urine parameters were estimated. Results: Result showed that ellipticine considerably enhanced the body weight and reduced the renal tissue weight. Ellipticine treatment significantly (P < 0.001) repressed the level of blood urea nitrogen, serum creatinine, uric acid, blood glucose and altered the lipid parameters. Ellipticine significantly (P < 0.001) repressed the level of malonaldehyde and boosted the glutathione, catalase, superoxide dismutase, and glutathione peroxidase. Ellipticine treatment significantly (P < 0.001) reduced the inflammatory cytokines and inflammatory mediators. Conclusions: Ellipticine could be a renal protective drug via attenuating the inflammatory reaction, fibrosis and oxidative stress in streptozotocin induced rats.

Divergent Pd-catalyzed Functionalization of 4-Oxazolin-2-ones and 4-Methylene-2-oxazolidinones and Synthesis of Heterocyclic-Fused Indoles.

Palladium-catalyzed functionalization was presently performed on two building blocks: 4-oxazolin-2-ones and 4-methylene-2-oxazolidinones. Direct Heck arylation of 4-oxazolin-2-ones led to a series of 5-aryl-4-oxazolin-2-ones, including analogues with N-chiral auxiliary, in an almost quantitative yield. The Pd(II)-catalyzed homocoupling reaction of 4-oxazolin-2-ones provided novel heterocyclic across-ring dienes. Meanwhile, the intramolecular cross-coupling of N-aryl-4-methylene-2-oxazolidinones furnished a series of oxazolo[3,4-a]indol-3-ones. Further functionalization of 4-methylene-2-oxazolidinones afforded substituted indoles and heterocyclic-fused indoles with aryl, bromo, carbinol, formyl, and vinyl groups. A computational study was carried out to account for the behavior of the formylated derivatives. The currently developed methodology was applied to a new formal total synthesis of ellipticine.

The potential mechanism of Chebulae Fructus in the treatment of hepatocellular carcinoma on the basis of network pharmacology.

INTRODUCTION AND OBJECTIVES: Hepatocellular carcinoma (HCC) ranks third on the list of the leading cause for cancer death globally. The treatment of HCC patients is unsatisfactory. However, the traditional Chinese medicine Chebulae Fructus has potential efficacy in the treatment of HCC. MATERIALS AND METHODS: We mined the active ingredients of Chebulae Fructus and its main targets from the Traditional Chinese Medicine Systems Pharmacology database. HCC-related datasets were downloaded from The Cancer Genome Atlas database and differentially expressed genes (DEGs) in HCC were obtained by differential expression analysis. Top10 small molecule compounds capable of reversing HCC pathology were screened by the Connectivity Map database based on DEGs. Ellipticine, an extract of Chebulae Fructus, had the potential to reverse HCC pathology. Protein-Protein Interaction (PPI) networks of DEGs in HCC were constructed using STRING. Eighteen potential targets of Chebulae Fructus for the treatment of HCC were obtained by taking intersection of DEGs in HCC with targets corresponding to the active constituents of Chebulae Fructus. In addition, MTT assay was also employed to examine the effect of ellipticine on HCC cell viability. RESULTS: It has been shown that ellipticine and ellagic acid have antitumor activity. Random Walk with Restart analysis of PPI networks was performed using potential targets as seeds, and the genes with the top 50 affinity coefficients were selected to construct a drug-active constituent-gene interaction network. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses of key genes involved in the treatment of HCC with Chebulae Fructus demonstrated that these genes were mainly enriched in signaling pathways related to tumor metabolism such as cAMP signaling pathway and Ras signaling pathway. Finally, it was verified by MTT assay that proliferation of HCC cells could be remarkably hindered. CONCLUSIONS: We excavated ellipticine, a key active constituent of Chebulae Fructus, by network pharmacology, and elucidated the signaling pathways involved in Chebulae Fructus, providing a theoretical basis for the use of Chebulae Fructus for HCC clinical application.

Regioselection Switch in Nucleophilic Addition to Isoquinolinequinones: Mechanism and Origin of the Regioselectivity in the Total Synthesis of Ellipticine.

