Study of nitrogen heterocycles as DNA/HSA binder, topoisomerase inhibitors and toxicological safety.

Four new nitrogen-containing heterocyclic derivatives (acridine, quinoline, indole, pyridine) were synthesized and their biological properties were evaluated. The compounds showed affinity for DNA and HSA, with CAIC and CAAC displaying higher binding constants (Kb) of 9.54 × 104 and 1.06 × 106, respectively. The fluorescence quenching assay (Ksv) revealed suppression values ranging from 0.34 to 0.64 × 103 M-1 for ethidium bromide (EB) and 0.1 to 0.34 × 103 M-1 for acridine orange (AO). Molecular docking confirmed the competition of the derivatives with intercalation probes at the same binding site. At 10 µM concentrations, the derivatives inhibited topoisomerase IIα activity. In the antiproliferative assays, the compounds demonstrated activity against MCF-7 and T47-D tumor cells and nonhemolytic profile. Regarding toxicity, no acute effects were observed in the embryos. However, some compounds caused enzymatic and cardiac changes, particularly the CAIC, which increased SOD activity and altered heart rate compared to the control. These findings suggest potential antitumor action of the derivatives and indicate that substituting the acridine core with different cores does not interfere with their interaction and topoisomerase inhibition. Further investigations are required to assess possible toxicological effects, including reactive oxygen species generation.

Biochemical and teratogenic effects of a mixture of pyriproxyfen and glyphosate.

The mixture of agrochemicals can be made to improve pest control or accidentally. In this way, the effects on non-target organisms are a critical aspect of the environment and heath. Thus, this work aimed to show how a mixture of pyriproxyfen, and glyphosate can impair biochemical routes and embryonic development. Zebrafish embryos 0-72 hpf were exposed to 0.001-1 µg/mL of pyriproxyfen, glyphosate, and a mixture of both pesticides. The ADMETox was evaluated in silico. The FET-test was used to estimate teratogenic effects. The biochemical effects were estimated using AChE, SOD, and CAT as parameters. ROS generation was estimated using 30 µM H2DCF-DA and 5 µM DHE. The ADMETox reveals that intestinal absorption and P-glycoprotein are the main sites for PPx and Gly adsorption. The distribution parameters were diverse. PPx + Gly at 0.1 µg/mL leads to 50 % of lethality and at 1 µg/mL 100 % of lethality. PPx + Gly leads to a 22 % of lack of somite formation at 1 µg/mL. The heart rate was reduced by >10 % in all concentrations tested. The AChE has a decrease with IC20 19.6 µM and IC50 261.5 µM. SOD showed a reduction of 28 % to PPx and CAT was reduced by 58 % to PPx + Gly and Gly at 1 µg/mL. Glyphosate does not increase unspecific ROS generation. The superoxide generation was 2× higher in the PPx + Gly at 1 µg/mL. Summarily, was observed that the mixture of PPx + Gly potentiated the toxic effects. This finding suggests a possible synergism between the PPx and Gly even at lower concentrations.

ACW-02 an Acridine Triazolidine Derivative Presents Antileishmanial Activity Mediated by DNA Interaction and Immunomodulation.

The present study proposed the synthesis of a novel acridine derivative not yet described in the literature, chemical characterization by NMR, MS, and IR, followed by investigations of its antileishmanial potential. In vitro assays were performed to assess its antileishmanial activity against L. amazonensis strains and cytotoxicity against macrophages through MTT assay and annexin V-FITC/PI, and the ability to perform an immunomodulatory action using CBA. To investigate possible molecular targets, its interaction with DNA in vitro and in silico targets were evaluated. As results, the compound showed good antileishmanial activity, with IC50 of 6.57 (amastigotes) and 94.97 (promastigotes) µg mL-1, associated with non-cytotoxicity to macrophages (CC50 > 256.00 µg mL-1). When assessed by flow cytometry, 99.8% of macrophages remained viable. The compound induced an antileishmanial effect in infected macrophages and altered TNF-α, IL-10 and IL-6 expression, suggesting a slight immunomodulatory activity. DNA assay showed an interaction with the minor grooves due to the hyperchromic effect of 47.53% and Kb 1.17 × 106 M-1, and was sustained by docking studies. Molecular dynamics simulations and MM-PBSA calculations propose cysteine protease B as a possible target. Therefore, this study demonstrates that the new compound is a promising molecule and contributes as a model for future works.

