Biochemical disorders induced by cytotoxic marine natural products in breast cancer cells as revealed by proton NMR spectroscopy-based metabolomics

International audienceMarine plants and animals are sources of a huge number of pharmacologically active compounds, some of which exhibit antineoplastic activity of clinical relevance. However the mechanism of action of marine natural products (MNPs) is poorly understood. In this study, proton NMR spectroscopy-based metabolomics was applied to unravel biochemical disorders induced in human MCF7 breast cancer cells by 3 lead candidate anticancer MNPs: ascididemin (Asc), lamellarin-D (Lam-D), and kahalalide F (KF). Asc, Lam-D, and KF provoked a severe decrease in DNA content in MCF7 cells after 24h treatment. Asc and Lam-D provoked apoptosis, whereas KF induced non-apoptotic cell death. Metabolite profiling revealed major biochemical disorders following treatment. The response of MCF7 tumor cells to Asc involved the accumulation of citrate (×17 the control level, <0.001), testifying enzyme blockade in citrate metabolism, and the accumulation of gluconate (×9.8, <0.005), a metabolite never reported at such concentration in tumor cells, probably testifying glycolysis shutdown. The response to Lam-D involved the accumulation of aspartate (×7.2, <0.05), glutamate (×14.7, <0.05), and lactate (×2.3, <0.05), probably in relation with the targeting of the malate-aspartate shuttle, as discussed. The response to KF involved increased lipid accumulation (polyunsaturated fatty acids ×9.8, <0.05), and phospholipid and acetate derivative alterations. Altogether, this study demonstrates the potential of proton NMR spectroscopy-based metabolomics to help uncover metabolic targets and elucidate the mechanism of cytotoxicity of candidate antineoplastic MNPs

Role of neo-adjuvant hormonal therapy in the treatment of breast cancer: a review of clinical trials

The clinical benefits of endocrine therapy for patients with hormonosensitive breast cancer are well established. For many years, 5 years of tamoxifen was the gold standard of adjuvant treatment. The recent development of new endocrine agents provides physicians with a more effective therapeutic approach. Nevertheless, the success of neoadjuvant endocrine therapy is much more recent and less reported in the literature. This article reviews the studies published about neoadjuvant endocrine treatment (tamoxifen and aromatase inhibitors). According to the literature, neoadjuvant endocrine therapy seems to be effective. In contrast to neoadjuvant chemotherapy, neoadjuvant endocrine therapy is well tolerated, with very few patients having to discontinue the treatment because of side effects. It does not constitute a standard treatment but could have potential for elderly women with operable, hormonosensitive, well differentiated and slowly progressing (SBR I) tumor or for patients with lobular MSBR 1 carcinoma (low chemosensitivity). The newer generation of aromatase inhibitors (letrozole, anastrozole, exemestane) appears to be more active (in terms of overall response rates and conservative surgery rate) than tamoxifen. Patients with an estrogen receptor Allred score of 6 and over are more likely to respond and gain a clinical benefit. The optimal duration of neoadjuvant therapy has not yet been investigated in detail. These preliminary results should be confirmed by further studies

Les dioxines (revue des connaissances, diaporama)

Parmi les polluants organiques persistants ayant un impact planétaire et donc présentant un intérêt majeur en santé publique, les dioxines sont un exemple typique de composés toxiques susceptibles de contaminer tous les compartiments environnementaux et, en final, l'homme. Après avoir analysé les principales caractéristiques des dioxines, leur processus de formation ainsi que les différentes sources d'émission de ces polluants, cette thèse précise les modalités d'exposition humaine, le mécanisme de toxicité impliquant le récepteur Ah et les effets observés chez l'homme et chez l'animal suite à une exposition subaiguë ou chronique. Ce document examine également l'imprégnation des populations et les doses acceptables de référence comme les doses journalières admissibles (DJA) mises en place pour prévenir les risques pour la santé humaine, les données de l'expérimentation animale (toxicologie) et les résultats d'études épidémiologiques. La présentation sous forme de diaporama comme outil didactique a été choisie dans le but d'être utilisée comme support d'enseignement en Santé Environnementale.GRENOBLE1-BU Médecine pharm. (385162101) / SudocSudocFranceF

Metabolomics Reveals Metabolic Targets and Biphasic Responses in Breast Cancer Cells Treated by Curcumin Alone and in Association with Docetaxel

