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Metastasis is the most lethal attribute of cancer,1 and activation of invasion and metastasis is one of the hallmarks of cancer cells.2 Proliferation, deregulated adhesion, increased invasiveness, motility, resistance to apoptotic signals, and the ability to induce angiogenesis are characteristics of metastatic cells.2 Recently, metastasis-associated in colon cancer 1 (MACC1) has been discovered as an independent prognostic indicator of metastasis formation and metastasis-free survival for colon carcinoma patients.3 MACC1 expression is upregulated in malignant colon cancer tissues compared with normal tissues, like colon mucosa or liver, and occurs mainly at the crucial step of transition from the benign to the malignant phenotype.4 It induces cell migration, invasion, proliferation, colony formation, wound healing, and hepatocyte growth factor (HGF)–triggered scattering of colon cancer cells in cell culture. In tumor xenografts it causes tumor growth and metastasis.3,5 It is also discussed that it might be involved in the regulation of apoptosis.6 Mining of databases revealed high MACC1 expression not only in colorectal cancer, but also in pancreatic, ovarian, endometrial, cervical, breast, lung, liver, esophageal, and head and neck cancers.7 Meanwhile, different tumor entities have been analyzed for MACC1 expression by quantitative real-time reverse transcription (RT ) PCR and immunohistochemistry.8 Increased MACC1 expression was shown in advanced colorectal carcinomas,9–11 peritoneal-disseminated gastric carcinomas,12 recurrent lung adenocarcinomas,13,14 and vascular invasive hepatocellular carcinomas.15,16 To the best of our knowledge, there are no data on MACC1 expression in human astrocytic tumors available yet. Diffuse low-grade astrocytomas (LGAs) of World Health Organization (WHO) grade II are well-differentiated tumors with diffuse infiltration of the adjacent brain parenchyma.17 Most of these tumors progress to anaplastic astrocytomas of WHO grade III or glioblastoma multiforme (GBM) of WHO grade IV within 4–5 years after diagnosis.18 This secondary GBM is distinguished from primary GBM, which arises de novo without detectable precursor lesion.19–21 However, GBM are the most prevalent, highly malignant, invasive, and difficult to treat primary brain tumors of adults. The treatment regimens of patients with GBM include neurosurgical tumor resection followed by local γ-irradiation and chemotherapy.17 However, in spite of multidisciplinary treatment, the median survival is only 14.6 months.22,23 Therefore, it is vital to identify new potential therapeutic targets for future therapies. Although these tumors rarely metastasize, their invasive and migratory behaviors are very similar to those of metastatic cells of tumors originating from other tissues. Thus, we hypothesized that MACC1 may also be involved in progression of human gliomas. This hypothesis was supported by the fact that HGF and its receptor c-Met have been shown to be upregulated in high-grade astrocytic tumors,24 thereby inducing migration of their cells.25–28 MACC1 binds to the Sp1 site of the c-Met promoter, controls c-Met promoter activity and expression, and thereby regulates c-Met–mediated signaling.29 Indeed, HGF/c-Met is one of the major signaling cascades regulated by MACC1 in colorectal cancer and is of major importance for both the malignant transformation and metastatic potential of colorectal tumor cells.30 Here we report that MACC1 overexpression promoted proliferation and migration of GBM cells in cell culture. Furthermore, MACC1 overexpression increased the rates of tumor formation in organotypic hippocampal slice cultures (OHSCs) derived from C57BL6/J mice, probably via upregulation of c-Met, as shown by crizotinib inhibitor experiments. In human malignant gliomas, MACC1 expression increased concomitantly with increasing WHO grade. Moreover, MACC1 expression levels allowed discrimination of dormant and recurrent LGAs and of primary and secondary GBM. Most importantly, strong MACC1 protein expression was associated with reduced patient survival. Taken together, these data indicate that MACC1 may represent a promising new biomarker for prognostication and a target for inhibition of proliferation and migration of astrocytic tumor cells.

Despite a sharp fall in GDP, German employment in terms of employees stayed remarkably robust during the Great Recession. At the same time, hours worked per employee declined significantly. This is seen as the core of the German employment ‘miracle’. A general discussion arose about the reasons behind this astonishing labour market performance and the role of short-time work (STW) as a kind of exportable panacea. In this article we look at the macroeconomic implications of STW and other measures of internal flexibility, in particular focusing on the quantification of safeguarded jobs during the crisis. We find that STW played an important role and helped to safeguard employment in Germany during the Great Recession. However, we show that other measures of internal flexibility (working time accounts, contractual arrangements on working time reductions, reduction of overtime) were equally important. Together with STW these instruments saved around 1 million jobs. To explain and understand the German success story, the features of the German core model – with a strong employer–employee relationship of mutual trust, strong employment protection, traditional standard working contracts and strong works councils at the firm level – are of key importance.

Colorectal cancer is one of the most common cancers worldwide and one of the leading causes of cancer-related death in the Western world. Tumor progression towards metastasis affects a large number of patients with colorectal cancer and seriously affects their clinical outcome. Therefore, considerable effort has been made towards the development of therapeutic strategies that can decrease or prevent colorectal cancer metastasis. Standard treatment of metastatic colorectal cancer with chemotherapy has been improved in the last 10 years by the addition of new targeted agents. The currently used antibodies bevacizumab, cetuximab and panitumumab target the VEGF and EGFR signaling pathways, which are crucial for tumor progression and metastasis. These antibodies have shown relevant efficacy in both first- and second-line treatment of metastatic colorectal cancer. Additionally, other signaling pathways, including the Wnt and HGF/Met pathways, have a well-established role in colorectal cancer progression and metastasis and constitute, therefore, promising targets for new therapeutic approaches. Several new drugs targeting these pathways, including different antibodies and small-molecule tyrosine kinase inhibitors, are currently being developed and tested in clinical trials. In this review, we summarize the new developments in this field, focusing on the inhibitors that show more promising results for use in colorectal cancer patients.

The discovery of direct cell reprogramming and induced pluripotent stem (iPS) cell technology opened up new avenues for the application of non-viral, transposon-based gene delivery systems. The Sleeping Beauty (SB) transposon is highly advanced for versatile genetic manipulations in mammalian cells. We established iPS cell reprogramming of mouse embryonic fibroblasts and human foreskin fibroblasts by transposition of OSKM (Oct4, Sox2, Klf4 and c-Myc) and OSKML (OSKM + Lin28) expression cassettes mobilized by the SB100X hyperactive transposase. The efficiency of iPS cell derivation with SB transposon system was in the range of that obtained with retroviral vectors. Co-expression of the miRNA302/367 cluster together with OSKM significantly improved reprogramming efficiency and accelerated the temporal kinetics of reprogramming. The iPS cells displayed a stable karyotype, and hallmarks of pluripotency including expression of stem cell markers and the ability to differentiate into embryoid bodies in vitro. We demonstrate Cre recombinase-mediated exchange allowing simultaneous removal of the reprogramming cassette and targeted knock-in of an expression cassette of interest into the transposon-tagged locus in mouse iPS cells. This strategy would allow correction of a genetic defect by site-specific insertion of a therapeutic gene construct into ‘safe harbor’ sites in the genomes of autologous, patient-derived iPS cells.