A simple method for obtaining human albumin and its use for in vitro interaction assays with indole-thiazole and indole-thiazolidinone derivatives.

This work aimed to develop a simple and low-cost method to obtain human serum albumin (HSA) and its consequent application for in vitro drug interaction assays. The HSA was purified by classic principles of plasma precipitation and thermocoagulation, using a multiple-stage fractionation. The quality of the final product was assessed by electrophoresis, protein dosage by the Lowry method and the pharmacopeial thermal stability. At the end, an isotonic solution of HSA with a total protein concentration of 2.7 mg·mL-1 was obtained, which was visualized as a single band corresponding to the molecular weight of 66 kDa. After the thermal stability test, there was no indication of turbidity or color change of the solution. Finally, the HSA was useful for interaction assays with indole-thiazole and indole-thiazolidinone derivatives through UV-vis absorption and fluorescence spectroscopic studies, as well as by docking molecular analysis. Derivatives quenched the intrinsic fluorescence of HSA, disrupted the tryptophan residues microenvironment, and probably bind at Sudlow's site I. Therefore, the simplified methodology developed in this work proved to be effective in obtaining HSA that can be applied to research goals including drug interaction assays.

Novel indole-thiazole and indole-thiazolidinone derivatives as DNA groove binders.

In this study, we report the synthesis of eight novel indole-thiazole and indole-thiazolidinone derivatives, as well as their ability to interact with DNA, analysed through the UV-vis absorption, fluorescence, circular dichroism (CD), viscosity techniques and molecular docking. The ctDNA interaction analysis demonstrated different spectroscopic effects and the affinity constants (Kb) calculated by the UV-vis absorption method were between 2.08 × 105 and 6.99 × 106 M-1, whereas in the fluorescence suppression constants (Ksv) ranged between 0.38 and 0.77 × 104 M-1 and 0.60-7.59 × 104 M-1 using Ethidium Bromide (EB) and 4',6-Diamidino-2-phenylindole (DAPI) as fluorescent probes, respectively. Most derivatives did not alter significantly the secondary structure of the ctDNA according to the CD results. None of the compounds was able to change the relative viscosity of the ctDNA. These results prove that compounds interact with ctDNA via groove binding, which was confirmed by A-T rich oligonucleotide sequence assay with compound JF-252, suggesting the importance of both the phenyl ring coupled to C-4 thiazole ring and the bromo-unsubstituted indole nucleus.

COVID-19 therapy: What weapons do we bring into battle?

Urgent treatments, in any modality, to fight SARS-CoV-2 infections are desired by society in general, by health professionals, by Estate-leaders and, mainly, by the scientific community, because one thing is certain amidst the numerous uncertainties regarding COVID-19: knowledge is the means to discover or to produce an effective treatment against this global disease. Scientists from several areas in the world are still committed to this mission, as shown by the accelerated scientific production in the first half of 2020 with over 25,000 published articles related to the new coronavirus. Three great lines of publications related to COVID-19 were identified for building this article: The first refers to knowledge production concerning the virus and pathophysiology of COVID-19; the second regards efforts to produce vaccines against SARS-CoV-2 at a speed without precedent in the history of science; the third comprehends the attempts to find a marketed drug that can be used to treat COVID-19 by drug repurposing. In this review, the drugs that have been repurposed so far are grouped according to their chemical class. Their structures will be presented to provide better understanding of their structural similarities and possible correlations with mechanisms of actions. This can help identifying anti-SARS-CoV-2 promising therapeutic agents.

Topoisomerase inhibition and albumin interaction studies of acridine-thiosemicarbazone derivatives.

