Structure-Activity Relationship and Biological Investigation of a REV-ERBα-Selective Agonist SR-29065 () for Autoimmune Disorders.

Autoimmune diseases affect 50 million Americans, predominantly women, and are thought to be one of the top 10 leading causes of death among women in age groups up to 65 years. A central role for T17 cells has been highlighted by genome-wide association studies (GWAS) linking genes preferentially expressed in T17 cells to several human autoimmune diseases. We and others have reported that the nuclear receptors REV-ERBα and β are cell-intrinsic repressors of T17 cell development and pathogenicity and might therefore be therapeutic targets for intervention.

High-throughput optofluidic screening of single B cells identifies novel cross-reactive antibodies as inhibitors of uPAR with antibody-dependent effector functions.

The urokinase-type plasminogen activator receptor (uPAR) is an essential regulator for cell signaling in tumor cell proliferation, adhesion, and metastasis. The ubiquitous nature of uPAR in many aggressive cancer types makes uPAR an attractive target for immunotherapy. Here, we present a rapid and successful workflow for developing cross-reactive anti-uPAR recombinant antibodies (rAbs) using high-throughput optofluidic screening of single B-cells from human uPAR-immunized mice.

Pyridyl aminothiazoles as potent inhibitors of Chk1 with slow dissociation rates.

Pyridyl aminothiazoles comprise a novel class of ATP-competitive Chk1 inhibitors with excellent inhibitory potential. Modification of the core with ethylenediamine amides provides compounds with low picomolar potency and very high residence times. Investigation of binding parameters of such compounds using X-ray crystallography and molecular dynamics simulations revealed multiple hydrogen bonds to the enzyme backbone as well as stabilization of the conserved water molecules network in the hydrophobic binding region.

Pyridyl aminothiazoles as potent Chk1 inhibitors: optimization of cellular activity.

Translation of significant biochemical activity of pyridyl aminothiazole class of Chk1 inhibitors into functional CEA potency required analysis and adjustment of both physical properties and kinase selectivity profile of the series. The steps toward optimization of cellular potency included elimination of CDK7 activity, reduction of molecular weight and polar surface area and increase in lipophilicity of the molecules in the series.

Biochemical and structural characterization of a novel class of inhibitors of the type 1 insulin-like growth factor and insulin receptor kinases.

The type 1 insulin-like growth factor receptor (IGF-1R) is often overexpressed on tumor cells and is believed to play an important role in anchorage-independent proliferation. Additionally, cell culture studies have indicated that IGF-1R confers increased resistance to apoptosis caused by radiation or chemotherapeutic agents. Thus, inhibitors of the intracellular kinase domain of this receptor may have utility for the clinical treatment of cancer.

Functional significance of type 1 insulin-like growth factor-mediated nuclear translocation of the insulin receptor substrate-1 and beta-catenin.

Previous work has shown that the transcriptional regulator beta-catenin can translocate to the nuclei when cells are stimulated with the type 1 insulin-like growth factor (IGF-1). We show by immunocoprecipitation and by confocal microscopy that beta-catenin binds to and co-localizes with the insulin receptor substrate-1 (IRS-1), a docking protein for both the insulin and the IGF-1 receptors. IRS-1 is required for IGF-1-mediated nuclear translocation of beta-catenin, resulting in the activation of the beta-catenin target genes.

24p3 in differentiation of myeloid cells.

24p3 is a secreted lipocalin that has been variously related to apoptosis, proliferation, and the neutrophil lineage of blood cells. We have investigated the expression of 24p3 mRNA and protein in myeloid cell lines induced to differentiate by insulin-like growth factor 1 (IGF-1) and the granulocytic-colony simulating factor (G-CSF). Both these growth factors, which cause myeloid cells to differentiate into granulocytes, induced a marked increase in the expression of both 24p3 protein and mRNA.

A modified tandem affinity purification tag technique for the purification of protein complexes in mammalian cells.

The tandem affinity purification (TAP) tag technique has been used with success to identify under nondenaturing conditions protein complexes in yeast. The technique can be used in mammalian cells, but we found that the original technique does not yield enough recovery for the identification of proteins when mammalian cells growing in monolayer have to be used. We present here a modified TAP tag technique that allows sufficient recovery of proteins from mouse fibroblasts growing in monolayer cultures.

Control of cell size through phosphorylation of upstream binding factor 1 by nuclear phosphatidylinositol 3-kinase.

The insulin-like growth factor I/insulin receptor substrate 1 axis controls, in a nonredundant way, approximately 50% of cell and body size in animals from Drosophila to mice and in cells in culture. Although other factors may also intervene, cell size is strongly dependent on ribosome biogenesis, which is under the control of RNA polymerase I activity. We have previously shown that insulin receptor substrate 1 (IRS-1) translocates to the nuclei and nucleoli, where it binds to the upstream binding factor (UBF) 1, a regulator of RNA polymerase I activity.

Nuclear translocation of insulin receptor substrate-1 by the simian virus 40 T antigen and the activated type 1 insulin-like growth factor receptor.

32D cells are a murine hemopoietic cell line that undergoes apoptosis upon withdrawal of interleukin-3 (IL-3) from the medium. 32D cells have low levels of the type 1 insulin-like growth factor (IGF-I) receptor and do not express insulin receptor substrate-1 (IRS-1) or IRS-2. Ectopic expression of IRS-1 delays apoptosis but cannot rescue 32D cells from IL-3 dependence.

Regulation of Id1 protein expression in mouse embryo fibroblasts by the type 1 insulin-like growth factor receptor.

The activated type 1 insulin-like growth factor (IGF-IR) increases the expression of Id1 proteins in mouse embryo fibroblasts (MEF). Up-regulation depends on a functional receptor and on multiple pathways originating from different domains of the receptor. In MEF, Id1 protein expression is also up-regulated by serum and certain oncogenes.