Purpose: We have previously identified the gene MACC1 as a strong prognostic biomarker for colorectal cancer metastasis and patient survival. Here, we report for the first time the generation of transgenic mouse models for MACC1. Experimental Design: We generated mice with transgenic overexpression of MACC1 in the intestine driven by the villin promoter (vil-MACC1) and crossed them with ApcMin mice (vil-MACC1/ApcMin). Results: vil-MACC1/ApcMin mice significantly increased the total number of tumors (P = 0.0056). This was particularly apparent in large tumors (≥3-mm diameter; P = 0.0024). A detailed histopathologic analysis of these lesions demonstrated that the tumors from the vil-MACC1/ApcMin mice had a more invasive phenotype and, consequently, showed a significantly reduced survival time than ApcMin mice (P = 0.03). Molecular analysis revealed an increased Wnt and pluripotency signaling in the tumors of vil-MACC1/ApcMin mice. Specifically, we observed a prominent upregulation of the pluripotency markers Oct4 and Nanog in these tumors compared with ApcMin controls. Finally, we could also validate that Oct4 and Nanog are regulated by MACC1 in vitro and strongly correlate with MACC1 levels in a cohort of 60 tumors of colorectal cancer patients (r = 0.7005 and r = 0.6808, respectively; P > 0.0001 and P > 0.0002, respectively). Conclusions: We provide proof of principle that MACC1-induced tumor progression in colorectal cancer acts, at least in part, via the newly discovered MACC1/Nanog/Oct4 axis. These findings might have important implications for the design of novel therapeutic intervention strategies to restrict tumor progression. Clin Cancer Res; 22(11); 2812–24. ©2016 AACR.

The aberrant activity of Wnt signaling is an early step in the transformation of normal intestinal cells to malignant tissue, leading to more aggressive tumors, and eventually metastases. In colorectal cancer (CRC), metastasis accounts for about 90% of patient deaths, representing the most lethal event during the course of the disease and is directly linked to patient survival, critically limiting successful therapy. This review focuses on our studies of the metastasis-inducing gene S100A4, which we identified as transcriptional target of β-catenin. S100A4 increased migration and invasion in vitro and metastasis in mice. In patient CRC samples, high S100A4 levels predict metastasis and reduced patient survival. Our results link pathways important for tumor progression and metastasis: the Wnt signaling pathway and S100A4, which regulates motility and invasiveness. S100A4 suppression by interdicting Wnt signaling has potential for therapeutic intervention. As proof of principle, we applied S100A4 shRNA systemically and prevented metastasis in mice. Furthermore, we identified small molecule inhibitors from high-throughput screens of pharmacologically active compounds employing an S100A4 promoter-driven reporter. Best hits act, as least in part, via intervening in the Wnt pathway and restricted metastasis in mouse models. We currently translate our findings on restricting S100A4-driven metastasis into clinical practice. The repositioned FDA-approved drug niclosamide, targeting Wnt signaling, is being tested in a prospective phase II clinical trial for treatment of CRC patients. Our assay for circulating S100A4 transcripts in patient blood is used to monitor treatment success.

Background Colorectal cancer is a preventable disease if caught at early stages. This disease is highly aggressive and has a higher incidence in African Americans. Several biomarkers and mutations of aggressive tumor behavior have been defined such as metastasis-associated in colon cancer 1 (MACC1) that was associated with metastasis in colorectal cancer patients. Here, we aim to assess colon tissue MACC1 protein and circulating MACC1 transcripts in colon preneoplastic and neoplastic African American patients. Methods Patients’ tissue samples (n = 143) have been arranged on three tissue microarrays for normal (n = 26), adenoma (n = 68) and cancer (n = 49) samples. Immunohistochemistry was used to detect MACC1 expression. Blood samples (n = 93) from normal (n = 45), hyperplastic (n = 15) and tubular adenoma (n = 33) patients were used to assess MACC1 transcripts using qRT-PCR. Distribution of continuous variables was tested between different diagnoses with Kruskal–Wallis test. Categorical variables were tested by Chi square test. We assessed the prognostic ability of IHC staining by calculating area under receiver operating characteristics curve (ROC) for adenoma and cancer separately. Differences between groups in terms of MACC1 transcript levels in plasma were calculated by using non-parametric (exact) Wilcoxon-Mann–Whitney tests. We performed all calculations with SPSS, version 21. Results In patient tissues, there was a statistically significant difference in MACC1 expression in normal vs. adenoma samples (p = 0.004) and normal vs. cancer samples (p

This paper considers the use of short-time work (STW) schemes as a device to mitigate the negative effects of the current global recession on employment levels. STW schemes have been regarded as a popular and successful counter-cyclical policy to maintain workers in employment. However, by comparing and contrasting the experience of Germany and Italy, we argue that the effectiveness of STW schemes should only be evaluated in relation to (i) the institutional set-up within which they operate, and (ii) the whole set of labour market policies adopted by each country.

The increasing unraveling of the molecular basis of cancer offers manifold novel options for intervention strategies. However, the discovery and development of new drugs for potential clinical applications is a tremendously time-consuming and costly process. Translating a novel lead candidate compound into an approved clinical drug takes often more than a decade, and the success rate is very low due to versatile efforts including defining its pharmacokinetics, pharmacodynamics, side effects as well as lack of sufficient efficacy. Thus, strategies are needed to minimize time and costs, while maximizing success rates. A very attractive strategy for novel cancer therapeutic options is the repositioning of already approved drugs. These medicines, approved for the treatment of non-malignant disorders, have already passed some early costs and time, have been tested in humans and are ready for clinical trials as anti-cancer drugs. Here we discuss the repositioning of nonsteroidal anti-inflammatory drugs (NSAID), statins, anti-psychotic drugs, anti-helminthic drugs and vitamin D as anti-tumor agents. We focus on their novel actions and potential for inhibition of cancer growth and metastasis by interfering with target molecules and pathways, which drive these malignant processes. Furthermore, important pre-clinical and clinical data are reviewed herein, which elucidate their therapeutic mechanisms which enable their repositioning for cancer therapy and disruption of metastasis.

Bacterial toxins are known to be effective for cancer therapy. Clostridium perfringens enterotoxin (CPE) is produced by the bacterial Clostridium type A strain. The transmembrane proteins claudin-3 and -4, often overexpressed in numerous human epithelial tumors (for example, colon, breast, pancreas, prostate and ovarian), are the targeted receptors for CPE. CPE binding to them triggers formation of membrane pore complexes leading to rapid cell death. In this study, we aimed at selective tumor cell killing by CPE gene transfer. We generated expression vectors bearing the bacterial wild-type CPE cDNA (wtCPE) or translation-optimized CPE (optCPE) cDNA for in vitro and in vivo gene therapy of claudin-3- and -4-overexpressing tumors. The CPE expression analysis at messenger RNA and protein level revealed more efficient expression of optCPE compared with wtCPE. Expression of optCPE showed rapid cytotoxic activity, hightened by CPE release as bystander effect. Cytotoxicity of up to 100% was observed 72 h after gene transfer and is restricted to claudin-3-and -4-expressing tumor lines. MCF-7 and HCT116 cells with high claudin-4 expression showed dramatic sensitivity toward CPE toxicity. The claudin-negative melanoma line SKMel-5, however, was insensitive toward CPE gene transfer. The non-viral intratumoral in vivo gene transfer of optCPE led to reduced tumor growth in MCF-7 and HCT116 tumor-bearing mice compared with the vector-transfected control groups. This novel approach demonstrates that CPE gene transfer can be employed for a targeted suicide gene therapy of claudin-3- and -4-overexpressing tumors, leading to the rapid and efficient tumor cell killing in vitro and in vivo.