In the present study, acridine-thiosemicarbazones (ATD) derivatives were tested for their interaction properties with BSA through UV-Vis absorption and fluorescence spectroscopic studies. Both hyperchromic and hypochromic effects, as well as red or blue shifts were demonstrated after the derivatives were added to the BSA. Values for the binding constant (Kb) ranged from 1.62 × 104 to 8.71 × 105 M-1 and quenching constant (KSV) from 3.46 × 102 to 7.83 × 103 M-1 indicating a good affinity to BSA protein. Complementary, two compounds were selected to assess their inhibition activity against topoisomerase IIα enzyme, of which derivative 3a presented the best result. Moreover, to evaluate protein-ligand interactions, as well as the antitopoisomerase potential of these compounds, tests of molecular modeling were performed between all compounds using the albumin and Topoisomerase IIα/DNA complex. Finally, in silico studies showed that all derivatives used in this research displayed good oral bioavailability potential.

5-Nitro-Thiophene-Thiosemicarbazone Derivatives Present Antitumor Activity Mediated by Apoptosis and DNA Intercalation.

BACKGROUND: Considering the need for the development of new antitumor drugs, associated with the great antitumor potential of thiophene and thiosemicarbazonic derivatives, in this work we promote molecular hybridization approach to synthesize new compounds with increased anticancer activity. OBJECTIVE: Investigate the antitumor activity and their likely mechanisms of action of a series of N-substituted 2-(5-nitro-thiophene)-thiosemicarbazone derivatives. METHODS: Methods were performed in vitro (cytotoxicity, cell cycle progression, morphological analysis, mitochondrial membrane potential evaluation and topoisomerase assay), spectroscopic (DNA interaction studies), and in silico studies (docking and molecular modelling). RESULTS: Most of the compounds presented significant inhibitory activity; the NCIH-292 cell line was the most resistant, and the HL-60 cell line was the most sensitive. The most promising compound was LNN-05 with IC50 values ranging from 0.5 to 1.9 µg.mL-1. The in vitro studies revealed that LNN-05 was able to depolarize (dose-dependently) the mitochondrial membrane, induceG1 phase cell cycle arrest noticeably, promote morphological cell changes associated with apoptosis in chronic human myelocytic leukaemia (K-562) cells, and presented no topoisomerase II inhibition. Spectroscopic UV-vis and molecular fluorescence studies showed that LNN compounds interact with ctDNA forming supramolecular complexes. Intercalation between nitrogenous bases was revealed through KI quenching and competitive ethidium bromide assays. Docking and Molecular Dynamics suggested that 5-nitro-thiophene-thiosemicarbazone compounds interact against the larger DNA groove, and corroborating the spectroscopic results, may assume an intercalating interaction mode. CONCLUSION: Our findings highlight 5-nitro-thiophene-thiosemicarbazone derivatives, especially LNN-05, as a promising new class of compounds for further studies to provide new anticancer therapies.

Spiro-acridine inhibiting tyrosinase enzyme: Kinetic, protein-ligand interaction and molecular docking studies.

Here, we evaluate spiroacridines as inhibitors of tyrosinase, a key enzyme to melanogenesis. For this purpose, the spiroacridines 3-(acridin-9-yl)-N-benzylidene-2-cyanoacrylohydrazide (AMTAC-01) and 3-(acridin-9-yl)-2-cyano-N-(4-metoxybenzylidene)-acrylohydrazide (AMTAC-02) were synthesized and their enzymatic inhibition types and mechanisms were investigated. In addition, the interaction of these compounds with the enzyme were studied by UV-Vis spectroscopy, spectrofluorimetry, 1H NMR titration as well as molecular docking. Spectroscopic results reveals that the acridine derivatives interact strongly (Ka ≅ 104 - 105 M-1) with the mushroom tyrosinase and the enzyme undergoes small structural modifications due to the interaction with AMTAC-01 compound. The interaction studies support the enzymatic inhibition results, which suggests that AMTAC-01 compounds inhibit the enzyme reversibly and follows a noncompetitive type (AMTAC-01) and mixed type (AMTAC-02) of inhibition. Nevertheless, AMTAC-02 (IC50 = 96.29 µM) inhibits the enzyme more effectively than AMTAC-01 (IC50 = 189.40 µM), which suggests a highly relevant role of AMTAC-02's methoxy group to the inhibition activity, which is confirmed by docking studies to mushroom tyrosinase. Docking also indicates this interaction to be absent in human tyrosinase. SIGNIFICANCE: Based on previous results which evidenced the relevant activity of two spiroacridinic compounds for cell growth inhibition against melanoma cells, here we improve our understanding about the spiroacridines in the biological media by exploring the molecular mechanism that govern the activities of these two compounds using mushroom tyrosinase (mTYR) enzyme as molecular target. The paper not only will have a major impact upon molecular mechanism that regulates melanin inhibition by spiroacridinic compounds, but also by guiding the search for enzyme inhibitors and the development of new anti-melanoma prophylaxis.

