Characterization and comparison of hypoxia inducing factors on tumor growth and metastasis between two- and three-dimensional cancer models.

The monocarboxylic acid transporter 4 (Mct-4), a downstream biomarker of hypoxia inducing factor (HIF)-1α, is involved in the cellular response to hypoxia, as indicated by the hypoxic response element in its promoter region. Using a tumorsphere assay as an in vitro 3-dimensional (3D) model generated using 384-well ultra-low attachment (ULA) plates for cell proliferation analysis using a plate-based image cytometer, we identify a hypoxic response in the tumorsphere model that is distinct from that of cells grown under 2-dimensional (2D) normoxic conditions and demonstrate a key role for Mct-4 in enabling 3D growth. The tumorsphere model yields evidence of an essential role for Mct-4 in multiple cell lines, which were genetically modified to underexpress and overexpress Mct-4, evidence not apparent in a standard 2D model of growth in the same cell lines.

A graft-to strategy of poly(vinylphosphonates) on dopazide-coated gold nanoparticles using catalyst activation.

A modular synthetic pathway for poly(diethyl vinylphosphonates) grafting-to gold nanoparticles is presented. Utilising an azide-dopamine derivative as nanoparticle coating agent, alkyne-azide click conditions were used to covalently tether the polymer to gold nanoparticles leading to stable and well distributed colloids for different applications.

Pulmonary siRNA Delivery with Sophisticated Amphiphilic Poly(Spermine Acrylamides) for the Treatment of Lung Fibrosis.

RNA interference (RNAi) is an efficient strategy to post-transcriptionally silence gene expression. While all siRNA drugs on the market target the liver, the lung offers a variety of currently undruggable targets, which can potentially be treated with RNA therapeutics. To achieve this goal, the synthesis of poly(spermine acrylamides) (P(SpAA) is reported herein.

RDEA119/BAY 869766: a potent, selective, allosteric inhibitor of MEK1/2 for the treatment of cancer.

The RAS-RAF-mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK) kinase (MEK)-ERK pathway provides numerous opportunities for targeted oncology therapeutics. In particular, the MEK enzyme is attractive due to high selectivity for its target ERK and the central role that activated ERK plays in driving cell proliferation. The structural, pharmacologic, and pharmacokinetic properties of RDEA119/BAY 869766, an allosteric MEK inhibitor, are presented.

Human CD4+CD25+ regulatory T cells: proteome analysis identifies galectin-10 as a novel marker essential for their anergy and suppressive function.

CD4(+)CD25(+)Foxp3(+) regulatory T cells (CD25(+) Treg cells) direct the maintenance of immunological self-tolerance by active suppression of autoaggressive T-cell populations. However, the molecules mediating the anergic state and regulatory function of CD25(+) Treg cells are still elusive. Using differential proteomics, we identified galectin-10, a member of the lectin family, as constitutively expressed in human CD25(+) Treg cells, while they are nearly absent in resting and activated CD4(+) T cells.

An enzyme module system for the synthesis of dTDP-activated deoxysugars from dTMP and sucrose.

A flexible enzyme module system is presented that allows preparative access to important dTDP-activated deoxyhexoses from dTMP and sucrose. The strategic combination of the recombinant enzymes dTMP-kinase and sucrose synthase (SuSy), and the enzymes RmlB (4,6-dehydratase), RmlC (3,5-epimerase) and RmlD (4-ketoreductase) from the biosynthetic pathway of dTDP-beta-L-rhamnose was optimized. The SuSy module (dTMP-kinase, SuSy, +/-RmlB) yielded the precursor dTDP-alpha-D-glucose (2) or the biosynthetic intermediate dTDP-6-deoxy-4-keto-alpha-D-glucose (3) on a 0.

Application of proteomics and protein analysis for biomarker and target finding for immunotherapy.

Regulatory T-cells play a central role in the maintenance of the immunological balance and are powerful inhibitors of T-cell activation both in vivo and in vitro. The enhancement of suppressor-cell function might be a target for immunotherapeutic approaches for the treatment of immune-mediated diseases like multiple sclerosis and Crohn's disease.The method of choice to elucidate the still unclear effector functions of regulatory T-cells is the differential proteome analyses performed with human and murine T-cell populations.

Association of dopamine D(3) receptors with actin-binding protein 280 (ABP-280).

Proteins that bind to G protein-coupled receptors have been identified as regulators of receptor localization and signaling. In our previous studies, a cytoskeletal protein, actin-binding protein 280 (ABP-280), was found to associate with the third cytoplasmic loop of dopamine D(2) receptors. In this study, we demonstrate that ABP-280 also interacts with dopamine D(3) receptors, but not with D(4) receptors.

Interaction of gamma-COP with a transport motif in the D1 receptor C-terminus.

Truncations at the carboxyl termini of G protein-coupled receptors result in defective receptor biogenesis and comprise a number of inherited disorders. In order to evaluate the structural role of the C-terminus in G protein-coupled receptor biogenesis, we generated a series of deletion and substitution mutations in the dopamine D1 receptor and visualized receptor subcellular localization by fusion to a green fluorescent protein. Alanine substitutions of several hydrophobic residues within the proximal C-terminus resulted in receptor transport arrest in the ER.

Protection of intracellular dopamine cytotoxicity by dopamine disposition and metabolism factors.

Dopamine has been hypothesized as a contributing factor for the selective degeneration of dopaminergic neurons in Parkinson's disease. However, the cytotoxic mechanisms of dopamine and its metabolites remain poorly understood. Using a stable aromatic amino acid decarboxylase (AADC) expressing a fibroblast cell line, we previously demonstrated a novel, non-oxidative cytotoxicity of intracellular dopamine.

Evidence for non-oxidative dopamine cytotoxicity: potent activation of NF-kappa B and lack of protection by anti-oxidants.

A stable aromatic acid decarboxylase expressing the Chinese hamster ovary cell line was developed to study the cytotoxic properties of intracellular and extracellular dopamine. The relative impermeability of cells to dopamine, but not to L-DOPA, allows the differentiation of extracellular and intracellular dopamine cytotoxicity. In contrast to extracellular dopamine, intracellular dopamine toxicity was resistant to antioxidant protection, and did not require melanin formation for its toxicity.

Analysis of genes involved in 6-deoxyhexose biosynthesis and transfer in Saccharopolyspora erythraea.

Glycosylation represents an attractive target for protein engineering of novel antibiotics, because specific attachment of one or more deoxysugars is required for the bioactivity of many antibiotic and antitumour polyketides. However, proper assessment of the potential of these enzymes for such combinatorial biosynthesis requires both more precise information on the enzymology of the pathways and also improved Escherichia coli-actinomycete shuttle vectors. New replicative vectors have been constructed and used to express independently the dnmU gene of Streptomyces peucetius and the eryBVII gene of Saccharopolyspora erythraea in an eryBVII deletion mutant of Sac.