Calcimycin restricts Wnt/β-catenin–regulated S100A4 expression, leading to reduced S100A4-mediated cell migration and invasion in colon cancer cells, as well as to inhibition of metastasis formation in xenografted mice., The calcium-binding protein S100A4 is a central mediator of metastasis formation in colon cancer. S100A4 is a target gene of the Wnt/β-catenin pathway, which is constitutively active in the majority of colon cancers. In this study a high-throughput screen was performed to identify small-molecule compounds targeting the S100A4-promoter activity. In this screen calcimycin was identified as a transcriptional inhibitor of S100A4. In colon cancer cells calcimycin treatment reduced S100A4 mRNA and protein expression in a dose- and time-dependent manner. S100A4-induced cellular processes associated with metastasis formation, such as cell migration and invasion, were inhibited by calcimycin in an S100A4-specific manner. Calcimycin reduced β-catenin mRNA and protein levels despite the expression of Δ45-mutated β-catenin. Consequently, calcimycin inhibited Wnt/β-catenin pathway activity and the expression of prominent β-catenin target genes such as S100A4, cyclin D1, c-myc, and dickkopf-1. Finally, calcimycin treatment of human colon cancer cells inhibited metastasis formation in xenografted immunodeficient mice. Our results demonstrate that targeting the expression of S100A4 with calcimycin provides a functional strategy to restrict cell motility in colon cancer cells. Therefore calcimycin may be useful for studying S100A4 biology, and these studies may serve as a lead for the development of treatments for colon cancer metastasis.

We have previously reported that over-expression of a panel of 119 genes correlates with the metastatic potential of pancreatic carcinoma cells. We sought to identify and functionally characterize candidate tumour metastasis promoting genes among this library using a secondary phenotype-assisted screen. Here we report the discovery of the metastasis-promoting function of a hitherto not characterized gene located on chromosome 14 (ORF138), which we have named 'novel metastasis-promoting gene 1' (NVM-1). The NVM-1 transcript is extensively alternatively spliced, is expressed endogenously in a number of different tissues, and is strongly over-expressed at the protein level in a variety of human tumour types. Importantly, NVM-1 expression stimulates the migratory and invasive behaviour of tumour cells and promotes metastasis formation in experimental animals in vivo. Up-regulation of FMNL2 and MT1E and down-regulation of TIMP4 and MHC-I is observed as a consequence of NVM-1 expression. Together these data identify NVM-1 as a gene that is functionally involved in tumour metastasis, and suggest that NVM-1 may constitute a promising therapeutic target for inhibition of tumour metastasis.

Early detection of tumors and metastases is critical for improving treatment strategies and patient outcomes. The development of molecular markers and simple tests that are clinically applicable for detection, prognostication, and therapy monitoring is strongly needed. The gene S100A4 has long been known to act as a metastasis inducer. High S100A4 levels in the primary tumor are prognostic for metachronous metastasis and correlate with reduced patient survival. We provide, for the first time, a plasma-based assay for transcript quantification of S100A4 in gastrointestinal patients' plasma. We conducted a study to define the diagnostic and prognostic power of S100A4 transcripts using 466 plasma samples from colon, rectal, and gastric cancer patients. Plasma was separated, RNA was isolated, and S100A4 mRNA was determined by quantitative RT-PCR. S100A4 transcripts were increased in cancer patients of each entity (P < 0.0001) and all disease stages (P < 0.05), compared with tumor-free volunteers (sensitivities of 96%, 74%, and 90% and specificities of 59%, 82%, and 71%, for colon, rectal, and gastric cancer patients, respectively). Prospectively analyzed follow-up patients who later experienced metastasis showed higher S100A4 levels than follow-up patients without metastasis. Disease-free survival was decreased in high S100A4-expressing follow-up colorectal cancer patients (P = 0.013). In summary, we developed a method for quantitative S100A4 transcript determination in plasma that allows clinical application routinely. We demonstrated the diagnostic and prognostic potential of this method for early defining cancer staging and patients' risk for metastasis.

Colon cancer metastasis is often associated with activation of the Wnt/β-catenin signaling pathway and high expression of the metastasis mediator S100A4. We previously demonstrated the transcriptional regulation of S100A4 by β-catenin and the importance of the interconnection of these cellular programs for metastasis. Here we probe the hypothesis that the nonsteroidal anti-inflammatory drug sulindac sulfide can inhibit colon cancer metastasis by intervening in β-catenin signaling and thereby interdicting S100A4. We treated colon cancer cell lines heterozygous for gain-of-function and wild-type β-catenin with sulindac. We analyzed sulindac's effects on β-catenin expression and subcellular localization, β-catenin binding to the T-cell factor (TCF)/S100A4 promoter complex, S100A4 promoter activity, S100A4 expression, cell motility, and proliferation. Mice intrasplenically transplanted with S100A4-overexpressing colon cancer cells were treated with sulindac. Tumor growth and metastasis, and their β-catenin and S100A4 expressions, were determined. We report the expression knockdown of β-catenin by sulindac, leading to its reduced nuclear accumulation. The binding of β-catenin to TCF was clearly lowered, resulting in reduced S100A4 promoter activity and expression. This correlated well with the inhibition of cell migration and invasion, which could be rescued by ectopic S100A4 expression. In mice, sulindac treatment resulted in reduced tumor growth in the spleen (P = .014) and decreased liver metastasis in a human colon cancer xenograft model (P = .025). Splenic tumors and liver metastases of sulindac-treated mice showed lowered β-catenin and S100A4 levels. These results suggest that modulators of β-catenin signaling such as sulindac offer potential as antimetastatic agents by interdicting S100A4 expression.

IDH1-mutation and MGMT-promoter methylation are established prognostic markers in Glioblastoma multiforme (GBM); however, invasive procedures are needed to obtain the required tumor tissue. Metastasis-associated in colon cancer-1 (MACC1) can easily be measured in peripheral blood and has been established as a prognostic plasma marker for several solid cancer entities. Further, MACC1 has been identified by us and others as a promising biomarker for prognostication for GBM, when detected in the tumor tissue. Here, we evaluated whether circulating MACC1 transcripts could be useful to predict clinical outcome and response to therapy in GBM. Plasma samples were collected from GBM patients (IDH1 wildtype (wt), IDH1R132H, and unspecified) before surgery. Samples from healthy volunteers served as controls. MACC1-transcript levels, determined by qRT-PCR, were correlated with patient overall survival (OS) and clinical data of known prognostic significance. Cut-off values for Kaplan-Meier analyses were determined by ROC calculations. Cluster analyses were performed. MACC1 transcripts were higher in plasma of GBM patients compared to healthy controls (P The inclusion of circulating MACC1 transcript levels into the diagnostic workup might improve the accuracy of outcome prediction and thus help to define more precise risk categories of GBM patients. Whether pharmaceutical inhibition of MACC1 expression might be of general benefit for the treatment response and survival of GBM patients remains to be determined. Citation Format: Carsten Hagemann, Nikolas Neuhaus, Mathias Dahlmann, Almut F. Kessler, Dennis Kobelt, Pia Herrmann, Matthias Eyrich, Benjamin Freitag, Thomas Linsenmann, Camelia Monoranu, Ralf-Ingo Ernestus, Mario Löhr, Ulrike S. Stein. Plasma levels of circulating MACC1 transcripts co-segregate with other prognostic markers and treatment response in glioblastoma patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5583.

e14536Background: Patients with metastasized colorectal cancer (mCRC) progressing under standard therapy have poor prognosis and need further treatment alternatives. The S100 calcium binding protei...