Ellipticine was synthesized in six steps and 20% global yield starting from the readily available 2,5-dimethoxy isoquinoline. Unprecedented regioselective control of the nucleophilic attack on the isoquinoline-5,8-dione is first described. Investigation of the possible pathways of this transformation through density functional theory calculations reveals unexpected N-oxide assistance in cascade tautomerizations, which was crucial for directing the nucleophilic attack and hastening the overall process. Using this strategy, we prepared the aniline-isoquinolinedione adduct and submitted it to an intramolecular double C-H cross-coupling activation to furnish ellipticinequinone, which gave ellipticine after a MeLi addition/BH3 reduction sequence.

Synthesis and cytotoxicity evaluation of olivacine-indole hybrids tethered by alkyl linkers.

In this work, eleven new derivatives were prepared of the alkaloid olivacine (1), which was isolated from the bark of Aspidosperma australe. These compounds (7a-k) are hybrids of olivacine and indoles or carbazole, tethered by alkyl chains of variable lengths (C-4, C-5 or C-6). Compounds 7a-k showed increased cytotoxicity towards a panel of four cell lines. The subcellular localization of olivacine and of the synthetic derivatives was studied by fluorescence microscopy. The cycles of K562 cells exposed to olivacine or compounds 7a-k were analysed by flow cytometry, and showed, for some of the new derivatives, a different profile of cell distribution among the phases of the cycle when compared to olivacine, which is indicative of lysosomal apoptosis.

Anticancer and Immunomodulatory Activities of a Novel Water-Soluble Derivative of Ellipticine.

Cancer still remains a major public health concern around the world and the search for new potential antitumor molecules is essential for fighting the disease. This study evaluated the anticancer and immunomodulatory potential of the newly synthetized ellipticine derivate: sodium bromo-5,11-dimethyl-6H-pyrido[4,3-b]carbazole-7-sulfonate (Br-Ell-SO3Na). It was prepared by the chlorosulfonation of 9-bromoellipticine. The ellipticine-7-sulfonic acid itself is not soluble, but its saponification with sodium hydroxide afforded a water-soluble sodium salt. The cytotoxicity of Br-Ell-SO3Na was tested against cancerous (K562 cell line) and non-cancerous cells (Vero cell line and human peripheral blood mononuclear cells (PBMC)) using a Methylthiazoletetrazolium (MTT) assay. Cell cycle arrest was assessed by flow cytometry and the immunomodulatory activity was analyzed through an enzyme-linked immunosorbent assay (ELISA). The results showed that the Br-Ell-SO3Na molecule has specific anticancer activity (IC50 = 35 µM) against the K562 cell line, once no cytotoxicity effect was verified against non-cancerous cells. Cell cycle analysis demonstrated that K562 cells treated with Br-Ell-SO3Na were arrested in the phase S. Moreover, the production of IL-6 increased and the expression of IL-8 was inhibited in the human PBMC treated with Br-Ell-SO3Na. The results demonstrated that Br-Ell-SO3Na is a promising anticancer molecule attested by its noteworthy activity against the K562 tumor cell line and immunomodulatory activity in human PBMC cells.

Modeling the Antileukemia Activity of Ellipticine-Related Compounds: QSAR and Molecular Docking Study.

The antileukemia cancer activity of organic compounds analogous to ellipticine representes a critical endpoint in the understanding of this dramatic disease. A molecular modeling simulation on a dataset of 23 compounds, all of which comply with Lipinski's rules and have a structure analogous to ellipticine, was performed using the quantitative structure activity relationship (QSAR) technique, followed by a detailed docking study on three different proteins significantly involved in this disease (PDB IDs: SYK, PI3K and BTK). As a result, a model with only four descriptors (HOMO, softness, AC1RABAMBID, and TS1KFABMID) was found to be robust enough for prediction of the antileukemia activity of the compounds studied in this work, with an R2 of 0.899 and Q2 of 0.730. A favorable interaction between the compounds and their target proteins was found in all cases; in particular, compounds 9 and 22 showed high activity and binding free energy values of around -10 kcal/mol. Theses compounds were evaluated in detail based on their molecular structure, and some modifications are suggested herein to enhance their biological activity. In particular, compounds 22_1, 22_2, 9_1, and 9_2 are indicated as possible new, potent ellipticine derivatives to be synthesized and biologically tested.

