AKT/mTOR pathway activation and BCL-2 family proteins modulate the sensitivity of human small cell lung cancer cells to RAD001

PURPOSE: The Akt/mammalian target of rapamycin (mTOR) pathway is frequently activated in human cancers and plays an important role in small cell lung cancer (SCLC) biology. We investigated the potential of targeting mTOR signaling as a novel antitumor approach in SCLC. EXPERIMENTAL DESIGN: The expression of mTOR in patient specimens and in a panel of SCLC cell lines was analyzed. The effects on SCLC cell survival and downstream signaling were determined following mTOR inhibition by the rapamycin derivative RAD001 (Everolimus) or down-regulation by small interfering RNA. RESULTS: We found elevated expression of mTOR in patient specimens and SCLC cell lines, compared with normal lung tissue and normal lung epithelial cells. RAD001 treatment impaired basal and growth factor-stimulated cell growth in a panel of SCLC cell lines. Cells with increased Akt pathway activation were more sensitive to RAD001. Accordingly, a constitutive activation of the Akt/mTOR pathway was sufficient to sensitize resistant SCLC cells to the cytotoxic effect of RAD001. In the sensitive cells, RAD001 showed a strong additive effect to the proapoptotic action of the chemotherapeutic agent etoposide. Intriguingly, we observed low Bcl-2 family proteins levels in the SCLC cells with a constitutive Akt pathway activation, whereas an increased expression was detected in the RAD001-resistant SCLC cells. An antisense construct targeting Bcl-2 or a Bcl-2-specific inhibitor was able to sensitize resistant SCLC cells to RAD001. Moreover, SCLC tumor growth in vivo was significantly inhibited by RAD001. CONCLUSION: Together, our data show that inhibiting mTOR signaling with RAD001 potently disrupts growth and survival signaling in human SCLC cells.

The tumor suppressor gene hypermethylated in cancer 1 is transcriptionally regulated by E2F1

The Hypermethylated in Cancer 1 (HIC1) gene encodes a zinc finger transcriptional repressor that cooperates with p53 to suppress cancer development. We and others recently showed that HIC1 is a transcriptional target of p53. To identify additional transcriptional regulators of HIC1, we screened a set of transcription factors for regulation of a human HIC1 promoter reporter. We found that E2F1 strongly activates the full-length HIC1 promoter reporter. Promoter deletions and mutations identified two E2F responsive elements in the HIC1 core promoter region. Moreover, in vivo binding of E2F1 to the HIC1 promoter was shown by chromatin immunoprecipitation assays in human TIG3 fibroblasts expressing tamoxifen-activated E2F1. In agreement, activation of E2F1 in TIG3-E2F1 cells markedly increased HIC1 expression. Interestingly, expression of E2F1 in the p53(-/-) hepatocellular carcinoma cell line Hep3B led to an increase of endogenous HIC1 mRNA, although bisulfite genomic sequencing of the HIC1 promoter revealed that the region bearing the two E2F1 binding sites is hypermethylated. In addition, endogenous E2F1 induced by etoposide treatment bound to the HIC1 promoter. Moreover, inhibition of E2F1 strongly reduced the expression of etoposide-induced HIC1. In conclusion, we identified HIC1 as novel E2F1 transcriptional target in DNA damage responses.

Complexity of CEBPA dysregulation in human acute myeloid leukemia

The transcription factor CCAAT enhancer binding protein alpha (CEBPA) is crucial for normal development of granulocytes. Various mechanisms have been identified how CEBPA function is dysregulated in patients with acute myeloid leukemia (AML). In particular, dominant-negative mutations located either at the N- or the C terminus of the CEBPA gene are observed in roughly 10% of AML patients, either in the combination on separate alleles or as sole mutation. Clinically significant complexity exists among AML with CEBPA mutations, and patients with double CEBPA mutations seem to have a more favorable course of the disease than patients with a single mutation. In addition, myeloid precursor cells of healthy carriers with a single germ-line CEBPA mutation evolve to overt AML by acquiring a second sporadic CEBPA mutation. This review summarizes recent reports on dysregulation of CEBPA function at various levels in human AML and therapeutic concepts targeting correction of CEBPA activity. The currently available data are persuasive evidence that impaired CEBPA function contributes directly to the development of AML, whereas restoring CEBPA function represents a promising target for novel therapeutic strategies in AML.

Activation of the unfolded protein response is associated with favorable prognosis in acute myeloid leukemia

PURPOSE: The unfolded protein response is triggered by the accumulation of misfolded proteins within the endoplasmic reticulum. Previous studies suggest that the unfolded protein response is activated in some cancer cell lines and involved in tumor development. The role of the unfolded protein response during leukemogenesis is unknown thus far. EXPERIMENTAL DESIGN: Here, we assessed the induction of key effectors of the unfolded protein response in leukemic cells at diagnosis of 105 acute myeloid leukemia (AML) patients comprising all subtypes. We determined the formation of the spliced variant of the X-box-binding protein 1 (XBP1) mRNA, as well as expression levels of calreticulin, GRP78, and CHOP mRNA. RESULTS: The formation of the spliced variant of XBP1s was detectable in 16.2% (17 of 105) of AML patients. Consistent with activated unfolded protein response, this group also had significantly increased expression of calreticulin, GRP78, and CHOP. AML patients with activated unfolded protein response had lower WBC counts, lactate dehydrogenase levels, and more frequently, secondary AML. The incidence of fms-related tyrosine kinase 3 (FLT3) mutations was significantly lower in patients with activated unfolded protein response. In addition, an association was observed between activated unfolded protein response and deletion of chromosome 7. Finally, the clinical course of AML patients with activated unfolded protein response was more favorable with lower relapse rate (P = 0.0182) and better overall (P = 0.041) and disease-free survival (P = 0.022). CONCLUSIONS: These results suggest that the unfolded protein response is activated in a considerable subset of AML patients. AML patients with activated unfolded protein response present specific clinical characteristics and a more favorable course of the disease.