Synthesis, DNA and protein interactions and human topoisomerase inhibition of novel Spiroacridine derivatives.

Nine new spiroacridine derivatives were synthetized by introducing cyano-N-acylhydrazone group between the acridine and phenyl-substituted rings followed by spontaneous cyclization. The new compounds were assayed for their DNA binding properties, human topoisomerase IIα inhibition and bovine serum albumin (BSA) interaction. Besides, docking analysis were performed in order to better understanding the biomolecule-compounds interactions. All compounds interacted with BSA which was demonstrated by the fluorescence suppression constant of 104 M-1. Compounds with chloro and NO2 substituents at that para-position on phenyl ring demonstrated the best results for BSA interaction. DNA binding constant determined by UV-vis data demonstrated high values for AMTAC-11 and AMTAC-14, 1.1 × 108 M-1 and 4.8 × 106 M-1, respectively, and all others presented constant values of 105 M-1. AMTAC-06 with chloro at para-position on phenyl ring presented a topoisomerase II inhibition of 84.34% in comparison to the positive controls used. Docking studies indicated that AMTAC-06 is able to intercalate the DNA base pairs at topoisomerase IIα active site, preventing DNA connection after break, in a process known as poisoning. Topoisomerase enzyme inhibition result was correlated to BSA interaction profile, since AMTAC-06 showed the best results in both analysis. The findings obtained here proved that methoxy or chloro substitution on phenyl ring at para-position is fundamental for in vitro activity of new spiroacridine derivatives, and indicates that AMTAC-06 is a promising entity and should serve as a lead compound in the development of new DNA and protein binders, as well as human topoisomerase II inhibitors.

Molecular profile of mannan-binding lectin in hepatitis C patients with MBL gene polymorphisms by a modified mannan-coated nitrocellulose assay.

The aim of this study was to develop an assay to analyze the serum profile of Mannose-binding lectin (MBL) through a simple and "in-house" method (called "dot-N-man"). Furthermore, the study attempted to associate molecular masses of MBL to the profile of MBL gene polymorphisms in patients with hepatitis C. Heterogeneity in molecular masses of MBL is due to the impairment of oligomers formation, which is linked to genetic polymorphisms in the MBL gene. Individuals with AA genotype (wild-type) produce high-molecular-mass proteins, whereas AO and OO individuals produce intermediate and low-molecular-mass proteins, respectively. Sera of thirty patients carrying the hepatitis C virus (HCV) were investigated using MBL binding assay with mannan-coated nitrocellulose (dot-N-man). Purified MBL was evaluated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. Dot-N-Man assay yielded MBL with molecular masses ranging between 55 and 320 kDa, comparable to low and high molecular mass forms of MBL. Nonreducing SDS-PAGE showed high molecular mass bands in all AA individuals while bands of 270 and 205 kDa were observed in sera for a number of patients with AO and OO genotypes, respectively. Immunoblotting confirmed the MBL samples obtained from the dot-N-man. These results provide new insights to understand the MBL molecular forms profile in patients infected with HCV- which could be useful in future investigations on the influence of the MBL structure/genotype on both the progression of infection and the response to hepatitis C therapy.

Lectin-carbohydrate complex evaluation by chemiluminescence.

