Loss of CUL4A expression is underlying cisplatin hypersensitivity in colorectal carcinoma cells with acquired trabectedin resistance.

BACKGROUND: Colorectal carcinoma (CRC) is the third most common cancer worldwide. Platinum-based anticancer compounds still constitute one mainstay of systemic CRC treatment despite limitations due to adverse effects and resistance development. Trabectedin has shown promising antitumor effects in CRC, however, again resistance development may occur. In this study, we aimed to develop strategies to circumvent or even exploit acquired trabectedin resistance in novel CRC treatment regimens. METHODS: Human HCT116 CRC cells were selected for acquired trabectedin resistance in vitro and characterised by cell biological as well as bioinformatic approaches. In vivo xenograft experiments were conducted. RESULTS: Selection of HCT116 cells for trabectedin resistance resulted in p53-independent hypersensitivity of the selected subline against cisplatin. Bioinformatic analyses of mRNA microarray data suggested deregulation of nucleotide excision repair and particularly loss of the ubiquitin ligase CUL4A in trabectedin-selected cells. Indeed, transient knockdown of CUL4A sensitised parental HCT116 cells towards cisplatin. Trabectedin selected but not parental HCT116 xenografts were significantly responsive towards cisplatin treatment. CONCLUSIONS: Trabectedin selection-mediated CUL4A loss generates an Achilles heel in CRC cancer cells enabling effective cisplatin treatment. Hence, inclusion of trabectedin in cisplatin-containing cancer treatment regimens might cause profound synergism based on reciprocal resistance prevention.

Rb/Sr isotopic and compositional retentivity of muscovite during deformation.

Permian metapegmatite muscovite from the Upper-Austroalpine Matsch Unit in Southern Tyrol (Italy) was investigated regarding its Rb/Sr and compositional retentivity during Cretaceous Upper-greenschist facies deformation. The data imply that microstructurally relic Permian magmatic muscovite largely maintained its major and trace element compositions during deformation, whereas the Rb/Sr geochronometer is strongly affected by a net loss of Sr. Lower Sr concentrations of muscovite correlate with higher 87Rb/86Sr and 87Sr/86Sr ratios. In most samples, the muscovite grain size- and magnetic-fractions with the lowest 87Rb/86Sr and 87Sr/86Sr ratios preserve a Permo-Triassic muscovite-whole rock Rb/Sr apparent age interpreted as to reflect formation during or cooling after pegmatite emplacement. Contrastingly, muscovite fractions with higher 87Rb/86Sr and 87Sr/86Sr ratios are arranged along a roughly linear array with a positive correlation of the 87Rb/86Sr and 87Sr/86Sr ratios in the 87Rb/86Sr vs 87Sr/86Sr space. They yield successively lower muscovite-whole rock Rb/Sr apparent ages. We explain the variations in the Rb/Sr isotopic character of microstructurally relic muscovite by a, presumably deformation-related, loss of Sr during the Cretaceous event. Contemporaneously, only very limited amounts of isotopically different Sr from the matrix reservoir might possibly have entered the muscovite. Consequently, the Rb/Sr of the relic muscovite is affected by a net loss of Sr. The results imply that at temperatures of < 500 °C, deformation is supposed to be the predominant factor in controlling the Rb/Sr geochronometer of relic muscovite, by significantly reducing the characteristic length scale for volume diffusion. However, variations of the major and trace element compositions within Permian relic muscovite are interpreted to rather reflect primary compositional instead of deformation-related variations.

[Air quality in schools - classroom levels of carbon dioxide (CO2), volatile organic compounds (VOC), aldehydes, endotoxins and cat allergen]. / Raumluftqualität in Schulen - Belastung von Klassenräumen mit Kohlendioxid (CO2), flüchtigen organischen Verbindungen (VOC), Aldehyden, Endotoxinen und Katzenallergenen.