Tamoxifen inhibits TRPV6 activity via estrogen receptor-independent pathways in TRPV6-expressing MCF-7 breast cancer cells

The epithelial calcium channel TRPV6 is upregulated in breast carcinoma compared with normal mammary gland tissue. The selective estrogen receptor modulator tamoxifen is widely used in breast cancer therapy. Previously, we showed that tamoxifen inhibits calcium uptake in TRPV6-transfected Xenopus oocytes. In this study, we examined the effect of tamoxifen on TRPV6 function and intracellular calcium homeostasis in MCF-7 breast cancer cells transiently transfected with EYFP-C1-TRPV6. TRPV6 activity was measured with fluorescence microscopy using Fura-2. The basal calcium level was higher in transfected cells compared with nontransfected cells in calcium-containing solution but not in nominally calcium-free buffer. Basal influxes of calcium and barium were also increased. In transfected cells, 10 mumol/L tamoxifen reduced the basal intracellular calcium concentration to the basal calcium level of nontransfected cells. Tamoxifen decreased the transport rates of calcium and barium in transfected cells by 50%. This inhibitory effect was not blocked by the estrogen receptor antagonist, ICI 182,720. Similarly, a tamoxifen-induced inhibitory effect was also observed in MDA-MB-231 estrogen receptor-negative cells. The effect of tamoxifen was completely blocked by activation of protein kinase C. Inhibiting protein kinase C with calphostin C decreased TRPV6 activity but did not alter the effect of tamoxifen. These findings illustrate how tamoxifen might be effective in estrogen receptor-negative breast carcinomas and suggest that the therapeutic effect of tamoxifen and protein kinase C inhibitors used in breast cancer therapy might involve TRPV6-mediated calcium entry. This study highlights a possible role of TRPV6 as therapeutic target in breast cancer therapy.

Einfluss einer dynamischen Auftragssteuerung nach dem Floating-Batch-Prinzip in zweistufigen Kommissioniersystemen mit Sortereinsatz

Im Rahmen eines Projektes der Deutschen Forschungsgesellschaft (DFG) wurden am Lehrstuhl für Förder- und Lagerwesen der Universität Dortmund die Auswirkungen einer dynamischen Auftragssteuerung nach dem Floating-Batch-Prinzip auf die Effizienz und das Systemverhalten zweistufiger Kommissioniersysteme untersucht. Hierbei wurden Potenziale, technische und organisatorische Voraussetzungen sowie Grenzen für dessen Einsetzbarkeit abgeleitet, sowie geeignete Lösungen für bei der Umsetzung auftretende Problemstellungen ermittelt. Grundlage dieser Untersuchungen bildet eine detaillierte Analyse realisierter zweistufiger Kommissioniersysteme.

Rollout of CampusContent

CampusContent (CC) is a DFG-funded competence center for eLearning with its own portal. It links content and people who support sharing and reuse of high quality learning materials and codified pedagogical know-how, such as learning objectives, pedagogical scenarios, recommended learning activities, and learning paths. The heart of the portal is a distributed repository whose contents are linked to various other CampusContent portals. Integrated into each portal are user-friendly tools for designing reusable learning content, exercises, and templates for learning units and courses. Specialized authoring tools permit the configuration, adaption, and automatic generation of interactive Flash animations using Adobe's Flexbuilder technology. More coarse-grained content components such as complete learning units and entire courses, in which contents and materials taken from the repository are embedded, can be created with XML-based authoring tools. Open service interface allow the deep or shallow integration of the portal provider's preferred authoring and learning tools. The portal is built on top of the Enterprise Content Management System Alfresco, which comes with social networking functionality that has been adapted to accommmodate collaboration, sharing and reuse within trusted communities of practice.

Ermittlung von Berechnungsverfahren für den erzielbaren Volumenstrom von stark geneigten Schneckenförderern

Die Dimensionierung und Auslegung von Schneckenförderern zur stark geneigten Förderung von Schüttgut beruht heute noch oft auf Erfahrungswerten der Anlagenhersteller. Um eine effiziente Auslegung auf Basis gesicherter Berechnungsverfahren zu gewährleisten, läuft am Lehrstuhl für Fördertechnik Materialfluss Logistik (fml) derzeit ein von der Deutschen Forschungsgemeinschaft (DFG) gefördertes Projekt im Rahmen dessen die Schüttgutförderung in geneigten Schneckenförderern untersucht wird. Entscheidender Bestandteil der Untersuchung ist dabei die Vorhersage des Förderverhaltens im Schneckenförderer und damit die Bestimmung des erzielbaren Volumenstroms in Abhängigkeit der vorliegenden Geometrie- und Betriebsparameter. Um dies zu gewährleisten werden Daten für den erzielten Volumenstrom für eine große Anzahl verschiedener Parameterkombinationen gewonnen und anschließend statistisch ausgewertet. Durch eine Regressionsanalyse wird schließlich auf einen berechenbaren Zusammenhang geschlossen.