In order to characterize the affinity between specific carbohydrate-binding proteins such as lectins, a model is proposed to study these interactions using a polysaccharide membrane to simulate such adsorption. Here, lectin-carbohydrate interactions were chemiluminescently investigated using lectins conjugated to acridinium ester (AE) and polysaccharides composed of their respective specific carbohydrates. The lectin-AE conjugates were incubated with discs (0.0314-0.6358 cm2) of phytagel, chitosan and carrageenan. The complex formation chemiluminescently detected followed the Langmuir isotherm from which constants were estimated. The association constant (Ka) and maximum binding sites on the membranes were 2.4 × 10-7 M-1 ±â€¯0.8 × 10-7 M-1 and 1.3 × 10-3 mol. mg-1 ± 0.3 × 10-3 mol. mg-1 (Con A); 0.9 × 10-6 M-1 ±â€¯0.4 × 10-6 M-1 and 0.021 × 10-3 mol. mg-1 ± 0.003 × 10-3 mol. mg-1 (WGA) and 2.0 × 10-6 M-1 ±â€¯0.9 × 10-6 M-1 and 0.069 × 10-3 mol. mg-1 ± 0.010 × 10-3 mol. mg-1 (PNA). The proposed model might be useful to study binding affinity and estimate the amount of binding not limited by the sugar content in the membrane.

New thiophene-acridine compounds: Synthesis, antileishmanial activity, DNA binding, chemometric, and molecular docking studies.

In this study, we synthesized eight new compounds containing the 2-amino-cycloalkyl[b]thiophene and acridine moieties (ACT01 and ACS01 -ACS07 ). None tested compounds presented human erythrocyte cytotoxicity. The new compounds presented antipromastigote activity, where ACS01 and ACS02 derivatives presented significant antileishmanial activity, with better performance than the reference drugs (tri and pentavalent antimonials), with respective IC50 values of 9.60 ± 3.19 and 10.95 ± 3.96 µm. Additionally, these two derivatives were effective against antimony-resistant Leishmania (Leishmania) amazonensis strains. In addition, binding and fragmentation DNA assays were performed. It was observed that the antileishmanial activity of ACS01 is not associated with DNA fragmentation of the promastigote forms. However, it interacted with DNA with a binding constant of 104  m-1 . In partial least-squares studies, it was observed that the most active compounds (ACS01 and ACS02 ) showed lower values of amphiphilic moment descriptor, but there was a correlation between the lipophilicity of the molecules and antileishmanial activity. Furthermore, the docking molecular studies showed interactions between thiophene-acridine derivatives and the active site of pyruvate kinase enzyme with the major contribution of asparagine 152 residue for the interaction with thiophene moiety. Thus, the results suggested that the new thiophene-acridine derivatives are promising molecules as potential drug candidates.

DNA binding and Topoisomerase inhibition: How can these mechanisms be explored to design more specific anticancer agents?

DNA is considered one of the most promising targets of molecules with anticancer activity potential. Its key role in various cell division mechanisms, which commands the intense multiplication of tumor cells, is considered in studies with compounds whose mechanisms of action suggest likeliness of interaction. In addition, inhibition of enzymes that actively participate in biological functions of cells such as Topoisomerase, is seen as a primary factor for conducting several events that result in cell death. Discovery of new anticancer chemotherapeutical capable of interacting with DNA and inhibiting Topoisomerase enzymes is highlighted in anticancer research. The present review aims at showing through distinct biological tests the performance of different candidates to anticancer drugs and their respective chemical modifications, which are crucial and/or determinant for DNA affinity and inhibition of important enzymes in cells' vital processe to either separately or synergistically optimize anticancer activity.

Thiosemicarbazones and 4-thiazolidinones indole-based derivatives: Synthesis, evaluation of antiproliferative activity, cell death mechanisms and topoisomerase inhibition assay.

