BiotecVisions 2013, February

Methylation of Cancer-Stem-Cell-Associated Wnt Target Genes Predicts Poor Prognosis in Colorectal Cancer Patients

Severe meningococcal disease in childhood

Immunotherapy Exposes Cancer Stem Cell Resistance and a New Synthetic Lethality

Targeting the Hedgehog and Notch Signaling Pathways

![][1] The Spanish National Cancer Institute (CNIO) Meeting on Development and Cancer took place between 4 and 6 February 2008 in Madrid, Spain, and was organized by K. Basler, G. Morata, E. Moreno and M. Torres. ![][2] Developmental biologists rightly argue that genes regulating developmental pathways are reiterated in cancer development. At the recent Spanish National Cancer Institute (CNIO) Meeting on Development and Cancer, novel concepts and processes that are considered to be common to both embryonic and cancer development were discussed. It is worth pointing out that tumour formation should be considered to be a form of aberrant organ development, in which many developmental pathways that control proliferation, differentiation and growth or morphogenesis are affected. This report is structured in three parts, describing the work presented at the meeting according to the stages of tumour development to which it pertains. First, the normal behaviour of cell populations that are the targets of transformation, in particular adult stem cells and the pathways that regulate their behaviour. Second, processes that occur at the early stages of cancer development before morphological malformations are detectable, in particular the process that is best known from studies in flies as ‘cell competition’. Third, developmental processes and pathways that are active in the growing tumour—which we consider to be an extrinsic organ—for example, the recruitment of blood vessels or the role of morphogens in controlling the size and shape of tumours, and the progression from hyperplasia to invasive tumours (Fig 1). Figure 1. Tumour development and genes involved in the different stages. Schematic representation of how stem cells (A) , by acquiring a mutation through cell competition ( B ) and by the selection of additional mutations, can give rise to the ‘tumour organ’ ( C ). The genes involved in these processes are listed on the right‐hand side, and most of them … [1]: /embed/graphic-1.gif [2]: /embed/graphic-2.gif

The human colorectal epithelium is maintained by multipotent stem cells that give rise to absorptive, mucous, and endocrine lineages. Recent evidence suggests that human colorectal cancers are likewise maintained by a minority population of so-called cancer stem cells. We have previously established a human colorectal cancer cell line with multipotent characteristics (HRA-19) and developed a serum-free medium that induces endocrine, mucous and absorptive lineage commitment by HRA-19 cells in vitro. In this study, we investigate the role of the β1 integrin family of cell surface extracellular matrix receptors in multilineage differentiation by these multipotent human colorectal cancer cells. We show that endocrine and mucous lineage commitment is blocked in the presence of function-blocking antibodies to β1 integrin. Function-blocking antibodies to α2 integrin also blocked both HRA-19 endocrine lineage commitment and enterocytic differentiation by Caco-2 human colon cancer cells; both effects being abrogated by the MEK inhibitor, PD98059, suggesting a role for ERK signaling in α2-mediated regulation of colorectal cancer cell differentiation. To further explore the role of α2 integrin in multilineage differentiation, we established multipotent cells expressing high levels of wild-type α2 integrin or a non-signaling chimeric α2 integrin. Overexpression of wild-type α2 integrin in HRA-19 cells significantly enhanced endocrine and mucous lineage commitment, while cells expressing the non-signaling chimeric α2 integrin had negligible ability for either endocrine or mucous lineage commitment. This study indicates that the collagen receptor α2β1 integrin is a regulator of cell fate in human multipotent colorectal cancer cells.

Kingdom-Agnostic Metagenomics and the Importance of Complete Characterization of Enteric Microbial Communities

Multidrug resistance and disease relapse is a challenging clinical problem in the treatment of breast cancer. In this study, we investigated the hyaluronan (HA)-induced interaction between CD44 (a primary HA receptor) and protein kinase Cepsilon (PKCepsilon), which regulates a number of human breast tumor cell functions. Our results indicate that HA binding to CD44 promotes PKCepsilon activation, which, in turn, increases the phosphorylation of the stem cell marker, Nanog, in the breast tumor cell line MCF-7. Phosphorylated Nanog is then translocated from the cytosol to the nucleus and becomes associated with RNase III DROSHA and the RNA helicase p68. This process leads to microRNA-21 (miR-21) production and a tumor suppressor protein (e.g. PDCD4 (program cell death 4)) reduction. All of these events contribute to up-regulation of inhibitors of apoptosis proteins (IAPs) and MDR1 (multidrug-resistant protein), resulting in anti-apoptosis and chemotherapy resistance. Transfection of MCF-7 cells with PKCepsilon or Nanog-specific small interfering RNAs effectively blocks HA-mediated PKCepsilon-Nanog signaling events, abrogates miR-21 production, and increases PDCD4 expression/eIF4A binding. Subsequently, this PKCepsilon-Nanog signaling inhibition causes IAP/MDR1 down-regulation, apoptosis, and chemosensitivity. To further evaluate the role of miR-21 in oncogenesis and chemoresistance, MCF-7 cells were also transfected with a specific anti-miR-21 inhibitor in order to silence miR-21 expression and inhibit its target functions. Our results indicate that anti-miR-21 inhibitor not only enhances PDCD4 expression/eIF4A binding but also blocks HA-CD44-mediated tumor cell behaviors. Thus, this newly discovered HA-CD44 signaling pathway should provide important drug targets for sensitizing tumor cell apoptosis and overcoming chemotherapy resistance in breast cancer cells.

