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

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 cells (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 towards PT. Multidrug-resistant MDA-MB-231-BCRP cells were even more sensitive towards PT than sensitive MDA-MB-231-pcDNA3 cells with a resistance degree of 0.07 (collateral sensitivity). To the best of our knowledge, hypersensitivity (collateral sensitivity) of BCRP-overexpressing tumor cells has been reported 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 towards 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, the profound cytotoxic activity of PT against various cancer cell lines particularly against BCRP-overexpressing tumor cells suggesting PT as novel candidate compound for cancer treatment

Biopiracy <i>versus </i>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

Molecular mechanism of action and pharmacogenomics of curcumin, curcumin synthetic derivatives and combinations with curcumin in cancer therapy

Curcumin has been shown to be active against various cancers and it has also been seen to exhibit good synergism with other nutraceutical for example resveratrol, piperine and genistein. In this study we have investigated nineteen new synthetic derivatives of curcumin for their anticancer activity and further assessed the effect on cancer cells of a combination of curcumin and ascorbic acid (AA). The first part of this work focused on overcoming multidrug resistance and the nineteen synthetic derivatives of curcumin were tested on acute lymphoblastic CCRF-CEM leukemia cells and the P-gp overexpressing subline CEM/ADR5000. The cytotoxicity of the new synthetic derivatives was established using the resazurin assay. Ability to inhibit P-gp function was also assessed by the doxorubicin uptake assay and in silico studies of the same have been carried out using molecular docking tools and QSAR studies. The compounds displayed varied levels of cytotoxicity with some of them exhibiting lower IC50 values in comparison to the parent compound curcumin. The new synthetic derivatives also showed ability to inhibit P-gp function of extrusion of doxorubicin from the cells and some of them inhibited P-gp better than the control drug verapamil. These derivatives can be used to design novel and better P-gp inhibitors. The second part of our study focused on the cytotoxic effect of a combination of curcumin with AA on varied cancer cell lines. Here we carried out the chemoprofiling of three different members of the curcuma species and observed that curcumin was present in all curcuma species while AA was only available in C. longa. The combination of curcumin and AA was tested for cytotoxicity on human cancer cell lines including CCRF-CEM and CEM/ADR5000 leukemia, HCT116p53+/+ and HCT116p53-/- colon cancer, and U87MG and U87MG.∆EGRF glioblastoma. The drug combination exhibited additive cytotoxicity in leukemia and colon cancer cell lines while in the glioblastoma cell lines additive to supra additive cytotoxicity was recorded. Further we assessed the pharmacogenomics of curcumin and AA by microarray- based mRNA expression, COMPARE analysis and hierarchical cluster analysis. Gene profiles were obtained and they were used to predict sensitivity and resistance of the tumor cells to curcumin and AA. From these gene profiles we also established the gene functions that our compounds affected by assessing up and down regulation patterns as exhibited by the color coded heat map analysis. Both curcumin and AA affected varied groups of genes and varied functions in the cell lines. The pharmacogenomics results further supports the cytotoxicity results of additive effects.Curcumin besitzt nicht nur eine Wirksamkeit gegen verschiedene Krebsarten, sondern es wurden auch Synergien in Verbindung mit anderen Nutraceuticals wie Reservatol, Piperin und Genistein nachgewiesen. In der vorliegenden Arbeit untersuchte ich 19 neue synthetische Curcumin-Derivate auf ihre Wirksamkeit gegen Krebs. Außerdem bewertete ich den Effekt einer Kombination von Curcumin und Askorbinsäure (AA) auf Krebszellen. Der erste Teil der Arbeit konzentriert sich auf die Überwindung von Multidrug-Resistenzen. Aus diesem Grund wurden die 19 synthetischen Curcumin-Derivate an einer Zelllinie der akuten lymphoplastischen Leukämie (CCRF-CEM) und ihrer P-Glykoprotein-überexprimierenden Sublinie (CEM/ADR 500) getestet. Die Zytotoxizität der Derivate wurde mit einem Resazurin-Assay bestimmt. Die Fähigkeit zur Hemmung der P-Glykoprotein-Wirkung wurde mit einem Doxorubicin-Aufnhame-Test gemessen sowie mittels in silico Studien, bei denen molekulare Docking-Methoden und Berechnungen zu quantitative Struktur-Wirkungsbeziehungen (QSAR) verwendet wurden. Die Substanzen zeigten unterschiedliche Grade der Zytotoxizität, wobei einige im Vergleich zur Ursprungssubstanz sogar niedrigere IC50 Werte besaßen. Die neuen synthetischen Derivate besaßen außerdem die Fähigkeit die P-Glykoprotein-Wirkung zur Verdrängung von Doxorubicin aus den Zellen zu hemmen, einige sogar in höherem Maße als die Kontrolldroge Verapamil. Der zweite Teil der Arbeit konzentrierte sich auf den zytotoxischen Effekt von Curcumin mit AA auf verschiedenen Krebsarten. Es wurden Chemoprofile der sekundären Pflanzenstoffe von drei verschiedenen Mitgliedern der Curcuma-Familie erstellt, wobei in allen drei Arten Curcumin gefunden wurde, jedoch nur in einer Art, C. longa, AA nachgewiesen wurde. Die Kombination von Curcumin mit AA wurde an verschiedenen menschlichen Krebszelllinien, unter anderem CCRF-CEM und CEM/ADR 5000 Leukämie-Zellen, HTC116p53+/+ und HTC116p53-/- Darmkrebs-Zellen, sowie U87MG und U87MG.∆EGRF Glioblastom-Zellen, getestet. Eine Kombination der Substanzen ergab einen additiven Effekt auf Leukämie- und Darmkrebs-Zellen, wohingegen die Glioblastom-Zelllinien additive bis supra additive Effekte zeigten. Desweiteren beschäftigte ich mich mit den pharmakogenomischen Eigenschaften von Curcumin und AA über Microarray-basierte mRNA Expressionsprofile, COMPARE-Analyse und hierarchische Cluster-Analyse. Genprofile wurden erstellt und zur Vorhersage der Empfindlichkeit bzw. Resistenz der Tumorzellen gegenüber Curcumin und AA benutzt. Diese Genprofile zeigten hoch- und nieder-exprimierte Gene in den verschiedenen Zelllinien. Es wurde ebenfalls eine color-coded heatmap analysis für die betroffenen Gene erstellt. Beide, Curcumin und AA beeinflussten vielfältige Gengruppen und unterschiedliche Funktionen innerhalb der Zelllinien. Die pharmakogenomischen Ergebnisse unterstützen die Ergebnisse zu dem additiven zytotoxischen Effekt der beiden Substanzen