Peritoneal metastasis (PM) represents a terminal tumor stage with limited therapy options for patients with colorectal cancer (CRC). To improve therapeutic efficacy and overall survival, availability of appropriate in vivo models of PM could improve the evaluation of chemosensitivity as well as identification of novel biomarkers and therapeutic targets to improve the therapeutic outcome of PM patients. In this context, we generated the first patient-derived xenograft (PDX) models of PM from CRC as platform to test chemotherapy response and to potentially analyze biomarkers. For the PDX establishment colorectal surgical specimens were subcutaneously (s.c.) transplanted onto immunocompromised NOG mice. The engrafted tumors were transferred to NMRI nu/nu mice for further passaging. They were characterized by histopathology, immunohistochemistry, and gene expression analyses using real-time RT-PCR. Chemosensitivity of PDX models was evaluated in vivo by application of a panel of conventional chemotherapeutic and of targeted drugs. For PDX establishment 68 CRC surgical specimens (taken from peritoneum and omentum) were transplanted, from which 22 PDX have engrafted. From those, 13 models of 10 patients have been analyzed. Their tumor doubling times ranged between 4 to 28 days. The histopathologic evaluation revealed maintenance of the original CRC histology in the PDX. The chemosensitivity testing of conventional and of targeted drugs in the 13 PM PDX models revealed the individual, diverging response of the PDX, such as for 5-FU, irinotecan, oxaliplatin, cetuximab, and erlotinib. More interestingly, different responses were observed in PDX of the omentum vs. peritoneum, which were derived from the same patient. Our results demonstrate that this novel panel of PDX maintains the morphology of the patient tumor in early passages, reflect heterogeneous response rates, and can be used as a preclinical in vivo platform for translational studies of potential clinical use. In conclusion, this PDX panel of PM of the CRC can be used for further studies to evaluate new biomarkers and to test novel therapies or combinations for PM of CRC. Citation Format: Wolfgang Walther, Eva Pachmayr, Bernadette Brzezicha, Britta Büttner, Beate Rau, Ulrike Stein. Novel patient-derived xenograft (PDX) models from peritoneal metastasis of colorectal carcinoma for drug testing and biomarker analysis [abstract]. In: Proceedings of the AACR Special Conference: Advances in Modeling Cancer in Mice: Technology, Biology, and Beyond; 2017 Sep 24-27; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(10 Suppl):Abstract nr A17.

Nachdem die Immobilienblase geplatzt war, hat sich die grose Rezession rasch uber Europa ausgebreitet. Deutschland, Italien und das Vereinigte Konigreich erlebten daraufhin fast einen gleich grosen Einbruch im Bruttoinlandsprodukt. Gleichzeitig blieb der Anstieg der Arbeitslosigkeit aber deutlich hinter den Erfahrungen vergangener Krisen zuruck. Dennoch sind die Lander unterschiedlich gut durch die Krise gekommen. Wahrend man mit Blick auf Deutschland vom Jobwunder spricht, ist die Arbeitslosigkeit im Vereinigten Konigreich trotz masigen Anstiegs so hoch wie seit Jahren nicht mehr. Italien belegt eine mittlere Position. Wie kann man diese Unterschiede erklaren? Im Vereinigten Konigreich spielte die Aktive Arbeitsmarktpolitik so gut wie keine Rolle. Ganz anders in Italien, wo Kurzarbeit eine lange Tradition hat. Sie wurde in der Krise wieder intensiv genutzt, doch es wird deutlich, dass sie kein Allheilmittel ist. Anders in Deutschland: Hier steht den Unternehmen gleichzeitig eine Vielzahl von Instrumenten der internen Flexibilitat zur Verfugung, die die Betriebe erganzend zur Kurzarbeit nutzen konnen. Gerade dieses Zusammenspiel scheint nun in der Krise den entscheidenden Unterschied auszumachen.

Therapy-inducible vectors are useful for conditional expression of therapeutic genes in gene therapy, which is based on the control of gene expression by conventional treatment modalities. By this approach, combination of chemotherapy, radiation or hyperthermia with gene therapy can result in considerable, additive or synergistic improvement of therapeutic efficacy. This concept has been successfully tested in particular for gene therapy of cancer. The identification of efficient heat-responsive gene promoters provided the rationale for heat-regulated gene therapy. The objective of this review is to provide insights into the cellular mechanisms of heat-shock response, as prerequisite for therapeutic actions of hyperthermia and into the field of heat-responsive gene therapy. Furthermore, the major strategies of heat-responsive gene therapy systems in particular for cancer treatment are summarized. The developments for heat-responsive vector systems for in vitro and in vivo approaches are discussed. This review will provide an overview for this gene therapy strategy and its potential for multimodal therapeutic concepts in the clinic.

Background & Aims Much is known about the genes and mutations that cause colorectal cancer (CRC), yet only a few have been associated with CRC metastasis. We performed expression-profiling experiments to identify genetic markers of risk and to elucidate the molecular mechanisms of CRC metastasis. Methods We compared gene expression patterns between metastatic and nonmetastatic stage-matched human colorectal carcinomas by microarray analysis. Correlations between BAMBI and metastasis-free survival were examined by quantitative real-time polymerase chain reaction (PCR) using an independent set of human colon carcinomas. Human colon cancer cell lines were analyzed for BAMBI regulation, cell motility, and experimental metastasis. Results We established a signature of 115 genes that differentiated metastatic from nonmetastatic primary tumors. Among these, the transforming growth factor (TGF) β inhibitor BAMBI was highly expressed in approximately half of metastatic primary tumors and metastases but not in nonmetastatic tumors. BAMBI is a target of canonical Wnt signaling that involves the β-catenin coactivator BCL9-2. We observed an inverse correlation between level of BAMBI expression and metastasis-free survival time of patients. BAMBI inhibits TGF-β signaling and increases migration in colon cancer cells. In mice, overexpression of BAMBI caused colon cancer cells to form tumors that metastasized more frequently to liver and lymph nodes than control cancer cells. Conclusions BAMBI regulates CRC metastasis by connecting the Wnt/β-catenin and TGF-β-signaling pathways. The metastatic expression signature we describe, along with BAMBI levels, can be used in prognosis. Developmental signaling pathways appear to act in hierarchies and cooperate in tumor cell migration, invasion, and metastasis.

We identified a previously undescribed gene associated with colon cancer by genome-wide expression analysis in primary and metastatic carcinomas: metastasis-associated in colon cancer-1, MACC1. MACC1 expression in tumor specimens is an independent prognostic indicator of metastasis formation and metastasis-free survival. We show that the gene encoding the hepatocyte growth factor (HGF) receptor, MET, is a transcriptional target of MACC1. MACC1 promotes proliferation, invasion and HGF-induced scattering of colon cancer cells in cell culture and tumor growth and metastasis in mouse models. These phenotypes are lost in cells expressing MACC1 mutants lacking the SH3 domain or the proline-rich motif. For clinical practice, MACC1 will be useful for the identification of poor prognosis subjects with colorectal cancer and is a promising new target for intervention in metastasis formation.

Purpose: This phase I clinical trial evaluated safety, feasibility, and efficiency of nonviral intratumoral jet-injection gene transfer in patients with skin metastases from melanoma and breast cancer. Experimental Design: Seventeen patients were enrolled. The patients received five jet injections with a total dose of 0.05 mg β-galactosidase (LacZ)-expressing plasmid DNA (pCMVβ) into a single cutaneous lesion. Clinical and laboratory safety monitoring were done. Systemic plasmid clearance was monitored by quantitative real-time PCR of blood samples throughout the study. All lesions were resected after 2 to 6 days. Intratumoral plasmid DNA load, DNA distribution, and LacZ expression was analyzed by quantitative real-time PCR, quantitative reverse transcription-PCR, Western blot, immunohistochemistry, and 5-bromo-4-chloro-3-indolyl-β-d-galactoside staining. Results: Jet injection of plasmid DNA was safely done in all patients. No serious side effects were observed. Thirty minutes after jet injection, peak plasmid DNA levels were detected in the blood followed by rapid decline and clearance. Plasmid DNA and LacZ mRNA and protein expression were detected in all treated lesions. Quantitative analysis revealed a correlation of plasmid DNA load and LacZ-mRNA expression confirmed by Western blot. Immunohistochemistry and 5-bromo-4-chloro-3-indolyl-β-d-galactoside staining showed LacZ-protein throughout the tumor. Transfected tumor areas were found close and distant to the jet-injection site with varying levels of DNA load and transgene expression. Conclusion: Intratumoral jet injection of plasmid DNA led to efficient LacZ reporter gene expression in all patients. No side effects were experienced, supporting safety and applicability of this novel nonviral approach. A next step with a therapeutic gene product should determine antitumor efficacy of jet-injection gene transfer.

