In this work, highly infiltrative brain tumors with a stem-like phenotype were established by xenotransplantation of human brain tumors in immunodeficient nude rats. These tumors coopted the host vasculature and presented as an aggressive disease without signs of angiogenesis. The malignant cells expressed neural stem cell markers, showed a migratory behavior similar to normal human neural stem cells, and gave rise to tumors in vivo after regrafting. Serial passages in animals gradually transformed the tumors into an angiogenesis-dependent phenotype. This process was characterized by a reduction in stem cells markers. Gene expression profiling combined with high throughput immunoblotting analyses of the angiogenic and nonangiogenic tumors identified distinct signaling networks in the two phenotypes. Furthermore, proinvasive genes were up-regulated and angiogenesis signaling genes were down-regulated in the stem-like tumors. In contrast, proinvasive genes were down-regulated in the angiogenesis-dependent tumors derived from the stem-like tumors. The described angiogenesis-independent tumor growth and the uncoupling of invasion and angiogenesis, represented by the stemlike cancer cells and the cells derived from them, respectively, point at two completely independent mechanisms that drive tumor progression. This article underlines the need for developing therapies that specifically target the stem-like cell pools in tumors.glioma ͉ invasiveness ͉ vessel cooption

The Notch signaling pathway regulates developmental cell-fate decisions and has recently also been linked to inflammatory diseases. Although therapies targeting Notch signaling in inflammation in theory are attractive, their design and implementation have proven difficult, at least partly due to the broad involvement of Notch signaling in regenerative and homeostatic processes. In this review, we summarize the supporting role of Notch signaling in various inflammation-driven diseases, and highlight efforts to intervene with this pathway by targeting Notch ligands and/or receptors with distinct therapeutic strategies, including antibody designs. We discuss this in light of lessons learned from Notch targeting in cancer treatment. Finally, we elaborate on the impact of individual Notch members in inflammation, which may lay the foundation for development of therapeutic strategies in chronic inflammatory diseases.

In contrast to other inflammatory cytokines, IL-33 selectively targets nonquiescent endothelial cells. By this novel concept, quiescent cells may remain nonresponsive to a proinflammatory stimulus that concomitantly triggers a powerful response in cells that have been released from contact inhibition.

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Colorectal cancer (CRC) represents the second most common cause of cancer mortality in the western world. The tumors frequently show metastatic spread which affects different organs such as lymph nodes, liver and lungs. Although the pattern of spread may vary, the initial step usually involves dissemination to regional lymph nodes. At present it is clear that neovessel formation, including lymphangiogenesis, represents key events in tumor progression. However, to what extent lymphangiogenesis contributes in the progression of CRC is unclear. This work focuses on recent progress within the field of tumor lymphangiogenesis with special reference to CRC, and on novel therapeutic strategies for anti-lymphangiogenic therapies. Inhibition of metastatic spread may be achieved by restriction of lymphatic vessel growth by using targeted therapeutic strategies towards molecules involved in lymphangiogenic signalling. Such adjuvant therapeutic approaches in addition to existing therapeutic strategies may represent a favourable treatment for CRCs with higher than average risk of disease recurrence and progression. ' 2007 Wiley-Liss, Inc.

BackgroundOrganotypic tumor spheroids, a 3D in vitro model derived from patient tumor material, preserve tissue heterogeneity and retain structural tissue elements, thus replicating the in vivo tumor more closely than commonly used 2D and 3D cell line models. Such structures harbour tumorigenic cells, as revealed by xenograft implantation studies in animal models and maintain the genetic makeup of the original tumor material. The aim of our work was a morphological and proteomic characterization of organotypic spheroids derived from colorectal cancer tissue in order to get insight into their composition and associated biology.ResultsMorphological analysis showed that spheroids were of about 250 μm in size and varied in structure, while the spheroid cells differed in shape and size and were tightly packed together by desmosomes and tight junctions. Our proteomic data revealed significant alterations in protein expression in organotypic tumor spheroids cultured as primary explants compared to primary colorectal cancer tissue. Components underlying cellular and tissue architecture were changed; nuclear DNA/ chromatin maintenance systems were up-regulated, whereas various mitochondrial components were down-regulated in spheroids. Most interestingly, the mesenchymal cells appear to be substantial component in such cellular assemblies. Thus the observed changes may partly occur in this cellular compartment. Finally, in the proteomics analysis stem cell-like characteristics were observed within the spheroid cellular assembly, reflected by accumulation of Alcam, Ctnnb1, Aldh1, Gpx2, and CD166. These findings were underlined by IHC analysis of Ctnnb1, CD24 and CD44, therefore warranting closer investigation of the tumorigenic compartment in this 3D culture model for tumor tissue.ConclusionsOur analysis of organotypic CRC tumor spheroids has identified biological processes associated with a mixture of cell types and states, including protein markers for mesenchymal and stem-like cells. This 3D tumor model in which tumor heterogeneity is preserved may represent an advantageous model system to investigate novel therapeutic approaches.

Tumour spheroids initiated from glioma biopsy specimens provide a valuable three-dimensional cell culture system that share several biological features of malignant brain tumours in situ. Upon xenotransplantation in immunodeficient rats, tumours derived from such spheroids exhibit a highly infiltrative growth. Successful cryopreservation of spheroid specimens therefore represents an excellent tool for future comparative studies of tumour growth and progression. Thus, if frozen stocks of human glioma spheroids can be established, similar to those obtained from cancer cell lines, it would ease the planning of biopsy-based experiments. In this context, it is crucial that cryopreservation does not alter the biological behaviour of the tumour spheroids. The biopsy spheroids were frozen to -40 degrees Celsius, stored for 1 week at -196 degrees Celsius, thawed rapidly and cultured for 1 week. The viability of the spheroids was compared against controls using a two-colour fluorescence assay, which demonstrated that cryopreservation was well tolerated. Using an in vitro invasion assay, it is shown that the freezing procedures did not affect the spheroids ability to invade a collagen gel. Cryopreserved and control tumour spheroids were equally tumourogenic, and produced overlapping survival curves when transplanted into the brains of immunocompromised rats. Immunohistochemical analyses showed no significant changes regarding microvessel density or proliferation index. Furthermore, gene expression profiling using a macroarray system detected no significant changes following cryopreservation. The present data show that cryopreservation is well tolerated, and represent a methodologically reliable storage method for biopsy spheroids that can be used in experimental studies at later time points.

Our findings reveal that NOTCH1 signaling supports the expression of a subset of inflammatory genes at the enhancer level and demonstrate how key signaling pathways converge on chromatin to coordinate the transition to an infla mmatory endothelial phenotype.