Children are assumed to be more vulnerable to health hazards and spend a large part of their time in schools. To assess the exposure situation in this microenvironment, we evaluated the indoor air quality in winter 2004/5 in 92 classrooms, and in 75 classrooms in summer 2005 in south Bavaria, Germany. Indoor air climate parameters (temperature, relative humidity), carbon dioxide (CO2) and various volatile organic compounds, aldehydes and ketones were measured. Additionally, cat allergen (Fel d1) and endotoxin (LAL-test) were analysed in the settled dust of school rooms. Data on room and building characteristics were collected by use of a standardised form. Only data collected during teaching hours were considered in analysis. The median indoor CO2 concentration in the classrooms ranged in the winter and summer period from 598 to 4 172 ppm and 480 to 1 875 ppm, respectively. While during the winter period in 92% of the classrooms the CO2 daily medians went above 1 000 ppm, the percentage of classrooms with increased CO2 concentration fell to 28% in summer. In winter, in 60% of classes the daily median CO2 concentration exceeded 1 500 ppm, while in summer this threshold was reached by only 9%. A high concentration of CO2 was associated with a high number of pupils, a low room surface area and a low room volume. The levels of total volatile organic compounds (TVOC) in classrooms ranged between 110 and 1 000 microg/m3 (median in winter 345 microg/m3, in summer 260 microg/m3). Acetone, formaldehyde and acetaldehyde were measured in concentrations from 14.0 to 911 microg/m3, from 3.1 to 46.1 microg/m3, and from 2.9 to 78 microg/m3, respectively. The other aldehydes were detected in minor amounts only. The median Fel d1 level in winter was 485 ng/g dust (20 to 45 160 ng/g) and in summer it was 417 ng/g (40-7 470 ng/g). We observed no marked differences between the two sampling periods and between smooth floors and rooms with carpeted floors. No differences were found according to room surface area and room volume. The median endotoxin contents in winter and summer were 19.7 EU/mg dust (6.6 to 154 EU/mg) and 32.2 EU/mg (9.6 to 219 EU/mg), respectively. The levels varied significantly between the sampling periods, but were independent of room surface area, room volume and surface floorings. Overall the results of VOC, aldehydes, ketones and endotoxin indicate, in general, a low exposure level in classrooms. The observed concentrations of cat allergens should be considered as a meaningful exposure route and thus could be tackled within preventive programs.

Multidrug-resistant cancer cells are preferential targets of the new antineoplastic lanthanum compound KP772 (FFC24).

Recently, we have introduced [tris(1,10-phenanthroline)lanthanum(III)] trithiocyanate (KP772, FFC24) as a new lanthanum compound which has promising anticancer properties in vivo and in vitro. Aim of this study was to investigate the impact of ABC transporter-mediated multidrug resistance (MDR) on the anticancer activity of KP772. Here, we demonstrate that all MDR cell models investigated, overexpressing ABCB1 (P-glycoprotein), ABCC1 (multidrug resistance protein 1), or ABCG2 (breast cancer resistance protein) either due to drug selection or gene transfection, were significantly hypersensitive against KP772. Using ABCB1-overexpressing KBC-1 cells as MDR model, KP772 hypersensitivity was demonstrated to be based on stronger apoptosis induction and/or cell cycle arrest at unaltered cellular drug accumulation. KP772 did neither stimulate ABCB1 ATPase activity nor alter rhodamine 123 accumulation arguing against a direct interaction with ABCB1. Accordingly, several drug resistance modulators did not sensitize but rather protect MDR cells against KP772-induced cytotoxicity. Moreover, long-term KP772 treatment of KBC-1 cells at subtoxic concentrations led within 20 passages to a complete loss of drug resistance based on blocked MDR1 gene expression. When exposing parental KB-3-1 cells to subtoxic, stepwise increasing KP772 concentrations, we observed, in contrast to several other metallo-drugs, no acquisition of KP772 resistance. Summarizing, our data demonstrate that KP772 is hyperactive in MDR cells and might have chemosensitizing properties by blocking ABCB1 expression. Together with the disability of tumor cells to acquire KP772 resistance, our data suggest that KP772 should be especially active against notoriously drug-resistant tumor types and as second line treatment after standard chemotherapy failure.