Multidrug resistance (MDR) is a major cause for cancer chemotherapy failure. Among the numerous strategies to overcome persistent action of proinflammatory cytokines, such as tumor necrosis factor α (TNF-α) permits downregulation of MDR-associated genes, including ATP-binding cassette, subfamily B 1 gene (ABCB1). A key regulator of ABCB1 expression is the transcription factor nuclear factor κ light chain enhancer (NF-κB)/p65. We analyzed diverging short- and long-term effects of TNF-α regarding modulation of NF-κB/p65 signaling and ABCB1 expression in colon cancer cells. Highly resistant ABCB1 overexpressing human HCT15 colorectal carcinoma cells were subjected to short- (30-120 min) or long-term (24-96 h) TNF-α treatment. TNF-α mediated modulation of ABCB1 expression was analyzed by real-time RT-PCR and western blot analysis. The TNF-mediated chemosensitization was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cytotoxicity assay. The involvement of TNF receptors and of NF-κB/p65 signaling was analyzed by western blot analysis, ABCB1 promoter analysis and electrophoretic mobility shift assay (EMSA). The study revealed, that long-term, but not short-term TNF-α treatment leads to TNF-receptor 1 (TNFR1) mediated downregulation of ABCB1 resulting in sensitization towards drug treatment. It dampens NF-κB/p65 activation and nuclear factor of κ light polypeptide gene enhancer in B-cells inhibitor α (IκBα) resynthesis, associated with reduced nuclear accumulation of NF-κB/p65 and reduced binding to its consensus sequence in the ABCB1 promoter. The study reveals the diverging effects of short- or long-term TNF-α action and provides novel insights on downregulation of ABCB1 expression by TNF-mediated repression of NF-κB signaling.

The suppression of genes involved in tumor progression, metastasis formation, or therapy resistance by RNA interference is a promising tool to treat cancer disease. Efficient delivery of interfering molecules and their sustained presence in tumor cells are required for therapeutic success. This chapter describes a method of systemic application of shRNA expression plasmid via tail vein injection in xenograft mice, causing the sustained reduction of target gene expression in the primary tumor. By choosing S100A4 as a metastasis driving target gene, this therapeutic approach restricted the formation of distant colorectal cancer metastases after intrasplenic transplantation. In vivo imaging of bioluminescent cancer cells allows the monitoring of tumor growth and metastasis formation over time. End point analysis of the trial included scoring of the metastatic burden and the quantification of target gene expression in the tumor. Average S100A4 expression in tumor tissues was reduced by 30 %, causing a 70 % decrease of liver metastases.

Expression of the small calcium-binding protein S100A4 is associated with poor prognosis in patients with thyroid cancer (TC). The authors have previously shown that S100A4 is a target for relaxin and insulin-like peptide 3 signaling in TC cells and that S100A4 is secreted from human TC cells. Although the pro-migratory role of intracellular S100A4 in binding to non-muscle myosin is well known, this study investigated here whether extracellular S100A4 contributes to TC migration.Human cell lines of follicular, papillary, and undifferentiated thyroid cancer, primary patient TC cells, and TC tissues were utilized to discover the presence of the receptor of advanced glycation end products (RAGE) in TC cells and TC tissues. Fluorescence imaging, protein pull-down assays, Western blot, siRNA protein silencing, small GTPase inhibitors, cell proliferation, and cell migration assays were used to investigate the interaction of extracellular S100A4 with RAGE in promoting a TC migratory response.It was demonstrated that RAGE served as receptor for extracellular S100A4 mediating cell migration in TC cells. The RAGE-mediated increase in cell migration was dependent on the intracellular RAGE signaling partner diaphanous-1 (Dia-1) and involved the activation of the small GTPases Cdc42 and RhoA. Although extracellular S100A4 consistently activated ERK signaling in TC cells, it was shown that ERK signaling was not mediated by RAGE and not essential for the migratory response in TC cells.The data have identified the RAGE/Dia-1 signaling system as a mediator for the pro-migratory response of extracellular S100A4 in human TC. Thus, therapeutic targeting of the RAGE/Dia-1/small GTPases signaling may successfully reduce local invasion and metastasis in TC.

MACC1 (metastasis associated in colon cancer 1) is a prognostic biomarker for tumor progression, metastasis and survival of a variety of solid cancers including colorectal cancer (CRC). Here we aimed to identify the MACC1-induced transcriptome and key players mediating the MACC1-induced effects in CRC. We performed microarray analyses using CRC cells ectopically overexpressing MACC1. We identified more than 1300 genes at least twofold differentially expressed, including the gene SPON2 (Spondin 2) as 90-fold upregulated transcriptional target of MACC1. MACC1-dependent SPON2 expression regulation was validated on mRNA and protein levels in MACC1 high (endogenously or ectopically) and low (endogenously or by knockdown) expressing cells. Chromatin immunoprecipitation analysis demonstrated the binding of MACC1 to the gene promoter of SPON2. In cell culture, ectopic SPON2 overexpression induced cell viability, migration, invasion and colony formation in endogenously MACC1 and SPON2 low expressing cells, whereas SPON2 knockdown reduced proliferative, migratory and invasive abilities in CRC cells with high endogenous MACC1 and SPON2 expression. In intrasplenically transplanted NOD/SCID mice, metastasis induction was analyzed with control or SPON2-overexpressing CRC cells. Tumors with SPON2 overexpression induced liver metastasis (vs control animals without any metastases, P=0.0036). In CRC patients, SPON2 expression was determined in primary tumors (stages I-III), and survival time was analyzed by Kaplan-Meier method. CRC patients with high SPON2 expressing primary tumors demonstrated 8 months shorter metastasis-free survival (MFS) compared with patients with low SPON2 levels (P=0.053). Combining high levels of SPON2 and MACC1 improved the identification of high-risk patients with a 20-month shorter MFS vs patients with low biomarker expression. In summary, SPON2 is a transcriptional target of the metastasis gene MACC1. SPON2 induces cell motility in vitro and CRC metastasis in mice. In patients, SPON2 serves as prognostic indicator for CRC metastasis and survival, and might represent a promising target for therapeutic approaches.

Aim: To evaluate the diagnostic and prognostic value of circulating Metastasis Associated in Colon Cancer 1 (MACC1) transcripts in plasma of gastric cancer patients. Methods: We provide for the first time a blood-based assay for transcript quantification of the metastasis inducer MACC1 in a prospective study of gastric cancer patient plasma. MACC1 is a strong prognostic biomarker for tumor progression and metastasis in a variety of solid cancers. We conducted a study to define the diagnostic and prognostic power of MACC1 transcripts using 76 plasma samples from gastric cancer patients, either newly diagnosed with gastric cancer, newly diagnosed with metachronous metastasis of gastric cancer, as well as follow-up patients. Findings were controlled by using plasma samples from 54 tumor-free volunteers. Plasma was separated, RNA was isolated, and levels of MACC1 as well as S100A4 transcripts were determined by quantitative RT-PCR. Results: Based on the levels of circulating MACC1 transcripts in plasma we significantly discriminated tumor-free volunteers and gastric cancer patients (P < 0.001). Levels of circulating MACC1 transcripts were increased in gastric cancer patients of each disease stage, compared to tumor-free volunteers: patients with tumors without metastasis (P = 0.005), with synchronous metastasis (P = 0.002), with metachronous metastasis (P = 0.005), and patients during follow-up (P = 0.021). Sensitivity was 0.68 (95%CI: 0.45-0.85) and specificity was 0.89 (95%CI: 0.77-0.95), respectively. Importantly, gastric cancer patients with high circulating MACC1 transcript levels in plasma demonstrated significantly shorter survival when compared with patients demonstrating low MACC1 levels (P = 0.0015). Furthermore, gastric cancer patients with high circulating transcript levels of MACC1 as well as of S100A4 in plasma demonstrated significantly shorter survival when compared with patients demonstrating low levels of both biomarkers or with only one biomarker elevated (P = 0.001). Conclusion: Levels of circulating MACC1 transcripts in plasma of gastric cancer patients are of diagnostic value and are prognostic for patient survival in a prospective study.