Synthesis and antiproliferative evaluation of new isomeric ellipticine quinones / Síntesis y evaluación de la actividad antiproliferativa de nuevas elipticinquinonas isomericas

The synthesis of new isomeric ellipticine quinones 3a-c and their in vitro antiproliferative activities on cancer cell lines is reported. The designed N-heterocyclic quinones 3a-c were synthesized through a three step sequence which involves: a) one-pot preparation of 4-methoxycarbonyl-3,4-dimethylisoquinoline-5,8-quinone 1 from 2,5-dihydroxyacetophenone, methyl aminocrotonate and silver (II) oxide; b) regioselective amination of 1 with arylamines to give aminoquinones 2a-c and c) palladium-catalyzed intramolecular oxidative coupling of 7-aminoisoquinoline-5,8-quinones 2a-c. The in vitro antiproliferative activity of the new angular quinones was evaluated againts one normal cell line (lung fibroblasts) and gastric, lung and bladder cancer cell lines in 72-h drug exposure assays. The new compounds displayed similar or higher antiproliferative activity with respect to their quinone precursors 2a-c. The isomeric ellipticine quinone 2b appears as the more active member on bladder cancer cell line (IC50: 2.4 uM), comparable to etoposide used as anticancer reference drug...

Se describe la síntesis de las nuevas quinonas 3a-c, isoméricas de elipticina, y sus actividades antiproliferativas in vitro en líneas de células de cáncer. Las quinonas N-heterocíclicas 3a-c se sintetizaron a través de una secuencia que involucra: a) preparación de 4- metoxicarbonil-3,4-dimetlisoquinolin-5,8-quinone 1 a partir de 2,5-dihidroxiacetofenona, aminocrotonato de metilo y óxido de plata (I); b) aminación regioselectiva de 1 con arilaminas para producir las aminoquinonas 2a-c y c) acoplamiento oxidante intramolecular de 7- aminoisoquinolin-5,8-quinonas 2a-c catalizado con paladio. La actividad antiproliferative in vitro de los nuevos compuestos fue evaluada en una línea celular normal (fibroblastos de pulmón) y líneas de células de cáncer gástrico, pulmón y vejiga en ensayos de exposición de 72 horas a la droga. Las quinonas 3a-c exhiben interesantes propiedades antiproliferativas destacando la elipticinquinona isomérica 2b en células de cáncer de vejiga (IC50: 2.4 uM) comparado con etopósido usada como droga anticancer de referencia. Los nuevos compuestos mostraron actividades antiproliferativa similar o mayor respecto de las correspondientes quinonas precursoras 2a-c. La elipticin quinona isomérica 2b corresponde al miembro más activo en células de câncer de vejiga (IC50: 2.4 uM), comparable a la del etopósido, usada como droga anticáncer de referencia...

Antiplasmodial activity of synthetic ellipticine derivatives and an isolated analog.

Ellipticine has been shown previously to exhibit excellent in vitro antiplasmodial activity and in vivo antimalarial properties that are comparable to those of the control drug chloroquine in a mouse malaria model. Ellipticine derivatives and analogs exhibit antimalarial potential however only a few have been studied to date. Herein, ellipticine and a structural analog were isolated from Aspidosperma vargasii bark. A-ring brominated and nitrated ellipticine derivatives exhibit good in vitro inhibition of Plasmodium falciparum K1 and 3D7 strains. Several of the compounds were found not to be toxic to human fetal lung fibroblasts. 9-Nitroellipticine (IC50=0.55µM) exhibits greater antiplasmodial activity than ellipticine. These results are further evidence of the antimalarial potential of ellipticine derivatives.

Comparative in vitro and in vivo antimalarial activity of the indole alkaloids ellipticine, olivacine, cryptolepine and a synthetic cryptolepine analog.