In this study, we report the synthesis and structural characterization of a series of thiosemicarbazone and 4-thiazolidinones derivatives, as well as their in vitro antiproliferative activity against eight human tumor cell lines. For the most potent compound further studies were performed evaluating cell death induction, cell cycle profile, ctDNA interaction and topoisomerase IIα inhibition. A synthetic three-step route was established for compounds (2a-e and 3a-d) with yields ranging from 32 to 95%. Regarding antiproliferative activity, compounds 2a-e and 3a-d showed mean GI50 values ranging between 1.1 µM (2b) - 84.65 µM (3d). Compound 2b was the most promising especially against colorectal adenocarcinoma (HT-29) and leukemia (K562) cells (GI50 = 0.01 µM for both cell lines). Mechanism studies demonstrated that 24 h-treatment with compound 2b (5 µM) induced phosphatidylserine residues exposition and G2/M arrest on HT-29 cells. Moreover, 2b (50 µM) was able to interact with ctDNA and inhibited topoisomerase IIα activity. These results demonstrate the importance of thiosemicarbazone, especially the derivative 2b, as a promising candidate for anticancer therapy.

Synthesis of novel indole derivatives as promising DNA-binding agents and evaluation of antitumor and antitopoisomerase I activities.

Molecules bearing indole nucleus present diverse biological properties such as antitumor and anti-inflammatory activities that can be associated both to DNA and protein interactions. This study focused on the synthesis of new indole derivatives with thiazolidines and imidazolidine rings condensed as side chains as well as the evaluation of their ability to interact with the DNA and antitumor and topoisomerase inhibition activities. All derivatives were successfully synthesized and their structures were elucidated by mass spectrometry (MS), infrared (IR), spectroscopy 1H NMR, 13C NMR, COSY 1H-1H and HSQC 1H-13C. The antitumor activity was evaluated against different cancer cell lines using the antiproliferative MTT assay. DNA binding ability was analyzed by absorption spectroscopy and fluorescence technique using ethidium bromide (EB) as a fluorescent probe. Changes were observed in spectroscopic properties of the compounds after interacting with ctDNA (calf thymus DNA), with hypochromic and hyperchromic effects, besides blue or red shifts in the maxima of spectra. The indole derivative 5-(1H-Indol-3-ylmethylene)-thiazolidin-2,4-dione (4c) presented the best results in antitumor assay against the breast line tested (T47D), with IC50 value lower than the positive control, doxorubicin (1.93 and 4.61 µM, respectively). On the other hand, the compound 3-amino-5-(1H-indol-3-ylmethylene)-2-thioxo-thiazolidin-4-one (4a) was active against leukemia cell lines (HL60 and K562) with the high value of the DNA binding constant, Kb of 5.69 × 104. However, this compound (4a) did not inhibit the topoisomerase-I activity evaluated by relaxation assay. These results show that the indole nucleus contribute to the incorporation of molecules into the DNA. Moreover, it was highlighted that basic side chains, such as thiazolidines and imidazolidines, and free amino group, are relevant for design of promising antitumor and DNA binding compounds.

Ultrastructural Assessment of 2-(acridin-9-ylmethylene)-N-phenylhydrazinecarbothioamide activity on human breast adenocarcinoma cells.

The aim of the present study was to investigate ultrastructural changes induced by (Z)-2-(acridin-9-ylmethylene)-N-phenylhydrazinecarbothioamide (APHCA) treatment on human breast adenocarcinoma cancer cells MCF-7, besides the evaluation of phosphatidylserine externalization and DNA fragmentation in treated cells. Cell viability analysis demonstrated concentration and time-manner cytotoxicity. Treated MCF-7 cells did not expose phosphatidylserine residues to the external plasma membrane surface and DNA fragmentation was not visualized by electrophoresis. Light microscopy showed compromised cell density and presence of vacuolization after APHCA treatment with 60µM. Scanning and transmission electron microscopies revealed hallmarks of autophagy, namely the presence of membrane bebbling and autophagosomes, besides shrunken cells and cell debris in treated MCF-7 cells. However, more specific tests such as the quantification of mammalian autophagy proteins are necessary to determine the kind of death that is trigged by APHCA.

Synthesis, DNA Binding, and Antiproliferative Activity of Novel Acridine-Thiosemicarbazone Derivatives.

