In Silico and In Vitro Screening of 50 Curcumin Compounds as EGFR and NF-κB Inhibitors.

The improvement of cancer chemotherapy remains a major challenge, and thus new drugs are urgently required to develop new treatment regimes. Curcumin, a polyphenolic antioxidant derived from the rhizome of turmeric (Curcuma longa L.), has undergone extensive preclinical investigations and, thereby, displayed remarkable efficacy in vitro and in vivo against cancer and other disorders. However, pharmacological limitations of curcumin stimulated the synthesis of numerous novel curcumin analogs, which need to be evaluated for their therapeutic potential. In the present study, we calculated the binding affinities of 50 curcumin derivatives to known cancer-related target proteins of curcumin, i.e., epidermal growth factor receptor (EGFR) and nuclear factor κB (NF-κB) by using a molecular docking approach. The binding energies for EGFR were in a range of −12.12 (±0.21) to −7.34 (±0.07) kcal/mol and those for NF-κB ranged from −12.97 (±0.47) to −6.24 (±0.06) kcal/mol, indicating similar binding affinities of the curcumin compounds for both target proteins. The predicted receptor-ligand binding constants for EGFR and curcumin derivatives were in a range of 0.00013 (±0.00006) to 3.45 (±0.10) µM and for NF-κB in a range of 0.0004 (±0.0003) to 10.05 (±4.03) µM, indicating that the receptor-ligand binding was more stable for EGFR than for NF-κB. Twenty out of 50 curcumin compounds showed binding energies to NF-κB smaller than −10 kcal/mol, while curcumin as a lead compound revealed free binding energies of >−10 kcal/mol. Comparable data were obtained for EGFR: 15 out of 50 curcumin compounds were bound to EGFR with free binding energies of <−10 kcal/mol, while the binding affinity of curcumin itself was >−10 kcal/mol. This indicates that the derivatization of curcumin may indeed be a promising strategy to improve targe specificity and to obtain more effective anticancer drug candidates. The in silico results have been exemplarily validated using microscale thermophoresis. The bioactivity has been further investigated by using resazurin cell viability assay, lactate dehydrogenase assay, flow cytometric measurement of reactive oxygen species, and annexin V/propidium iodide assay. In conclusion, molecular docking represents a valuable approach to facilitate and speed up the identification of novel targeted curcumin-based drugs to treat cancer.

Collateral Sensitivity of Parthenolide via NF-κB and HIF-α Inhibition and Epigenetic Changes in Drug-Resistant Cancer Cell Lines.

Parthenolide (PT) is a sesquiterpene lactone isolated from Tanacetum parthenium. In this study, PT showed varying cytotoxic effects against different solid tumor cell lines. HCT116 (p53+/+) colon carcinoma cells and their parental HCT116 knockout p53 (p53-/-) cell lines showed a resistance degree of 2.36. On the other hand, wild-type U87.MG cells or cells transfected with a deletion-activated EGFR cDNA (U87.MGΔEGFR) exhibited slight sensitivity toward PT. Multidrug-resistant MDA-MB-231-BCRP cells were even more sensitive toward PT than sensitive MDA-MB-231-pcDNA cells with a resistance degree of 0.07 (collateral sensitivity). To the best of our knowledge, hypersensitivity (collateral sensitivity) in MDA-MB-231-BCRP cell line is reported in this study for the first time. We attempted to identify the mechanism of collateral sensitivity. Firstly, we found that PT bound to IKK preventing IκBα degradation and eventually inhibition of the nuclear factor kappa B (NF-κB) pathway. Down-regulation of hypoxia inducing factor 1-alpha (HIF-1α) in MDA-MB-231-BCRP resistant cells may be a second mechanism, since it is a target gene of NF-κB. Moreover, PT also showed epigenetic effect by inhibition of HDAC activity as shown using both molecular docking and HDAC activity assay. Based on COMPARE and hierarchical cluster analyses, we found gene expression profiles that predicted sensitivity or resistance of 47 tumor cell lines toward PT. Interestingly, pathway analyses of gene expression profiles revealed NF-κB and HIF signaling as top networks of these genes, cellular functions and canonical pathways influencing the activity of PT against tumor cells. In conclusion, PT exerted profound cytotoxic activity against various cancer cell lines mainly against BCRP-overexpressing tumor cells, suggesting PT as novel candidate for cancer treatment.

Biopiracy versus One-World Medicine-From colonial relicts to global collaborative concepts.