Integrated Exposure Assessment Survey (INES) exposure to persistent and bioaccumulative chemicals in Bavaria, Germany.

The Integrated Exposure Assessment Survey (INES) was started in the year 2005. Altogether 50 healthy adults living in Bavaria, Germany, were included into the study. Monitoring was conducted in accordance with relevant routes of human exposure (inhalation, ingestion) and integrated different pathways (indoor air, food, house dust). This approach consisted of a combination of external measurements of contaminants with the determination of these substances or their metabolites in body fluids. The target substances were phthalates, perfluorinated compounds (PFC), polychlorinated biphenyls (PCBs), polychlorinated dibenzo-p-dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs), and polybrominated diphenylethers (PBDEs). This paper gives a brief description of the objectives and the concept of INES as well as methods of sampling and analyses of target compounds. Some preliminary results of biomonitoring data for PFC and phthalates as well as of the dietary intake of DEHP will be discussed.

Tissue distribution after a single subcutaneous administration of 2,3,7,8-tetrabromodibenzo-p-dioxin in comparison with toxicokinetics of 2,3,7,8-tetrachlorodibenzo-p-dioxin in female Wistar rats.

Tissue concentrations of 2,3,7,8-tetrabromodibenzo-p-dioxin (TBDD) and induction of ethoxyresorufin O-deethylase (EROD) were determined in female Wistar rats following a single subcutaneous (s.c.) injection of TBDD. Two sets of experiments were performed in order to study (a) the time course after a single s.c. administration of 600 ng TBDD/kg body wt up to 78 days, and (b) the dose-response seven days after a single s.c. injection of different doses of TBDD (3 to 3,000 ng/kg body wt). The results obtained on toxicokinetics and enzyme induction were compared with those following a single s.c. administration of 300 ng/kg body wt 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Three days after the injection, approximately 93% of TBDD and 90% of TCDD had been absorbed. Fourteen days after s.c. injection less than 1% of the administered dose of both substances remained at the injection site. Three days after a single s.c. injection of 600 ng TBDD/kg body wt and 300 ng TCDD/kg body wt, the maximum tissue concentrations in the liver amounted to (M +/- S.D.) 5.7 +/- 0.8 and 4.7 +/- 0.9 ng/g wet weight, respectively. In adipose tissue, the peak concentration was 3.2 +/- 0.2 ng/g wet weight for TBDD on day 14, and 0.8 +/- 0.1 ng/g for TCDD on day 7. Throughout the study, the concentration ratio in the TCDD-treated group was always at least twice as high as that in the TBDD-treated group. The elimination half-life (t1/2) of TBDD and of TCDD in the liver was 13.3 and 13.6 days, respectively. In the adipose tissue the t1/2 of TCDD was 24.5 days but no reliable t1/2 could be calculated for TBDD (t1/2 = 39.4 days with a 95% confidence interval of 25.9 to 82.4 days). Tissue content of TBDD and TCDD in liver and adipose tissue increased dose-dependently, and the linear regression in a double-logarithmic plot showed a straight line. Time course of the induction of hepatic EROD activity after treatment with 600 ng TBDD/kg body wt was almost identical with that observed following a single dose of 300 ng TCDD/kg body wt. The induction of hepatic EROD activity was linearly correlated in a double-logarithmic plot to the hepatic concentrations of the congeners (both TBDD and TCDD). The slopes of the dose-response curves after administration of TBDD and TCDD were almost parallel for tissue concentrations ranging from 0.1 to 30 ng/g wet weight.

Phenolic xenoestrogens in surface water, sediments, and sewage sludge from Baden-Württemberg, south-west Germany.