In this work, the acridine nucleus was used as a lead-compound for structural modification by adding different substituted thiosemicarbazide moieties. Eight new (Z)-2-(acridin-9-ylmethylene)-N-phenylhydrazinecarbothioamide derivatives (3a-h) were synthesized, their antiproliferative activities were evaluated, and DNA binding properties were performed with calf thymus DNA (ctDNA) by electronic absorption and fluorescence spectroscopies. Both hyperchromic and hypochromic effects, as well as red or blue shifts were demonstrated by addition of ctDNA to the derivatives. The calculated binding constants ranged from 1.74 × 10(4) to 1.0 × 10(6) M(-1) and quenching constants from -0.2 × 10(4) to 2.18 × 10(4) M(-1) indicating high affinity to ctDNA base pairs. The most efficient compound in binding to ctDNA in vitro was (Z)-2-(acridin-9-ylmethylene)-N- (4-chlorophenyl) hydrazinecarbothioamide (3f), while the most active compound in antiproliferative assay was (Z)-2-(acridin-9-ylmethylene)-N-phenylhydrazinecarbothioamide (3a). There was no correlation between DNA-binding and in vitro antiproliferative activity, but the results suggest that DNA binding can be involved in the biological activity mechanism. This study may guide the choice of the size and shape of the intercalating part of the ligand and the strategic selection of substituents that increase DNA-binding or antiproliferative properties.

Different cell death responses induced by eupomatenoid-5 in MCF-7 and 786-0 tumor cell lines.

Natural products remain an important source of new drugs, including anticancer drugs. Recently, our group reported the anticancer activity of eupomatenoid-5 (eup-5), a neolignan isolated from Piper regnellii (Miq.) C. DC. var. regnellii leaves. In vitro studies demonstrated that MCF-7 (breast) and 786-0 (kidney) were among the cancer cell lines most sensitive to eup-5 treatment. The current results demonstrate that mitochondrial membrane depolarization and generation of reactive oxygen species are implicated in eup-5-mediated cytotoxic effects on these cancer cells lines. In MCF-7 cells, eup-5 led to phosphatidylserine externalization and caspase activation, whereas the same did not occur in 786-0 cells. Scanning electron microscopy revealed a reduction of microvilli density, as well as cell morphology alterations. Moreover, treated MCF-7 cells exhibited well-characterized apoptosis alterations, while treated 786-0 cells exhibited characteristics of programmed necroptosis process. These findings support the possibility that different mechanisms may be targeted by eup-5 in cell death response.

Evaluation of glycophenotype in prostatic neoplasm by chemiluminescent assay.

This work aimed to evaluate the glycophenotype in normal prostate, bening prostatic hyperplasia (BPH) and prostatic adenocarcinoma (PCa) tissues by a chemiluminescent method. Concanavalin A (Con A), Ulex europaeus agglutinin (UEA-I) and Peanut agglutinin (PNA) lectins were conjugated to acridinium ester (lectins-AE). These conjugates remained capable to recognize their specific carbohydrates. Tissue samples were incubated with lectins-AE. The chemiluminescence of the tissue-lectin-AE complex was expressed in relative light units (RLU). Transformed tissues (0.25 cm(2) by 8 µm of thickness) showed statistical significant lower α-D-glucose/mannose (BPH: 226,931 ± 17,436; PCa: 239,520 ± 12,398) and Gal-ß(1-3)-GalNAc (BPH: 28,754 ± 2,157; PCa: 16,728 ± 1,204) expression than normal tissues (367,566 ± 48,550 and 409,289 ± 22,336, respectively). However, higher α-L-fucose expression was observed in PCa (251,118 ± 14,193) in relation to normal (200,979 ± 21,318) and BHP (169,758 ± 10,264) tissues. It was observed an expressive decreasing of the values of RLU by inhibition of the interaction between tissues and lectins-AE using their specific carbohydrates. The relationship between RLU and tissue area showed a linear correlation for all lectin-AE in both transformed tissues. These results indicated that the used method is an efficient tool for specific, sensitive and quantitative analyses of prostatic glycophenotype.