BACKGROUND: Practices of biopiracy to use genetic resources and indigenous knowledge by Western companies without benefit-sharing of those, who generated the traditional knowledge, can be understood as form of neocolonialism. HYPOTHESIS: The One-World Medicine concept attempts to merge the best of traditional medicine from developing countries and conventional Western medicine for the sake of patients around the globe. STUDY DESIGN: Based on literature searches in several databases, a concept paper has been written. Legislative initiatives of the United Nations culminated in the Nagoya protocol aim to protect traditional knowledge and regulate benefit-sharing with indigenous communities. The European community adopted the Nagoya protocol, and the corresponding regulations will be implemented into national legislation among the member states. Despite pleasing progress, infrastructural problems of the health care systems in developing countries still remain. Current approaches to secure primary health care offer only fragmentary solutions at best. Conventional medicine from industrialized countries cannot be afforded by the impoverished population in the Third World. Confronted with exploding costs, even health systems in Western countries are endangered to burst. Complementary and alternative medicine (CAM) is popular among the general public in industrialized countries, although the efficacy is not sufficiently proven according to the standards of evidence-based medicine. CAM is often available without prescription as over-the-counter products with non-calculated risks concerning erroneous self-medication and safety/toxicity issues. The concept of integrative medicine attempts to combine holistic CAM approaches with evidence-based principles of conventional medicine. CONCLUSION: To realize the concept of One-World Medicine, a number of standards have to be set to assure safety, efficacy and applicability of traditional medicine, e.g. sustainable production and quality control of herbal products, performance of placebo-controlled, double-blind, randomized clinical trials, phytovigilance, as well as education of health professionals and patients.

Pharmacogenomic Characterization and Isobologram Analysis of the Combination of Ascorbic Acid and Curcumin-Two Main Metabolites of Curcuma longa-in Cancer Cells.

Curcuma longa has long been used in China and India as anti-inflammatory agent to treat a wide variety of conditions and also as a spice for varied curry preparations. The chemoprofile of the Curcuma species exhibits the presence of varied phytochemicals with curcumin being present in all three species but AA only being shown in C. longa. This study explored the effect of a curcumin/AA combination on human cancer cell lines. The curcumin/AA combination was assessed by isobologram analysis using the Loewe additivity drug interaction model. The drug combination showed additive cytotoxicity toward CCRF-CEM and CEM/ADR5000 leukemia cell lines and HCT116p53+/+ and HCT116p53-/- colon cancer cell line, while the glioblastoma cell lines U87MG and U87MG.ΔEGFR showed additive to supra-additive cytotoxicity. Gene expression profiles predicting sensitivity and resistance of tumor cells to induction by curcumin and AA were determined by microarray-based mRNA expressions, COMPARE, and hierarchical cluster analyses. Numerous genes involved in transcription (TFAM, TCERG1, RGS13, C11orf31), apoptosis-regulation (CRADD, CDK7, CDK19, CD81, TOM1) signal transduction (NR1D2, HMGN1, ABCA1, DE4ND4B, TRIM27) DNA repair (TOPBP1, RPA2), mRNA metabolism (RBBP4, HNRNPR, SRSF4, NR2F2, PDK1, TGM2), and transporter genes (ABCA1) correlated with cellular responsiveness to curcumin and ascorbic acid. In conclusion, this study shows the effect of the curcumin/AA combination and identifies several candidate genes that may regulate the response of varied cancer cells to curcumin and AA.

Modulation of P-glycoprotein activity by novel synthetic curcumin derivatives in sensitive and multidrug-resistant T-cell acute lymphoblastic leukemia cell lines.

BACKGROUND: Multidrug resistance (MDR) and drug transporter P-glycoprotein (P-gp) represent major obstacles in cancer chemotherapy. We investigated 19 synthetic curcumin derivatives in drug-sensitive acute lymphoblastic CCRF-CEM leukemia cells and their multidrug-resistant P-gp-overexpressing subline, CEM/ADR5000. MATERIAL AND METHODS: Cytotoxicity was tested by resazurin assays. Doxorubicin uptake was assessed by flow cytometry. Binding modes of compounds to P-gp were analyzed by molecular docking. Chemical features responsible for bioactivity were studied by quantitative structure activity relationship (QSAR) analyses. A 7-descriptor QSAR model was correlated with doxorubicin uptake values, IC50 values and binding energies. RESULTS: The compounds displayed IC50 values between 0.7±0.03 and 20.2±0.25µM. CEM/ADR5000 cells exhibited cross-resistance to 10 compounds, collateral sensitivity to three compounds and regular sensitivity to the remaining six curcumins. Molecular docking studies at the intra-channel transmembrane domain of human P-gp resulted in lowest binding energies ranging from -9.00±0.10 to -6.20±0.02kcal/mol and pKi values from 0.24±0.04 to 29.17±0.88µM. At the ATP-binding site of P-gp, lowest binding energies ranged from -9.78±0.17 to -6.79±0.01kcal/mol and pKi values from 0.07±0.02 to 0.03±0.03µM. CEM/ADR5000 cells accumulated approximately 4-fold less doxorubicin than CCRF-CEM cells. The control P-gp inhibitor, verapamil, partially increased doxorubicin uptake in CEM/ADR5000 cells. Six curcumins increased doxorubicin uptake in resistant cells or even exceeded uptake levels compared to sensitive one. QSAR yielded good activity prediction (R=0.797 and R=0.794 for training and test sets). CONCLUSION: Selected derivatives may serve to guide future design of novel P-gp inhibitors and collateral sensitive drugs to combat MDR.