The suppression of genes involved in tumor progression, metastasis formation, or therapy resistance by RNA interference is a promising tool to treat cancer disease. Efficient delivery of interfering molecules and their sustained presence in tumor cells are required for therapeutic success. This chapter describes a method of systemic application of shRNA expression plasmid via tail vein injection in xenograft mice, causing the sustained reduction of target gene expression in the primary tumor. By choosing S100A4 as a metastasis driving target gene, this therapeutic approach restricted the formation of distant colorectal cancer metastases after intrasplenic transplantation. In vivo imaging of bioluminescent cancer cells allows the monitoring of tumor growth and metastasis formation over time. End point analysis of the trial included scoring of the metastatic burden and the quantification of target gene expression in the tumor. Average S100A4 expression in tumor tissues was reduced by 30 %, causing a 70 % decrease of liver metastases.

Nonviral jet injection is an applicable technology for in vivo gene transfer of naked DNA. However, little is known about the biodistribution and clearance of jet-injected DNA, or about its localization within tissue and cells. Therefore, in this study we analyzed the intratumoral and systemic biodistribution of jet-injected naked DNA in human colon carcinoma-bearing NCr-nu/nu mice, which were jet-injected with the pCMVbeta plasmid DNA. Intratumoral and systemic plasmid DNA biodistribution was analyzed 5, 10, 20, and 40 min and 3, 6, 24, 48, and 72 hr after jet injection, using quantitative real-time polymerase chain reaction. In the tumors, a rapid drop in naked DNA load within 24 hr of jet injection was shown. Detailed analysis of intratumoral distribution of rhodamine-labeled DNA revealed the presence of plasmid DNA within tumor cells 5 min after jet injection and further accumulation of significant DNA amounts in the cell nuclei 30 to 60 min after jet injection. In the blood, DNA amounts rapidly dropped within 10 to 40 min of jet injection to less than 0.001 pg of plasmid per 250 ng of tissue DNA and only minimal plasmid DNA dissemination was detected in liver, lung, spleen, kidney, and ovaries, which was cleared 3 to 6 hr after jet injection. By contrast, in heart, bone marrow, and brain almost no plasmid DNA was detectable.

Multidrug resistance (MDR) remains the major cause of failure of chemotherapeutic treatment in cancer therapy. Thus, reversal of MDR in cancer is a desperately needed clinical requirement and a scientific challenge. The MDR phenotype can be mediated by adenosine triphosphate (ATP)-binding cassette (ABC) transporter molecules, which lower the intracellular accumulation of an entire panel of structurally and functionally unrelated anticancer drugs. Detailed knowledge of the transporter molecules, their sequence polymorphisms, their regulation at the transcriptional, translational, and post-translational levels, as well as their functional characteristics for substrate binding and transport, represent the basis for specific and, thus, successful intervention strategies for reversal of ABC transporter-dependent MDR in the clinic. Initial data obtained with third-generation inhibitors of ABC transporter-dependent MDR in clinical trials, and new developments with tools that interfere with expression of MDR genes, thereby leading to restored chemosensitization, are very promising attempts that provide grounds for cautious optimism. This article presents an overview of different strategies for overcoming ABC transporter-dependent MDR. Approaches targeting the function as well as the expression of MDR-related ABC transporters are summarized. Clinical trials evaluating the impact of compounds for reversal of ABC transporter-dependent MDR are discussed. The question remains, is reversal of MDR in cancer truly an achievable goal, given the different tumor entities and tumor heterogeneity, different tumor resistance profiles, and different therapeutic regimens? If so, MDR reversal will probably be successful only when tailored to an individual patient with an individual tumor, resistance profile and response prediction, i.e. individual prevention, circumvention, and reversal of MDR. Tailor-made MDR reversal strategies are the ultimate goal — and probably a realistic option.

In our previous papers we described the ability of methylxanthine pentoxifylline (PTX) to depress the P-glycoprotein (P-gp) mediated multidrug resistance (MDR) of the mouse leukemic cell line L1210/VCR. Other methylxanthines like caffeine and theophylline were found to be ineffective in this respect. In the present paper we have analysed the capability of 25 methylxanthines to depress MDR of L1210/VCR cells. These methylxanthines structurally differ in substituents located in positions N1, N3, N7 and C8. The results indicate that for an effective reversal of P-gp mediated MDR of our cells the existence of a longer polar substituent in the position N1 plays a crucial role. The elongation of the substituent in the positions N3 and N7 (from methyl to propyl) increases and in the position C8 (from H to propyl) decreases the efficacy of xanthines to reverse the vincristine resistance of L1210/VCR cells. The multiple linear regression for effectiveness of methylxanthines in reversal of P-gp mediated MDR of L1210/VCR cells (expressed as respective IC(50r) values) has been computed, with molar weight: M(w), molar volume: V(M), molar refractivity: R(M), crystal density: d and partition coefficient n-octanol/water: logP as descriptors. A high intercorrelation of M(W), V(M) and R(M) was found for the tested group of methylxanthines indicating that only one of these parameters is necessary for testing a potential correlation. The best fit in the multiple linear regression was obtained for R(M) applied together with d and logP and resulted in a QSAR model given by the following equation: IC(50r)=-[(32.3+/-7.2)x10(-3)xR(M)]+[(10.1+/-2.3)xd]+[(0.74+/-0.10)xlogP]-[10.5+/-3.2]. Model revealed that: (i) the molar refractivity influences the effectiveness of xanthine positively; (ii) the crystal density and partition coefficient influence the MDR reversal effectiveness of xanthine negatively.

The activation of the RET signaling pathway during embryogenesis is a crucial prerequisite for a directional migration of enteric nervous system progenitor cells. Loss-of-function germline mutations of the RET proto-oncogene are reported in familial and sporadic cases of Hirschsprung disease (HSCR) with a variable frequency. Furthermore, variants of several RET polymorphisms are over- or under-represented in HSCR populations. Specifically, the c.135A RET variant has been previously shown to be strongly associated with the HSCR phenotype. We have reported an HSCR-phenotype modifying effect of the RET c.135G>A polymorphism due to a within-gene interaction in patients harboring RET germline mutations, yet the function of the c.135G>A variant is unknown. The basic RET promoter region was investigated by DNA sequencing approach in 80 HSCR patients. Identified polymorphisms were genotyped in the HSCR and in a control population and haplotypes were reconstructed. The dual-luciferase assay was used to evaluate the activity of different RET promoter haplotypes. We demonstrate that variants of two RET promoter polymorphisms -5G>A and -1C>A from the transcription start site are associated with HSCR. Furthermore, the -5G>A polymorphism is in strong linkage disequilibrium with the c.135G>A polymorphism. The promoter haplotype -5/-1AC associated with HSCR has a significantly lower activity in an in vitro dual-luciferase expression assay compared with those haplotypes identified in the majority of normal controls. These data suggest a role for RET haplotypes containing the -5A promoter variant in the etiology of HSCR.