Indole alkaloids ellipticine (1), cryptolepine triflate (2a), rationally designed 11-(4-piperidinamino)cryptolepine hydrogen dichloride (2b) and olivacine (3) (an isomer of 1) were evaluated in vitro against Plasmodium falciparum and in vivo in Plasmodium berghei-infected mice. 1-3 inhibited P. falciparum (IC50≤1.4 µM, order of activity: 2b>1>2a>3). In vitro toxicity to murine macrophages was evaluated and revealed selectivity indices (SI) of 10-12 for 2a and SI>2.8×10² for 1, 2b and 3. 1 administered orally at 50mg/kg/day was highly active against P. berghei (in vivo inhibition compared to untreated control (IVI)=100%, mean survival time (MST)>40 days, comparable activity to chloroquine control). 1 administered orally and subcutaneously was active at 10 mg/kg/day (IVI=70-77%; MST=27-29 days). 3 exhibited high oral activity at ≥50 mg/kg/day (IVI=90-97%, MST=23-27 days). Cryptolepine (2a) administered orally and subcutaneously exhibited moderate activity at 50mg/kg/day (IVI=43-63%, MST=24-25 days). At 50 mg/kg/day, 2b administered subcutaneously was lethal to infected mice (MST=3 days) and moderately active when administered orally (IVI=45-55%, MST=25 days). 1 and 3 are promising compounds for development of antimalarials.

Docking studies on DNA-ligand interactions: building and application of a protocol to identify the binding mode.

Despite DNA being an important target for several drugs, most of the docking programs are validated only for proteins and their ligands. In this paper, we used AutoDock 4.0 to perform self-dockings and cross dockings between two DNA ligands (a minor groove binder and an intercalator) and four distinct receptors: 1) crystallographic DNA without intercalation gap; 2) crystallographic DNA with intercalation gap; 3) canonical B-DNA; and 4) modified B-DNA with intercalation gap. Besides being efficient in self-dockings, AutoDock is capable of correctly identifying two of the main DNA binding modes with the condition that the target possesses an artificial intercalation gap. Therefore, we suggest a default protocol to identify DNA binding modes which uses a modified canonical DNA (with gap) as receptor. This protocol was applied to dock two different Troger bases to DNA and the predicted binding modes agree with those suggested, yet not established, by experimental data. We also applied the protocol to dock aflatoxin B(1) exo-8,9-epoxide, and the results are in complete agreement with experimental data from the literature. We propose that this approach can be used to investigate other ligands whose binding mode to DNA remains unknown, yielding a suitable starting point for further theoretical studies such as molecular dynamics simulations.

Pattern recognition methods investigation of ellipticines structure-activity relationships.

Ellipticine is a molecule derived from the natural extract Ochrosia elliptica. This molecule and its derivatives are highly cytotoxic to malignant cultured cells. The relatively simple structure of ellipticine has prompted chemists to design various structural modifications in order to obtain either more active derivatives or information on the structural moieties required for pharmacological activities. In the present work we report theoretical structure-activity relationship studies for 40 ellipticine derivatives using pattern-recognition methods such as electronics indices methodology (EIM), principal component analysis (PCA) and hierarchical clustering analysis (HCA) with molecular descriptors obtained from semiempirical parametric method 3 (PM3) calculations. By applying selected molecular descriptors it was possible to classify active and inactive compounds with accuracy up to 92% and also to suggest the activity of new untested molecules. These descriptors have been only recently discussed in the literature as new possible universal parameters for defining the biological activity of several classes of compounds.

Interaction effects of 5-azacytidine with topoisomerase II inhibitors on CHO cells, as detected by cytogenetic analysis.

Different cell treatment protocols with the hypomethylating agent 5 azacytidine (5-aza C) were used in exponentially growing Chinese hamster ovary (CHO) cells in order to test its influence on the induction of chromosomal aberrations (CAs) induced by topoisomerase II inhibitors, ellipticine (EPC) and teniposide (VM-26). Cells pre-treated with 1 microg/ml 5-aza C for 1 h during the S-phase and post-treated in the last 2 h of incubation with 0.6 microg/ml EPC or 0.04 microg/ml VM-26 showed a reduction of 48% and 45%, respectively, in the frequencies of CAs as compared to the sum value of the frequencies obtained for each drug alone. 5-aza C added to the cultures for the last 2 h before cell fixation after a 30-min pulse treatment with EPC or VM-26 caused a 38% and 28% reduction, respectively. Simultaneous treatments with 5-aza C plus EPC, or 5-aza C plus VM-26 during the last 2 h of incubation (G2-phase), showed a significant effect of CA reduction (24%) only for the combination of 5-aza C + EPC. Preliminary assays with 5-aza C alone added to the cultures at different times demonstrated its effectiveness in inducing chromosome damage during the S-phase. Since S-phase-treated CHO cells showed a higher degree of reduction in the frequencies of CAs induced by EPC and VM-26, we suggest that 5-aza C incorporation into DNA may change the topo II cleavage sites, protecting the DNA from the induction of damage, or that the hypomethylation induced by incorporation of 5-aza C into DNA may change the chromatin structure facilitating the access to DNA repair enzymes. An alternative possibility is that 5-azaC can reactivate methylated genes involved in the repair of DNA double-strand breaks induced by topo II inhibitors.