Biopiracy of natural products and good bioprospecting practice.

BACKGROUND: Biopiracy mainly focuses on the use of biological resources and/or knowledge of indigenous tribes or communities without allowing them to share the revenues generated out of economic exploitation or other non-monetary incentives associated with the resource/knowledge. METHODS: Based on collaborations of scientists from five continents, we have created a communication platform to discuss not only scientific topics, but also more general issues with social relevance. This platform was termed 'PhytCancer -Phytotherapy to Fight Cancer' (www.phyt-cancer.uni-mainz.de). As a starting point, we have chosen the topic "biopiracy", since we feel this is of pragmatic significance for scientists working with medicinal plants. RESULTS: It was argued that the patenting of herbs or natural products by pharmaceutical corporations disregarded the ownership of the knowledge possessed by the indigenous communities on how these substances worked. Despite numerous court decisions in U.S.A. and Europe, several international treaties, (e.g. from United Nations, World Health Organization, World Trade Organization, the African Unity and others), sharing of a rational set of benefits amongst producers (mainly pharmaceutical companies) and indigenous communities is yet a distant reality. In this paper, we present an overview of the legal frameworks, discuss some exemplary cases of biopiracy and bioprospecting as excellent forms of utilization of natural resources. CONCLUSIONS: We suggest certain perspectives, by which we as scientists, may contribute towards prevention of biopiracy and also to foster the fair utilization of natural resources. We discuss ways, in which the interests of indigenous people especially from developing countries can be secured.

Artemisinin derivatives induce iron-dependent cell death (ferroptosis) in tumor cells.

BACKGROUND: Apoptosis and other forms of cell death have been intensively investigated in the past years to explain the mode of action of synthetic anticancer drugs and natural products. Recently, a new form of cell death emerged, which was termed ferroptosis, because it depends on intracellular iron. Here, the role of genes involved in iron metabolism and homeostasis for the cytotoxicity of ten artemisinin derivatives have been systematically investigated. MATERIAL AND METHODS: Log10IC50 values of 10 artemisinin derivatives (artesunate, artemether, arteether, artenimol, artemisitene, arteanuin B, another monomeric artemisinin derivative and three artemisinin dimer molecules) were correlated to the microarray-based mRNA expression of 30 iron-related genes in 60 cell lines of the National Cancer Institute (NCI, USA) as determined in 218 different microarray hybridization experiments. The effect of desferoxamine and ferrostatin-1 on the cytotoxicity of artenimol of CCRF-CEM cells was determined by resazurin assays. The mRNA expression of TFRC was exemplarily validated by immunohistochemical detection of transferrin receptor protein expression. RESULTS: The mRNA expression of 20 genes represented by 59 different cDNA clones significantly correlated to the log10IC50 values for the artemisinins, including genes encoding transferrin (TF), transferrin receptors 1 and 2 (TFRC, TFR2), cerulopasmin (CP), lactoferrin (LTF) and others. The ferroptosis inhibitor ferrostatin-1 and the iron chelator deferoxamine led to a significantly reduced cytotoxicity of artenimol, indicating ferroptosis as cell death mode. CONCLUSION: The numerous iron-related genes, whose expression correlated with the response to artemisinin derivatives speak in factor for the relevance of iron for the cytotoxic activity of these compounds. Treatment with ferroptosis-inducing agents such as artemisinin derivatives represents an attractive strategy for cancer therapy. Pre-therapeutic determination of iron-related genes may indicate tumor sensitivity to artemisinins. Ferroptosis induced by artemisinin-type drugs deserve further investigation for individualized tumor therapy.