Aims: To investigate the prognostic significance of chromosomal alterations in colorectal cancer patients. Histopathological tumour classification is still considered to be the gold standard for the characterization of solid tumours. However, it is well known that such established parameters do not satisfactorily predict the clinical outcome in individual cases. Markers that reliably predict survival are needed. These markers should guide the clinical treatment of neoplastic disease. Methods and results: Chromosomal imbalances in 61 colorectal carcinoma specimens in 37 patients determined by comparative genomic hybridization were correlated with patient survival using custom-made computer software which enabled the assessment of individual chromosomal loci. Kaplan–Meier analysis revealed that over-representations of 2p14-15, 6q23-6q24, 15q22-15q23, 22q11.2 and deletions of 1p36.1-36.2, 4q31.3, 4q35, 8q12-q21, 8p11.2 and 9p22 were significantly associated with shorter disease-specific survival, whereas over-expression of 20q13.3 and deletion of 18q11.2 were significantly associated with longer disease-specific survival in this collection of colorectal cancers. Multivariate Cox proportional hazards regression models consistently identified gains of 2p14-15, 15q22-23, 22q11.2 and losses of 1p36.1-36.2 and 4q35 as independent markers of shorter patient survival carrying greater significance than the classical clinicopathological parameters of nodal status and tumour grade. Conclusions: These five markers allow a molecular categorization of patients into high and low clinical risk groups. Thus, the genomic data have refined the histopathological classification highlighting the necessity for a supplementary genetically based stratification of colorectal cancer.

Jet-injection has become an applicable technology among other established nonviral delivery systems, such as particle bombardment or in vivo electroporation. The low-volume jet injector employed in this study uses compressed air to inject solutions of 1.5-10 microL containing naked DNA into the desired tissue. The novel design of this prototype makes multiple jet-injections possible. Therefore, repeated jet-injections into one target tissue can be performed easily. This jet-injector hand-held system was used for the direct in vivo gene transfer of plasmid DNA into tumors to achieve efficient expression of reporter genes (beta-galactosidase, green fluorescent protein [GFP]) and of therapeutic genes (TNF-alpha) in different tumor models. The study presented here revealed the key parameters of efficient in vivo jet-injection (jet-injection volume, pressure, jet penetration, DNA stability) to define the optimal conditions for a jet-injection-aided nonviral gene therapy.

Overexpression of the c-erbB-2 protein (also called HER-2/neu) is observed in a variety of malignancies including colorectal cancer (CRC). In this study we aimed to evaluate the rate of c-erbB-2 overexpression in our tumor collection and to clarify its correlation with the chromosomal status at the c-erbB-2 locus in CRC. Additionally we correlated the c-erbB-2 overexpression and the chromosomal gain of 17q with patient survival. Seventy-four specimens were analyzed immunohistochemically using a polyclonal c-erbB-2 antibody (DAKO) and the staining was scored according to the Clinical Trial Assay recommendations (0–3+). Of these, 45 cases were analyzed by comparative genomic hybridization (CGH) and immunohistochemistry (IHC). Overexpression was observed in 51% of the cases (score ≥2). Chromosomal gains at the c-erbB-2 locus were clearly correlated with overexpression of the gene (P=0.0009). Furthermore Kaplan–Meier analysis showed that overexpression of c-erbB-2 was significantly associated with poor survival and thus could serve as a prognostic marker. We conclude that c-erbB-2 is related with tumor progression in CRC which can be observed on protein level and reflects chromosomal gain at the locus at 17q.

Colorectal carcinoma (CRC) is associated with high incidence and mortality rate, particularly if metastasized to distant sites, such as the peritoneum. At time of first diagnosis 4 to 7 percent of the patients suffer from peritoneal metastasis (PM) of CRC. The PM is associated with poor prognosis and limited therapeutic options. Therefore, availability of adequate in vivo models for PM will promote the search for novel prognostic or even predictive biomarkers as well as the evaluation of chemosensitivity of PM towards standard, targeted and novel drugs. In this regard such models could be employed for more individualized concepts to improve the therapeutic outcome of PM patients. During the last decade patient-derived xenograft (PDX) mouse models have gained importance, since they closely resemble the molecular and biological features of the original primary tumors. However, until now no PM PDX models have been established from CRC. We therefore focused on the establishment and characterization of a novel CRC PM panel of PDX as useful platform for preclinical studies. For the PDX establishment colorectal surgical specimens were subcutaneously (s.c.) transplanted onto immunocompromized NOD scid gamma (NSG) mice. The successfully engrafted tumors were transferred to NMRI nu/nu mice for further passaging. Engrafted tumors were characterized by histopathology, immunohistochemistry and gene expression analyses using real-time RT-PCR. Chemosensitivity of PDX models was evaluated in vivo by application of a panel of conventional chemotherapeutic and of targeted drugs. For PDX establishment 68 CRC surgical specimens were transplanted onto NSG mice. From those, currently 22 PDX have engrafted and are stably growing on NMRI nu/nu mice. From those, 13 models of 10 patients have been analyzed. Their tumor doubling times ranged between 4 to 28 days. The histopathological evaluation revealed maintenance of the original CRC histology in the PDX. The chemosensitivity testing of conventional and of targeted drugs in the 13 PM PDX models revealed the individual, diverging response of the PDX, such as for 5-FU, irinotecan, oxaliplatin, cetuximab, and erlotinib. For selected PDX, orthotopic transplantation into the peritoneum revealed their potential to form disseminated tumors in kidney, ovaries and abdominal diaphragm. Our results demonstrate, that this novel panel of PDX maintains the morphology of the patient tumor in early passages, reflect heterogeneous response rates, and can be used as preclinical in vivo platform for translational studies of potential clinical use. In conclusion, this PDX panel of PM of the CRC can be used for further studies to evaluate new biomarker and to test novel therapies or combinations for PM of CRC. Citation Format: Wolfgang Walther, Eva Pachmayr, Bernadette Brzezicha, Britta Büttner, Beate Rau, Ulrike S. Stein. Patient-derived xenograft models from peritoneal metastasis of colorectal carcinoma as novel platform for biomarker analysis and drug testing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3853. doi:10.1158/1538-7445.AM2017-3853

We have previously identified the gene Metastasis-Associated in Colon Cancer 1 (MACC1). MACC1 acts a prognostic biomarker for tumor progression, metastasis and patient survival for a broad variety of solid cancer types. Here we assessed if the MACC1 gene could separate stage II colon cancer patients with proficient mismatch repair (pMMR) into high- and low-risk groups who might benefit from or be spared adjuvant chemotherapy based on their prognosis. In the Charité 1 discovery cohort (n=61), MACC1 expression and MSI status were assayed by qRT-PCR in cryo-preserved tumors from CRC patients. MSS/MSI-low/MACC1-low tumors showed better survival vs. MSS/MSI-low/MACC1-high (P Citation Format: Ulrich-Peter Rohr, Pia Herrmann, Katharina Ilm, Hai Zhang, Sabine Lohmann, Astrid Reiser, Andrea Muranyi, Janice Smith, Susen Burock, Marc Osterland, Katherine Leith, Shalini Singh, Patrick Brunhoeber, Rebecca Bowermaster, Jeanne Tie, Michael Christie, Hui-Li Wong, Paul Waring, Kandavel Shanmugam, Peter Gibbs, Ulrike S. Stein. Combination of DNA mismatch repair status and MACC1 expression in patients with stage II colon cancer: The BIOGRID studies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2778. doi:10.1158/1538-7445.AM2017-2778