Nine structurally different phenolic chemicals, which have been reported to mimic estrogen effects, were determined in various aquatic environmental compartments. Twenty-three water samples from five streams and rivers showed levels up to 458 ng/l for 4-nonylphenol (4NP), 189 ng/l for 4-t-octylphenol (4tOP), 272 ng/l for bisphenol A (BPA) and 47 ng/l for 2-hydroxybiphenyl (2OHBiP). Elevated levels of these compounds in a stream with a high load of effluents of sewage treatment plants (STPs), compared to a brook free of sewage, identified STPs as major sources. With a similar order, 4NP (10-259 micrograms/kg dry matter), 4tOP (< 0.5-8 micrograms/kg), BPA (< 0.5-15 micrograms/kg), and 2OHBiP (2-69 micrograms/kg) were also detected regularly in riverine sediment (n = 11). Levels in sewage sludge were one order of magnitude higher than in sediments. 4-Hydroxybiphenyl and 4-chloro-3-methylphenol were found predominantly in sludge and sediment in the lower ppb range.

Development of a sensitive E-screen assay for quantitative analysis of estrogenic activity in municipal sewage plant effluents.

A simplified proliferation test with human estrogen receptor-positive MCF-7 breast cancer cells (E-screen assay) was optimized and validated for the sensitive quantitative determination of total estrogenic activity in effluent samples from municipal sewage plants. After solid phase extraction of 1 l sewage on either 0.2 g polystyrene copolymer (ENV+) or 1 g RP-C18 material and removal of the solvent, analysis of the extracts in the E-screen assay could be performed without any clean-up step. This was even possible with untreated sewage. Parallel extraction of four sewage samples on both different solid phase materials gave comparable quantitative results in the E-screen. A blank sample did not induce cell proliferation. As additive behaviour of the estrogenic response of single compounds was proven for two different mixtures each containing three xenoestrogens, total estrogenic activity in the sewage samples, expressed as 17 beta-estradiol equivalent concentration (EEQ), could be calculated comparing the EC50 values of the samples with those of the positive control 17 beta-estradiol. The detection limit of the E-screen method was 0.05 pmol EEQ/l (0.014 ng EEQ/l), the limit of quantification 0.25-0.5 pmol EEQ/l (0.07-0.14 ng EEQ/l). In total, extracts of nine effluent and one influent sample from five different municipal sewage plants in South Germany were analyzed in the E-screen. All samples strongly induced cell proliferation in a dose-dependent manner which was completely inhibited by coincubation with 5 nM of the estrogen receptor-antagonist ICI 182,780. The proliferative effect relative to the positive control 17 beta-estradiol (RPE) was between 30 and 101%. 17 beta-Estradiol equivalent concentrations were between 2.5 and 25 ng/l indicating a significant input of estrogenic substances via sewage treatment plants into rivers.

Development and validation of a GC/MS method for determination of phenolic xenoestrogens in aquatic samples.

A simple and sensitive GC/MS method for the quantitative determination of the estrogenic phenolic compounds 4-nonylphenol, 4-t-octylphenol, bisphenol A, 3-t-butyl-4-hydroxyanisole, 2-t-butyl-4-methylphenol, 4-hydroxybiphenyl, 2-hydroxybiphenyl, 4-chloro-3-methylphenol, and 4-chloro-2-methylphenol in aquatic samples was developed. The method for assessing their occurrence in sewage, surface and drinking waters consists of solid phase extraction (SPE) using a polystyrene copolymer phase. After methylation of the extract HRGC/LRMS analysis was possible without any clean up, even in raw sewage samples. Limits of detection and determination were between <0.01 and 0.05 ng/l and 0.01 and 0.05 ng/l, respectively. Recoveries were above 70% with exception of 3-t-butyl-4-hydroxyanisole.

Concentrations and profiles of PCDDs and PCDFs in human mammary carcinoma tissue.

PCDD/PCDF concentrations in eight mammary carcinoma tissue samples obtained after surgical excision were similar to those found in two healthy breast glandular tissue samples from autopsy material. These levels agree well with mean concentrations in human adipose tissue from German adults. An analogous consistency was found for the congener profiles of the normalized concentrations, also in comparison with mothers' milk from Germany. In spite of similar congener profiles the concentrations in four axillary adipose tissue samples corresponding to the carcinoma samples were about 40% lower. This discrepancy was not found in one tissue pair from a healthy breast.

Input/output balance of estrogenic active compounds in a major municipal sewage plant in Germany.

