Modeling colorectal cancer: A bio-resource of 50 patient-derived organoid lines.

Background And Aim: Colorectal cancer (CRC) is the second leading cause of cancer death worldwide. To improve outcomes for these patients, we need to develop new treatment strategies. Personalized cancer medicine, where patients are treated based on the characteristics of their own tumor, has gained significant interest for its promise to improve outcomes and reduce unnecessary side effects.

Identifying subpopulations in multicellular systems by quantitative chemical imaging using label-free hyperspectral CARS microscopy.

Quantitative hyperspectral coherent Raman scattering microscopy merges imaging with spectroscopy and utilises quantitative data analysis algorithms to extract physically meaningful chemical components, spectrally and spatially-resolved, with sub-cellular resolution. This label-free non-invasive method has the potential to significantly advance our understanding of the complexity of living multicellular systems. Here, we have applied an in-house developed hyperspectral coherent anti-Stokes Raman scattering (CARS) microscope, combined with a quantitative data analysis pipeline, to imaging living mouse liver organoids as well as fixed mouse brain tissue sections xenografted with glioblastoma cells.

3D imaging of colorectal cancer organoids identifies responses to Tankyrase inhibitors.

Aberrant activation of the Wnt signalling pathway is required for tumour initiation and survival in the majority of colorectal cancers. The development of inhibitors of Wnt signalling has been the focus of multiple drug discovery programs targeting colorectal cancer and other malignancies associated with aberrant pathway activation. However, progression of new clinical entities targeting the Wnt pathway has been slow.

Developmentally regulated splice variants mediate transcriptional repressor functions during eye formation.

Tcf7l2 mediates Wnt/β-Catenin signalling during development and is implicated in cancer and type-2 diabetes. The mechanisms by which Tcf7l2 and Wnt/β-Catenin signalling elicit such a diversity of biological outcomes are poorly understood. Here, we study the function of zebrafish alternative splice variants and show that only variants that include exon five or an analogous human variant can effectively provide compensatory repressor function to restore eye formation in embryos lacking function.

Wnt ligand and receptor patterning in the liver.

In the liver, the tight spatiotemporal regulation of Wnt/β-catenin signaling is required to establish and maintain a metabolic form of tissue polarity termed zonation. In this review, we discuss the latest technologies applied in the study of liver zonation and provide a summary of the Wnt ligand and receptor expression patterns in the hepatic lobule. We further discuss the mechanisms, by which Wnt instructive cues might be spatially confined and propagated along the central vein-portal triad axis.

Cell Permeable Stapled Peptide Inhibitor of Wnt Signaling that Targets β-Catenin Protein-Protein Interactions.

The Wnt signaling pathway plays a critical role in cell proliferation and differentiation, thus it is often associated with diseases such as cancers. Unfortunately, although attractive, developing anti-cancer strategy targeting Wnt signaling has been challenging given that the most attractive targets are involved in protein-protein interactions (PPIs). Here, we develop a stapled peptide inhibitor that targets the interaction between β-catenin and T cell factor/lymphoid enhancer-binding factor transcription factors, which are crucially involved in Wnt signaling.

Assessing the mechanism and therapeutic potential of modulators of the human Mediator complex-associated protein kinases.

Mediator-associated kinases CDK8/19 are context-dependent drivers or suppressors of tumorigenesis. Their inhibition is predicted to have pleiotropic effects, but it is unclear whether this will impact on the clinical utility of CDK8/19 inhibitors. We discovered two series of potent chemical probes with high selectivity for CDK8/19.

Wnt and Neuregulin1/ErbB signalling extends 3D culture of hormone responsive mammary organoids.

The development of in vitro culture systems quantitatively and qualitatively recapitulating normal breast biology is key to the understanding of mammary gland biology. Current three-dimensional mammary culture systems have not demonstrated concurrent proliferation and functional differentiation ex vivo in any system for longer than 2 weeks. Here, we identify conditions including Neuregulin1 and R-spondin 1, allowing maintenance and expansion of mammary organoids for 2.

A functional connectome: regulation of Wnt/TCF-dependent transcription by pairs of pathway activators.