<div><p>Background</p><p>Curcumin (CUR) has deserved extensive research due to its anti-inflammatory properties, of interest in human diseases including cancer. However, pleiotropic even paradoxical responses of tumor cells have been reported, and the mechanisms of action of CUR remain uncompletely elucidated.</p> <p>Methodology/Principal Findings</p><p>¹H-NMR spectroscopy-based metabolomics was applied to get novel insight into responses of MCF7 and MDA-MB-231 breast cancer cells to CUR alone, and MCF7 cells to CUR in cotreatment with docetaxel (DTX). In both cell types, a major target of CUR was glutathione metabolism. Total glutathione (GSx) increased at low dose CUR (≤ 10 mg.l⁻¹–28 µM-) (up to +121% in MCF7 cells, P<0.01, and +138% in MDA-MB-231 cells, P<0.01), but decreased at high dose (≥ 25 mg.l⁻¹ −70 µM-) (−49%, in MCF7 cells, P<0.02, and −56% in MDA-MB-231 cells, P<0.025). At high dose, in both cell types, GSx-related metabolites decreased, including homocystein, creatine and taurine (−60 to −80%, all, P<0.05). Together with glutathione-S-transferase actvity, data established that GSx biosynthesis was upregulated at low dose, and GSx consumption activated at high dose. Another major target, in both cell types, was lipid metabolism involving, at high doses, accumulation of polyunsaturated and total free fatty acids (between ×4.5 and ×11, P<0.025), and decrease of glycerophospho-ethanolamine and -choline (about −60%, P<0.025). Multivariate statistical analyses showed a metabolic transition, even a biphasic behavior of some metabolites including GSx, between low and high doses. In addition, CUR at 10 mg.l⁻¹ in cotreatment with DTX induced modifications in glutathione metabolism, lipid metabolism, and glucose utilization. Some of these changes were biphasic depending on the duration of exposure to CUR.</p> <p>Conclusions/Significance</p><p>Metabolomics reveals major metabolic targets of CUR in breast cancer cells, and biphasic responses that challenge the widely accepted beneficial effects of the phytochemical.</p> </div

Potentialisation de la chimiothérapie par docétaxel par la curcumine dans les cancers du sein métastatiques (études expérimentales et clinique)

Le traitement des cancers du sein au stade métastatique repose notamment sur l'utilisation de la chimiothérapie. Parmi les traitements cytotoxiques les plus efficaces, le docétaxel est utilisé en monothérapie en 1ère ligne de traitement de ces cancers métastatiques. Cependant, son efficacité n'est pas optimale car moins de 5% des patients traités sont en réponse complète et 1/3 des patients auront une rechute métastatique. L'objectif de ce travail est d'optimiser le traitement du cancer du sein métastatique, et d'approfondir les connaissances sur la réponse des cellules tumorales à la chimiothérapie par docétaxel. Pour potentialiser l'efficacité de la chimiothérapie, nous avons utilisé une approche nouvelle, la nutrithérapie, qui consiste à utiliser les propriétés des phytonutriments ou de composés d'origine alimentaire. Nous avons choisi la curcumine, un dérivé polyphénolique extrait du curcuma, largement étudié in vitro et in vivo pour ses effets chimiopréventifs. Dans un 1er temps, nous avons étudié par une analyse métabolomique l'impact du docétaxel, de la curcumine et de la combinaison des 2 agents sur le métabolisme des cellules humaines MCF-7 de tumeur mammaire par spectroscopie RMN du proton. Dans un 2nd temps, nous avons conduit un essai clinique de phase I dont l'objectif principal était d'étudier la tolérance et la faisabilité de l'association du docétaxel à une supplémentation orale de curcumine chez des patients présentant un cancer du sein métastatique. Les résultats prometteurs en terme d'efficacité antitumorale et anti-angiogénique nous orientent ainsi vers une étude clinique de phase II évaluant l'efficacité de cette association.Chemotherapy is mainly used in metastatic breast cancer (MBC) treatment. Among chemotherapeutics, docetaxel is the most effective agent indicated as a first line monotherapy in MBC. However, as only 5% of complete responses are reported in treated patients and 30% of them would have metastatic relapse; efficacy of chemotherapy has to be improved. The aim of our work was to optimize therapy of MBC in expanding knowledge on the tumor cell response to docetaxel. The new approach nutritherapy consisting in the use of phytochemicals properties was applied to potentiate docetaxel chemotherapy. Curcumin was selected because of its pleiotropic effects, mostly in cancer cells. First, the metabolic variations of human MCF- breast cancer cells were studied in response to docetaxel and curcumin separately, and to the combinaison of both agents. Metabolomic analysis using proton NMR based-spectroscopy was performed to generate hypotheses in order to characterize time-and dose-dependent phenotypic variations in response to each treatment. Second, a phase I clinical trial was conducted to evaluate feasibility and tolerability of docetaxel/curcumin association in MBC patients. Promising clinical data in terms of antitumor and anti-angiogenic efficacy led us to initiate a comparative phase II clinical trial in order to evaluate the efficacy of such combinaison.CLERMONT FD-BCIU-Santé (631132104) / SudocSudocFranceF

CUR dose-related data in MCF7 cells (fold variation <i>vs.</i> 24 h controls).

<p>Mean control value is set to 1 while dispersion of control data is maintained. Metabolite abbreviations, see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0057971#pone-0057971-t001" target="_blank">Table 1</a>. SD, standard deviation. #, P<0.05; *, P<0.025; , P<0.01, Mann-Whitney test.

Mean control value is set to 1 while dispersion of control data is maintained. Metabolite abbreviations, see Table 1. SD, standard deviation. *, P < 0.025; , P < 0.01, Mann-Whitney test.</p

Dose-dependent response to CUR.