24 h samples of untreated and treated wastewater were taken in parallel from a modern municipal sewage plant in southern Germany in March and June 1998. After solid phase extraction, total estrogenic activity was quantitatively measured with a miniaturized E-screen assay and the levels of nine estrogenic phenolic chemicals analyzed by HRGC/LRMS. 17Beta-estradiol equivalent concentrations (EEQ) were 58 and 70 ng/l in the influent and 6 ng/l in the effluent, indicating that the load of estrogenic activity of the wastewater was reduced by about 90% in the sewage plant. Less than 3% of the estrogenic activity was found in the sludge. 4-t-octylphenol, 4-nonylphenol, bisphenol A, 2-hydroxybiphenyl, and 4-chloro-3-methylphenol were detected in the untreated wastewater at levels from 0.13 to 3.6 microg/l. 4-t-octylphenol, 4-nonylphenol, and bisphenol A were present in the effluent at concentrations from 0.16 to 0.36 microg/l, 2-hydroxybiphenyl and 4-chloro-3-methylphenol were not detectable. The contribution of the quantified levels of phenolic xenoestrogens to total estrogenic activity in the sewage was 0.7-4.3%.

Validation and application of a rapid in vitro assay for assessing the estrogenic potency of halogenated phenolic chemicals.

The E-Screen assay serves as an in vitro tool for the detection of estrogenic activity of chemicals and extracts of environmental samples. Based on the induction of proliferation in human estrogen receptor-positive MCF-7 breast cancer cells we could substantially simplify the assay. As one important step of validation we applied the modified assay for testing nine known xenoestrogens. We could confirm the results of other groups assuring the reproducibility of the E-Screen assay. The results provide evidence that the E-Screen assay is suitable for determination of estradiol equivalency factors (EEFs) for environmental estrogens to rank their estrogenic potency relative to the natural estrogen 17 beta-estradiol. Further, we used the optimized proliferation test to screen nine halogenated phenolic compounds for their possible estrogenic potency. Three widely applied chemicals expressed a weak receptor-mediated estrogenic activity: the flame retardant Tetrabromo-Bisphenol-A, the disinfectant 4-chloro-3-methylphenol, and the herbicide educt 4-chloro-2-methylphenol. Their estrogenic potencies were five to six orders of magnitude lower than that of 17 beta-estradiol.

Substances with estrogenic activity in effluents of sewage treatment plants in southwestern Germany. 1. Chemical analysis.

A gas chromatography/mass spectrometry method for the simultaneous quantitative determination of natural and synthetic estrogens (17beta-estradiol, estrone, 17alpha-ethinylestradiol, and mestranol), phytoestrogens (genistein and beta-sitosterol), and xenoestrogens (benzyl butyl phthalate, dibutyl phthalate, bisphenol A, 4-nonylphenol [NP], 4-nonylphenoxyacetic acid [NP1EC], 4-nonylphenol diethoxylate [NP2EO], and alpha-endosulfan) in effluents of sewage treatment plants (STPs) was developed. Identification and quantification were carried out with the standard addition method using analyte-specific and, in some cases, deuterium-labeled internal standards. The effluents of 18 STPs were investigated. Apart from alpha-endosulfan and mestranol, all selected substances were detected in the majority of samples. The median concentrations of steroidal estrogens were between 0.4 ng/L (17alpha-ethinylestradiol) and 1.6 ng/L (17beta-estradiol). The metabolites of the nonylphenol polyethoxylates, NP, NPIEC, and NP2EO were found in concentrations ranging from the upper-ng/L-range (NP) to the lower-microg/L range (NP1EC). For all substances except mestranol and alpha-endosulfan, median values were calculated and compared to the results of other investigations in Europe and the United States. Possible dependencies of measured concentrations on the geographical location, the capacity, the influent composition, and the technical fitting of the STPs are discussed.

Substances with estrogenic activity in effluents of sewage treatment plants in southwestern Germany. 2. Biological analysis.