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. 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

The CINAMR (Clinical Information Network-Antimicrobial Resistance) Project: A pilot microbial surveillance using hospitals linked to regional laboratories in Kenya: Study Protocol [version 1; peer review: 2 approved]

Background: Antimicrobial resistance (AMR) is a global threat and is thought to be acute in low-and middle-income country (LMIC) settings, including in Kenya, but there is limited unbiased surveillance that can provide reliable estimates of its burden. Current efforts to build capacity for microbiology testing in Kenya are unlikely to result in systematic routine microbiological testing in the near term. Therefore, there is little prospect for microbiological support to inform clinical diagnoses nor for indicating the burden of AMR and for guiding empirical choice of antibiotics. Objective: We aim to build on an existing collaboration, the Clinical Information Network (CIN), to pilot microbiological surveillance using a ‘hub-and-spoke’ model where selected hospitals are linked to high quality microbiology research laboratories. Methods: Children admitted to paediatric wards of 12 participating hospitals will have a sample taken for blood culture at admission before antibiotics are started. Indication for blood culture will be a clinician’s prescription of antibiotics. Samples will then be transported daily to the research laboratories for culture and antibiotic susceptibility testing and results relayed back to clinicians for patient management. The surveillance will take place for 6 months in each hospital. Separately, we shall conduct semi-structured interviews with frontline health workers to explore the feasibility and utility of this approach. We will also seek to understand how the availability of microbiology results might inform antibiotic stewardship, and as an interim step to the development of better national or regional laboratories linked to routine surveillance. Conclusions: If feasible, this approach is less costly and periodic ‘hub-and-spoke’ surveillance can be used to track AMR trends and to broadly guide empirical antibiotic guidance meaning it is likely to be more sustainable than establishing functional microbiological facilities in each hospital in a LMIC setting

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 securedDeutsche Forschungsgemeinschaft/[GRK2015/1]//AlemaniaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Centro de Investigación en Enfermedades Tropicales (CIET