Background: Wnt/β-catenin signaling is often portrayed as a simple pathway that is initiated by Wnt ligand at the cell surface leading, via linear series of interactions between 'core pathway' members, to the induction of nuclear transcription from genes flanked by β-catenin/TCF transcription factor binding sites. Wnt/β-catenin signaling is also regulated by a much larger set of 'non-core regulators'. However the relationship between 'non-core regulators' is currently not well understood.

H-Prune through GSK-3β interaction sustains canonical WNT/β-catenin signaling enhancing cancer progression in NSCLC.

H-Prune hydrolyzes short-chain polyphosphates (PPase activity) together with an hitherto cAMP-phosphodiesterase (PDE), the latest influencing different human cancers by its overexpression. H-Prune promotes cell migration in cooperation with glycogen synthase kinase-3 (Gsk-3β). Gsk-3β is a negative regulator of canonical WNT/β-catenin signaling.

Huwe1-mediated ubiquitylation of dishevelled defines a negative feedback loop in the Wnt signaling pathway.

Wnt signaling plays a central role in development, adult tissue homeostasis, and cancer. Several steps in the canonical Wnt/β-catenin signaling cascade are regulated by ubiquitylation, a protein modification that influences the stability, subcellular localization, or interactions of target proteins. To identify regulators of the Wnt/β-catenin pathway, we performed an RNA interference screen in Caenorhabditis elegans and identified the HECT domain-containing ubiquitin ligase EEL-1 as an inhibitor of Wnt signaling.

Toward a quantitative understanding of the Wnt/β-catenin pathway through simulation and experiment.

Wnt signaling regulates cell survival, proliferation, and differentiation throughout development and is aberrantly regulated in cancer. The pathway is activated when Wnt ligands bind to specific receptors on the cell surface, resulting in the stabilization and nuclear accumulation of the transcriptional co-activator β-catenin. Mathematical and computational models have been used to study the spatial and temporal regulation of the Wnt/β-catenin pathway and to investigate the functional impact of mutations in key components.

Conditional disruption of Axin1 leads to development of liver tumors in mice.

Background & Aims: Mutations in components of the Wnt signaling pathway, including β-catenin and AXIN1, are found in more than 50% of human hepatocellular carcinomas (HCCs). Disruption of Axin1 causes embryonic lethality in mice. We generated mice with conditional disruption of Axin1 to study its function specifically in adult liver.

Evolutionary routes from a prebiotic ANA-world.

Recent experimental support has been generated for a model of prebiotic development that postulates a role for Amyloid-Nucleic Acid (ANA)-fibers as the earliest replicating entities capable of undergoing Darwinian evolution. Here, this new model is compared with existing RNA-world models with a particular focus on trajectories that lead to evolutionary-beneficial interactions between nucleic acid, protein and lipid components. This analysis suggests a number of new areas for fruitful experimental studies.

Amyloid-associated nucleic acid hybridisation.

Nucleic acids promote amyloid formation in diseases including Alzheimer's and Creutzfeldt-Jakob disease. However, it remains unclear whether the close interactions between amyloid and nucleic acid allow nucleic acid secondary structure to play a role in modulating amyloid structure and function. Here we have used a simplified system of short basic peptides with alternating hydrophobic and hydrophilic amino acid residues to study nucleic acid - amyloid interactions.

The potential for targeting oncogenic WNT/beta-catenin signaling in therapy.

There has been a surge of interest in the therapeutic targeting of the Wnt pathway following the demonstration that it is activated in a wide variety of tumors and that blocking aberrant signaling promoted tumor cell apoptosis or differentiation. This review describes recent therapeutic approaches and discusses potential opportunities for intervention at multiple levels within the Wnt pathway.

Deficiency of Mbd2 attenuates Wnt signaling.

We have previously shown that deficiency of the methyl binding domain protein Mbd2 dramatically reduces adenoma burden on an Apc(Min/+) background. To investigate the mechanism underlying this phenomenon, we have determined the effect of Mbd2 deficiency upon the phenotypes imposed by the conditional deletion of Apc in the small intestine. Microarray analysis demonstrated a partial suppression of the Wnt pathway in the absence of Mbd2.

Two functionally distinct Axin-like proteins regulate canonical Wnt signaling in C. elegans.