The proliferation test with human estrogen receptor-positive MCF-7 breast cancer cells (E-Screen assay) was applied for quantitative determination of total estrogenic activity in 24-h composite effluent samples from 16 municipal and two industrial sewage treatment plants (STPs) in the state of Baden-Württemberg, southwestern Germany. The estrogenic efficacy relative to the positive control, 17beta-estradiol, was between 26 and 74% (median, 48%) for the 16 municipal STPs. Estradiol equivalent concentrations (EEQs) were between 0.2 and 7.8 ng/L (median, 1.6 ng/L) and, thereby, were lower than those found in a pilot study, which revealed EEQs of greater than 10 ng/L in the effluents of two other STPs. The EEQs in 14 of the 16 effluent samples were very similar (0.9-3.3 ng/L), indicating a rather constant input of estrogenic substances via STPs into rivers. Additional activated charcoal filtration turned out to be very efficient in further eliminating estrogenic activity from effluents. The EEQs of the E-Screen assay and those calculated from the results of extensive chemical analysis using the estradiol equivalency factors determined for 13 natural and synthetic estrogenic substances were comparable for most of the effluent samples. 17beta-Estradiol, 17alpha-ethinylestradiol, and, to a lesser extent, estrone contributed to 90% or more of the EEQ value.

New aspects of the tolerance of retinol in humans.

A review is given of the evaluation of 517 cases of acute vitamin A intoxication and chronic hypervitaminosis A. Whereas acute vitamin A intoxication has lost numerical importance during the past 15 years, chronic hypervitaminosis-A, especially that caused by self-medication, has provoked world-wide discussion concerning the safety of retinol intake. Evaluation of the 132 cases of chronic hypervitaminosis-A showed that approximately one quarter cannot be considered as genuine cases, whereas the rest is divided almost equally between vitamin A prescription and self-medication. A significant positive correlation could be shown between the dose administered and the duration of treatment. Moreover, with comparable doses the symptoms of chronic hypervitaminosis-A appear significantly earlier (by a factor of 6) after emulsified or equivalent preparations than after oily emulsions because of their better absorbability. As soon as the liver vitamin A storage capacity is exhausted, hypervitaminosis-A symptoms appear and the liver vitamin A concentration is at least 10-fold the normal. The calculated vitamin A concentration in the liver of the individual cases of hypervitaminosis-A is highly correlated with the daily intake of vitamin A per kg of body-weight and its duration.

[Competence center for insurance sciences]. / Zum Kompetenzzentrum Versicherungswissenschaften.

The insurance companies located in Hannover have launched an initiative for a "Compentence Center for Insurance Sciences" whose participants include the Hannover Medical School (MHH/HMS), the University of Göttingen and the University of Hannover. A chair of insurance medicine has been established at the MHH/HMS, a professorship for insurance mathematics in Hannover and a professorship for insurance law in Göttingen. In cooperation with the chair of insurance economics, the above-mentioned participants are preparing to open a competence center that will operate as a limited liability company (GmbH), coordinating activities with economic relevance and providing information to encourage interdisciplinary cooperation among the university institutes and the insurance industry.

Nanoformulation improves activity of the (pre)clinical anticancer ruthenium complex KP1019.

Ruthenium anticancer drugs belong to the most promising non-platinum anticancer metal compounds in clinical evaluation. However, although the clinical results are promising regarding both activity and very low adverse effects, the clinical application is currently hampered by the limited solubility and stability of the drug in aqueous solution. Here, we present a new nanoparticle formulation based on polymer-based micelles loaded with the anticancer lead ruthenium compound KP1019. Nanoprepared KP1019 was characterised by enhanced stability in aqueous solutions. Moreover, the nanoparticle formulation facilitated cellular accumulation of KP1019 (determined by ICP-MS measurements) resulting in significantly lowered IC50 values. With regard to the mode of action, increased cell cycle arrest in G2/M phase (PI-staining), DNA damage (Comet assay) as well as enhanced levels of apoptotic cell death (caspase 7 and PARP cleavage) were found in HCT116 cells treated with the new nanoformulation of KP1019. Summarizing, we present for the first time evidence that nanoformulation is a feasible strategy for improving the stability as well as activity of experimental anticancer ruthenium compounds.