Analogues and prodrugs, permanent useful alternatives to improve the effectiveness of natural anticancer compounds: Enhanced activity of slightly soluble salts of olivacine in L1210 leukemia

Several derivatives of well-known natural anticancer drugs have been synthesized and will continue to be, to overcome their delivering problems, or effectiveness. Characteristics such as poor solubility, metabolic inactivation, resistance development, rapid clearance or short half-life, adverse side effects like nausea, alopecia, anorexia are common undesirable limitations to their clinical use. Drug toxicity to less sensitive tumors as those in liver or brain is another aimed goal. Recent efforts to reverse undesirable physicochemical properties of well-established natural anticancer compounds are focused. Improved antitumor action of olivacine in L(1210) leukemia with slightly soluble halides with increased life span as well as "cured" animals was observed.

Clastogenic action of ellipticine over the cell cycle of human lymphocytes and influence of posttreatments with caffeine and ara-C at G2.

The clastogenic potential of the intercalating compound ellipticine, an antitumor alkaloid, has been demonstrated in mammalian cells. To characterize the mechanism of action of this drug over the cell cycle, human lymphocyte cultures from 2 healthy donors were treated with 3 micrograms/ml ellipticine in 30-min pulses during different phases of the cell cycle and analyzed for chromosomal aberrations and sister-chromatid exchanges. The G2 phase was most sensitive in terms of induction of aberrations, followed by S and G1. Chromatid-type aberrations were the most common type of chromosomal damage. Induction of SCEs was significantly high only after treatment at G1, when the frequencies of SCEs doubled. The post-treatment effect of lymphocytes with inhibitors of DNA repair, 10(-3) M caffeine and 5 x 10(-6) M 1-beta-D-arabinofuranosylcytosine, was also tested by adding 3 micrograms/ml ellipticine at G2 in 30-min pulses and immediately followed by caffeine and/or ara-C during the last 3 h before harvesting. Three experiments performed on blood from 3 donors showed a moderate potentiation effect on the frequency of chromatid-type aberrations (about 2-3 times) by both inhibitors. Likewise, a 3-fold increase was observed in the frequencies of chromosomal aberrations when caffeine and ara-C were combined. The present data demonstrate that posttreatment with caffeine and ara-C at G2 can modify the response of human lymphocytes treated with ellipticine by increasing the clastogenic action of this compound or by changing the cell-cycle progression.

Clastogenic effect of the plant alkaloid ellipticine on bone marrow cells of Wistar rats and on human peripheral blood lymphocytes.

Ellipticine (EPC), a natural alkaloid extracted from Aspidosperma williansii (Apocynaceae), is known to have antitumor and cytotoxic activities on various types of tumors. This drug showed a strong clastogenic effect on bone marrow cells of Wistar rats treated in vivo (7.75-31.00 mg/kg body weight). EPC was also tested in vitro using the human peripheral blood lymphocyte system, at concentrations 100 times lower than those used in the in vivo test on rats, since the cytotoxic effect on lymphocytes was very strong. At the 2 highest concentrations used (7.75 X 10(-1) and 1.55 X 10(-1) micrograms/ml culture medium), EPC induced a statistically significant increase in the frequency of chromosome aberrations and sister-chromatid exchanges in lymphocytes. Based on data reported in the literature, we have tried to establish relationships between the clastogenic effect observed and the process of EPC intercalation into DNA and the formation of protein-associated DNA-strand breaks probably promoted by topoisomerase enzymes.

[A new quaternary alkaloid from Aspidosperma gilbertii]. / Um novo alcalóide quaternário de Aspidosperma gilbertii.

The present work describes the structure determination of a new quaternary carbozol type alkaloid, isolated from Aspidosperma gilbertii: N,N-Dimethyltetrahydroellipticinium Hydroxyde.