Axin is a central component of the canonical Wnt signaling pathway that interacts with the adenomatous polyposis coli protein APC and the kinase GSK3beta to downregulate the effector beta-catenin. In the nematode Caenorhabditis elegans, canonical Wnt signaling is negatively regulated by the highly divergent Axin ortholog PRY-1. Mutation of pry-1 leads to constitutive activation of BAR-1/beta-catenin-dependent Wnt signaling and results in a range of developmental defects.

Dishevelled (Dvl-2) activates canonical Wnt signalling in the absence of cytoplasmic puncta.

Dishevelled family proteins are multidomain intracellular transducers of Wnt signals. Ectopically expressed mammalian Dishevelled 2 (Dvl-2) activates downstream signalling and localises to cytoplasmic puncta. It has been suggested that these Dvl-2-containing structures correspond to intracellular vesicles and may be involved in the Wnt signal transduction process.

Contribution of the ABC transporters Bcrp1 and Mdr1a/1b to the side population phenotype in mammary gland and bone marrow of mice.

The ability of cells to export Hoechst 33342 can be used to identify a subpopulation of cells (side population [SP]) with characteristics of stem cells in many tissues. The ATP-binding cassette transporters Bcrp1 (Abcg2) and Mdr1a/1b (Abcb1a/1b) have been implicated as being responsible for this phenotype. To further explore the involvement of these transporters in the SP phenotype, we have generated Bcrp1/Mdr1a/1b triple knockout mice and studied the effect of their absence on the SP in bone marrow and mammary gland.

The breast cancer resistance protein BCRP (ABCG2) concentrates drugs and carcinogenic xenotoxins into milk.

Contamination of milk with drugs, pesticides and other xenotoxins can pose a major health risk to breast-fed infants and dairy consumers. Here we show that the multidrug transporter BCRP (encoded by ABCG2) is strongly induced in the mammary gland of mice, cows and humans during lactation and that it is responsible for the active secretion of clinically and toxicologically important substrates such as the dietary carcinogen PhIP, the anticancer drug topotecan and the antiulcerative cimetidine into mouse milk.

Protein kinase B/Akt acts via glycogen synthase kinase 3 to regulate recycling of alpha v beta 3 and alpha 5 beta 1 integrins.

Protein kinase B (PKB)/Akt is known to promote cell migration, and this may contribute to the enhanced invasiveness of malignant cells. To elucidate potential mechanisms by which PKB/Akt promotes the migration phenotype, we have investigated its role in the endosomal transport and recycling of integrins. Whereas the internalization of alpha v beta 3 and alpha 5 beta 1 integrins and their transport to the recycling compartment were independent of PKB/Akt, the return of these integrins (but not internalized transferrin) to the plasma membrane was regulated by phosphatidylinositol 3-kinases and PKB/Akt.

A divergent canonical WNT-signaling pathway regulates microtubule dynamics: dishevelled signals locally to stabilize microtubules.

Dishevelled (DVL) is associated with axonal microtubules and regulates microtubule stability through the inhibition of the serine/threonine kinase, glycogen synthase kinase 3beta (GSK-3beta). In the canonical WNT pathway, the negative regulator Axin forms a complex with beta-catenin and GSK-3beta, resulting in beta-catenin degradation. Inhibition of GSK-3beta by DVL increases beta-catenin stability and TCF transcriptional activation.

Functional and molecular characterisation of mammary side population cells.

Background: Breast cancer is thought to arise in mammary epithelial stem cells. However, the identity of these stem cells is unknown.

Methods: Studies in the haematopoetic and muscle systems show that stem cells have the ability to efflux the dye Hoechst 33342.

Structural basis for recruitment of glycogen synthase kinase 3beta to the axin-APC scaffold complex.

Glycogen synthase kinase 3beta (GSK3beta) is a serine/threonine kinase involved in insulin, growth factor and Wnt signalling. In Wnt signalling, GSK3beta is recruited to a multiprotein complex via interaction with axin, where it hyperphosphorylates beta-catenin, marking it for ubiquitylation and destruction. We have now determined the crystal structure of GSK3beta in complex with a minimal GSK3beta-binding segment of axin, at 2.