<p>A- Cellular DNA content in attached MCF7 (black bars) and MDA-MB-231 (white bars) breast cancer cells exposed to increasing concentrations (0.5, 2.5, 10, 25, and 50 mg.l⁻¹) of CUR for 24 h, measured using the Hoechst fluorescence intensity assay. *, P<0.05, **, P<0.01, CUR <i>vs.</i> CTL, Mann-Whitney test. B- DNA fragmentation expressed in tail DNA in MCF7 tumor cells exposed to increasing concentrations (0.5, 2.5, 10, 25, and 50 mg.l⁻¹) of CUR for 24 h, measured using the alkaline Comet assay. *, P<0.05, CUR <i>vs.</i> CTL, Mann-Whitney test. C- Total glutathione <i>S-</i>transferase (GST) activity in MCF7 tumor cells exposed to increasing concentrations (0.5, 2.5, 10, 25, and 50 mg.l⁻¹) of CUR for 24 h, measured using the Cayman Chemical Company assay kit. **, P<0.01, CUR <i>vs.</i> CTL, Mann-Whitney test. D- Set of typical 1D HRMAS ¹H-NMR spectra of MCF7 breast cancer cells in the 0.5–4.5 ppm spectral range. From bottom to top, cells exposed to 0, 0.5, 2.5, 10, 25, and 50 mg.l⁻¹ CUR. Thin arrows, signals from glutathione (GSx) at positions 2.25, 2.55, 2.98, and 3.78 ppm. Grey arrowheads, signal from total free fatty acids (tFA) at positions 1.30 and 0.89 ppm. E- Set of typical 1D HRMAS ¹H-NMR spectra of MDA-MB-231 breast cancer cells. From bottom to top, cells exposed to 0, 10, and 50 mg.l⁻¹ CUR. Arrows, as in Fig. 1D. F- Two-dimensional HRMAS ¹H-NMR spectra of intact MCF7 tumor cells, here from the 25 mg.l⁻¹ CUR group. Thin arrows, cross-peaks of glutathione (GSx) at positions 2.25×2.55, 2.25×3.78, 2.55×3.78, and 2.98×4.55 ppm×ppm. Square, area containing gluconate (Gna) signals. G- Square area magnified from the spectrum in Fig. 1F. Arrowheads, cross-peaks from Gna at positions 4.13×4.02, 4.02×3.76, and 3.76×3.66 ppm×ppm. Gna was quantified from the first 2 cross-peaks.</p

Co-mapping of dose-related metabolite changes with metabolic schemes.

<p>A- Glutathione cycle and metabolism. Abbreviations: GS-X, conjugated GSH; GSSG, oxidized GSH; GS-L, lactoyl-glutathione; Glu-Cys, glutamylcysteine; Cys, cysteine; Met, methionine; Ser, serine; GST, glutathione <i>S-</i>transferase; GSR, glutathione <i>S-</i>reductase; GLO1, glyoxalase-1; GS, glutathione synthetase; GCL, glutamate-cysteine ligase; OPase, oxoprolinase, CBS, cystathionine beta-synthase. Other metabolite abbreviations, as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0057971#pone-0057971-t001" target="_blank">Table 1</a>. Italicized, enzymes; Grey, undetected/unmeasured metabolites; #, P<0.05; *, P<0.025; , P<0.01, Mann-Whitney test. B- Glycolysis and lipid metabolism. Abbreviations: PtE, phosphatidylethanolamine; Glc, glucose; DAG, diacylglycerol; Pyr, pyruvate; ACoA, acetyl-CoA; PG, prostaglandin; CHK, choline kinase; CCT, choline cytidylyltransferase; PDH, pyruvate dehydrogenase; PLA₂, phospholipase A₂; FASN, fatty acid synthase; COX-2, cyclooxygenase-2. Other metabolite abbreviations, as in Table 1. #, P<0.05; *, P<0.025; , P<0.01, Mann-Whitney test.</p

Duration of exposure-related response to CUR.

<p>Cells were treated with 10 mg.l⁻¹ CUR and followed for 96 h. A- Cellular DNA content in attached MCF7 tumor cells relative to time-matched CTL. DNA content was measured using the Hoechst fluorescence intensity assay. *, P<0.05, **, P<0.01 CUR vs. CTL, Mann-Whitney test. B- Tail DNA relative to time-matched CTL. DNA. Tail DNA was assessed using the alkaline Comet assay. *, P<0.05, CUR vs. CTL, Mann-Whitney test. C- Total GST activity relative to time-matched CTL. GST activity was measured using the Cayman Chemical Company assay kit. **, P<0.01, CUR vs. CTL, Mann-Whitney test. D- Metabolite variations relative to time-matched CTL. #, P<0.05; *, P<0.025; $, P<0.01, CUR <i>vs.</i> CTL, Mann-Whitney test. E- PCA of duration of exposure-related data. The scores plot is displayed with individual scores labeled by their duration of exposure, between 24 h and 96 h, with CTL data also displayed. F1 and F2, first 2 principal components with the percentage of total variance of data they account for. Grey arrows, duration of exposure-related trajectory. F- Corresponding loading plot of PCA of duration of exposure-related data. Metabolite abbreviations, as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0057971#pone-0057971-t001" target="_blank">Table 1</a>.</p