Background The metastasis-associated in colon cancer 1 (MACC1) gene has been identified as prognostic biomarker for colorectal cancer (CRC). Here, we aimed at the refinement of risk assessment by separate and combined survival analyses of MACC1 expression with any of the markers KRAS mutated in codon 12 (KRAS G12) or codon 13 (KRAS G13), BRAF V600 mutation and MSI status in a retrospective study of 99 CRC patients with tumors UICC staged I, II and III. Findings We showed that only high MACC1 expression (HR: 6.09, 95% CI: 2.50-14.85, P

// Mathias Dahlmann 1 , Anna Okhrimenko 1 , Patrick Marcinkowski 1 , Marc Osterland 1 , Pia Herrmann 1 , Janice Smith 2 , Claus W. Heizmann 3 , Peter M. Schlag 1,4 , Ulrike Stein 1 1 Experimental and Clinical Research Center, Charite University Medicine Berlin and Max-Delbruck-Center for Molecular Medicine, Robert-Rossle-Strase 10, 13125 Berlin, Germany 2 Max-Delbruck-Center for Molecular Medicine, Robert-Rossle-Strase 10, 13125 Berlin, Germany 3 University Children’s Hospital, Division of Clinical Chemistry and Biochemistry, Steinwiesstrasse 75, 8032 Zurich, Switzerland 4 Charite Comprehensive Cancer Center, Charite University Medicine, Invalidenstrase 80, 10117 Berlin, Germany Correspondence: Ulrike Stein, email: // Keywords : Colorectal cancer, metastasis, S100A4, RAGE, signaling Received : February 03, 2014 Accepted : April 16, 2014 Published : April 17, 2014 Abstract Survival of colorectal cancer patients is strongly dependent on development of distant metastases. S100A4 is a prognostic biomarker and inducer for colorectal cancer metastasis. Besides exerting intracellular functions, S100A4 is secreted extracellularly. The receptor for advanced glycation end products (RAGE) is one of its interaction partners. The impact of the S100A4-RAGE interaction for cell motility and metastasis formation in colorectal cancer has not been elucidated so far. Here we demonstrate the RAGE-dependent increase in migratory and invasive capabilities of colorectal cancer cells via binding to extracellular S100A4. We show the direct interaction of S100A4 and RAGE, leading to hyperactivated MAPK/ERK and hypoxia signaling. The S100A4-RAGE axis increased cell migration (P

Das IMK analysiert regelmäßig mittels Daten von Eurostat die Entwicklung der Arbeitskosten und der Lohnstückkosten in Europa. In dieser aktuellen Auswertung wird zunächst die Entwicklung der Arbeitskosten in der Privatwirtschaft, dem privaten Dienstleistungssektor und dem Verarbeitenden Gewerbe in wichtigen europäischen Ländern, dem Euroraum und der Europäischen Union als Ganzes dargestellt. Zudem wird die Entwicklung der Arbeitskosten im öffentlichen Dienstleistungssektor aufgezeigt. Im Anschluss daran wird – unter Berücksichtigung der Arbeitsproduktivität – die Entwicklung der Lohnstückkosten untersucht. Im Jahr 2013 kostete in Deutschland eine Arbeitsstunde in der Privatwirtschaft 31,3 Euro. Wie im Vorjahr belegt Deutschland damit im europäischen Länderranking den achten Platz. Mit einer Veränderungsrate von 1,4% war die Arbeitskostenentwicklung 2013 schwach und lag nur knapp über dem europäischen Durchschnitt. Nach einem starken Anstieg im Jahr 2012 waren die deutschen Arbeitskosten im privaten Dienstleistungssektor in Höhe von 28,7 Euro 2013 wieder unterdurchschnittlich gestiegen und auch deutlich schwächer als im Verarbeitenden Gewerbe. Dabei liegen sie schon um rund ein Fünftel unterhalb der Arbeitskosten im Verarbeitenden Gewerbe. Die Krisenländer haben ihre Arbeits- und Lohnstückkosten in den letzten Jahren teilweise in dramatischer Weise bereits soweit nach unten korrigiert, dass sie mit der Zielinflationsrate der EZB von mittelfristig knapp 2 % kompatibel sind. Dagegen müssten die Löhne in Deutschland über mehrere Jahre hinweg um mehr als 3 % zulegen, um so den Anpassungsprozess der Krisenländer zu unterstützen und Deflationsgefahren im Euroraum zu begegnen.

Genotoxic stress leads to nuclear translocation of the Y-box transcription factor YB-1 and enhanced expression of the multidrug resistance gene MDR1. Because hyperthermia is used for the treatment of colon cancer in combination with chemoradiotherapy, we investigated the influence of hyperthermia on YB-1 activity and the expression of multidrug resistance-related genes. Here we report that hyperthermia causes YB-1 translocation from the cytoplasm into the nucleus of human colon carcinoma cells HCT15 and HCT116. Nuclear translocation of YB-1 was associated with increased MDR1 and MRP1 gene activity, which is reflected in strong efflux pump activity. However, a combination of hyperthermia and drug treatment effectively reduced cell survival of the HCT15 and HCT116 cells. These results demonstrate that activation of MDR1 and MRP1 gene expression and increased efflux pump activity after hyperthermia were insufficient to cause an increase in drug resistance in colon cancer cell lines. The ability of hyperthermia to abrogate drug resistance in the presence of an increase in functional MDR proteins may provide an explanation for the efficacious results seen in the clinic in colon cancer patients treated with a combination of hyperthermia and chemotherapy.

The jet-injection technology has developed as an applicable alternative to viral or liposomal gene delivery systems. In this study a novel, low-volume, 'high-speed jet injector' hand-held system was used for the direct gene transfer of naked DNA into tumors. Lewis-lung carcinoma bearing mice were jet-injected with the beta-galactosidase (LacZ), the green fluorescence (GFP) or the human tumor necrosis factor alpha (TNF-alpha) gene carrying vector plasmids. The animals received five jet injections into the tumor at a pressure of 3.0 bar, delivering 3--5 microl plasmid DNA (1 microg DNA/microl in water) per single jet injection. The jet injection of DNA leads to a widespread expression pattern within tumor tissues with penetration depths of 5--10 mm. Analysis of tumor cryosections revealed moderate LacZ or GFP expression at 48 h and strong reporter gene expression 72 h and 96 h after jet injection. The simultaneous jet injection of the TNF-alpha and LacZ carrying vectors demonstrated efficient expression and secretion of both the cytokine, as well as LacZ expression within the tumor 24 h, 48 h, 72 h, 96 h and 120 h after jet injection. These studies demonstrate the applicability of jet injection for the efficient in vivo gene transfer into tumors for nonviral gene therapy of cancer using minimal amounts of naked DNA.

Cancer gene therapy approaches are often designed as single-agent treatments; however, greater therapeutic effect might be obtained if combined with an established conventional treatment regimen such as chemotherapy. In this context, conditional promoters are useful tools, because they may be induced by therapeutic modalities. The human multidrug resistance gene (mdr1) promoter is inducible by cytostatic drugs and can be employed for the chemotherapy-regulated expression of therapeutic genes. In this in vivo study, the human mdr1 promoter fragment (-207 to +158) was used for drug-inducible expression of human tumor necrosis factor-alpha (TNF-alpha) in the vector construct pM3mdr-p-hTNF. The single doxorubicin and vincristine treatment of nude mice xenografted with pM3mdr-p-hTNF-transduced MCF-7 mammary tumors resulted in drug-induced and time-dependent elevation of intratumoral TNF-alpha expression at the mRNA and protein level. The highest drug induction was achieved at 2 days after drug application, as reflected by a maximum 25-fold increase in TNF-alpha secretion in the tumor. This drug-induced TNF-alpha expression is more effective in inhibiting tumor growth compared with the growth of tumors transduced with constitutively TNF-alpha-expressing vectors in combination with chemotherapy.