Disorders of hemostasis and excessive menstrual bleeding: prevalence and clinical impact

Escherichia coli yaeT encodes an essential, conserved outer membrane (OM) protein that is an ortholog of Neisseria meningitidis Omp85. Conflicting data with N. meningitidis indicate that Omp85 functions either in assembly of OM proteins or in export of OM lipids. The role of YaeT in E. coli was investigated with a new temperature-sensitive mutant harboring nine amino acid substitutions. The mutant stops growing after 60 min at 44 degrees C. After 30 min at 44 degrees C, incorporation of [35S]methionine into newly synthesized OM proteins is selectively inhibited. Synthesis and export of OM phospholipids and lipopolysaccharide are not impaired. OM protein levels are low, even at 30 degrees C, and the buoyant density of the OM is correspondingly lower. By Western blotting, we show that levels of the major OM protein OmpA are lower in the mutant in whole cells, membranes, and the growth medium. SecA functions as a multicopy suppressor of the temperature-sensitive phenotype and partially restores OM proteins. Our data are consistent with a critical role for YaeT in OM protein assembly in E. coli.

Current research and future needs in allergic rhinitis and asthma

Asymptomatic versus symptomatic persistent atrial fibrillation: clinical and noninvasive characteristics (Journal of Internal Medicine 2002; 250: 390-7).

Based on an improved isolation procedure using perfusion chromatography, trimeric Photosystem 1 (PS1) complexes have been isolated from various deletion mutants of the mesophilic cyanobacterium Synechocystis PCC 6803. These mutants are only deficient in the deleted subunits, which was carefully checked by high resolution gel electrophoresis in combination with immunoblotting. These highly purified and well characterized PS1 particles were then examined by electron microscopy, followed by computer-aided image processing with single particle averaging techniques as described earlier (Kruip, J., Boekema, E. J., Bald, D., Boonstra, A. F., and Rögner, M. (1993) J. Biol. Chem. 268, 23353-23360). This precise methodological approach allowed a confident localization of the PS1 subunits PsaC, -D, -E, -F, and -J; it also shows shape and size of these subunits once integrated in the PS1 complex. Subunits PsaC, -D, and -E form a ridge on the stromal site, with PsaE toward the edge of each monomer within the trimer and PsaD extending toward the trimeric center, leaving PsaC in between. PsaF (near PsaE) and PsaJ are close together on the outer edge of each monomer; their proximity is also supported by chemical cross-linking, using the zero-length cross-linker EDC. This localization of PsaF contradicts the position suggested by the published low resolution x-ray analysis and shows for the first time the existence of at least one transmembrane alpha-helix for PsaF. A topographic three-dimensional map has been drawn from this set of results showing the location of the major PS1 subunits (besides PsaA and PsaB). These data also led to the assignment of electron density in the recent medium resolution x-ray structure for PS1 (Krauss, N., Schubert, W.-D., Klukas, O., Fromme, P., Witt, H. T., Saenger, W. (1996) Nat. Struct. Biol. 3, 965-973).

Quorum sensing mediated by specific signal compounds (autoinducers) allows bacteria to monitor their cell density and enables a synchronized regulation of target gene sets. The best studied group of autoinducers are the acylhomoserine lactones (AHSLs), which are central to the regulation of virulence in many plant and animal pathogens. Variation of the acyl side chain of the AHSLs underlies the observed species specificity of this communication system. Here we show that even different strains of the plant pathogen Erwinia carotovora employ different dialects of this language and demonstrate the molecular basis for the acyl chain length specificity of distinct AHSL synthases. Under physiological concentrations, only the cognate AHSL with the "right" acyl chain is recognized as a signal that will switch on virulence genes. Mutagenesis of the AHSL synthase gene expI(SCC1) identified the changes M127T and F69L as sufficient to effectively alter ExpI(SCC1) (an N-3-oxohexanoyl-l-homoserine lactone producer) substrate specificity to that of an N-3-oxooctanoyl-l-homoserine lactone producer. Our data identify critical residues that define the size of the substrate-binding pocket of the AHSL synthase and will help in understanding and manipulating this bacterial language.

Rescue therapy for drug resistant hepatitis B: Another argument for combination chemotherapy?