A high-throughput screen for pre-mRNA splicing modulators

In this effort, we have aimed to 1) identify specific modulators of LMNA pre-mRNA splicing; these compounds are leads in the search for drugs with therapeutic potential in HGPS and 2) identify general pre-mRNA splicing inhibitors; these compounds will be useful as experimental tools in the study of pre-mRNA splicing mechanisms.

High throughput screening for SMA

A screen of this assay with a compound library can identify compounds that increase SMN2 levels by three mechanisms: modulating alternative splicing of SMN exon 7, increasing transcription from the SMN2 promoter, or stabilizing the SMN protein.

High Throughput Screening for Small Molecule Inhibitors of Heparin-induced Tau Fibril Formation

To identify novel inhibitors of tau fibrillization.

HTS for Identification of Inhibitors against the ERK Signaling Pathway using a Homogenous Cell-based Assay

The goal of this project was to evaluate this technology and screen the MLPCN’s compound collection (the MLSMR) to identify novel cell-membrane permeable inhibitors of the ERK signaling pathway.

Antagonists of IAP-family anti-apoptotic proteins

In this study we investigated small molecule compounds that mimicked the effect of SMAC in antagonizing IAPs by causing them to release Caspases.

uHTS for the identification of compounds that potentiate TRAIL-induced apoptosis of cancer cells

Based on the multi-component regulation of TRAIL-mediated apoptotic signaling we believe that by employing high throughput screening (HTS) techniques we will identify BCCG CPR TRAIL CID3380841 Page 3 of 17 Template H chemical hits and develop them into research tools and, ultimately, drugs that will efficiently, specifically, and safely sensitize different tumor cell lines to TRAIL-induced apoptosis.

Therapeutic Inhibitors of Phosphomannose Isomerase

We proposed that CDG-Ia patients will benefit from dietary mannose if we simultaneously reduce PMI activity with a non-competitive or un-competitive inhibitor. This would allow a modest intracellular accumulation of Man-6-P and drive metabolic flux into the glycosylation pathway using the residual PMM2 activity.

Placental Alkaline Phosphatase (PLAP) Luminescent HTS assay

the identification of PLAP-specific inhibitors with selectivity over tissue non-specific alkaline phosphatase (TNAP) and intestinal alkaline phosphatase (IAP) will provide the necessary tools to characterize its biological role. Currently, inhibitors of PLAP lack either potency or selectivity.

qHTS Assay for Inhibitors of 15-hLO-1 (15-human lipoxygenase 1)

The overall goal is to screen against three lipoxygenase isozymes; reticulocyte 15hLO-1, epidermal 15hLO-2 and platelet 12hLO, with the aim of finding selective inhibitors specific for each isozyme.

Profiling Report: Detergent-sensitive Inhibitors of Cruzain (Aggregators)

Using the cruzain enzymatic assay as a model screening system of an enzyme carrying a reactive cysteine catalytic residue, we attempt to profile the MLSMR collection with respect to several sources of false-positive or promiscuous types of inhibition: compound autofluorescence (1), colloidal aggregation (2), and reactive compounds.

Promiscuous and Specific Inhibitors of Cruzain (6-(3,5-difluorophenylamino)-9-ethyl-9H-purine-2-carbonitrile)

We aim to discover and develop novel cruzain inhibitors to ultimately aid in the development of antitrypanosomal agents.

Promiscuous and Specific Inhibitors of Cruzain (2-oxo-1,2-diphenylethyl 2-(cyclohexanecarboxamido)acetate)

We aim to discover and develop novel cruzain inhibitors to ultimately aid in the development of antitrypanosomal agents.

Probe Report for Nox1 Inhibitors

The identification of potent, selective inhibitors of Nox1 may lead to potential candidates for excess cell proliferation, cancer, and IBD. The known Nox inhibitors are of low micromolar potencies and are non-selective.

Therapeutic Inhibitors of Phosphomannose Isomerase

To develop novel phosphomannose isomerase (PMI) inhibitors for further characterization and therapeutics of Congenital Disorders of Glycosylation (CDG).

HTS identification of compounds activating TNAP at an intermediate concentration of phosphate acceptor detected in luminescent assay

The specific aims of this project were to identify small molecule compounds in the MLSMR that were highly specific activators of TNAP using a luminescence-based assay; test the confirmed positives in a secondary assay with natural substrates of TNAP, check for specificity against other recombinant phosphatases and test confirmed positives for their ability to increase calcification in osteoblast cultures. The novel chemical probes identified in this way may ultimately lead to a novel therapy for osteoporosis patients.

The Role of PHOSPHO1 in the Initiation of Skeletal Calcification

The main aim of this project was to screen large comprehensive chemical libraries to identify lead compounds for PHOSPHO1-specific inhibitors that will enable the elucidation of the functional involvement of this enzyme in skeletal mineralization and related biological phenomena.

Placental Alkaline Phosphatase (PLAP) Luminescent HTS assay

The identification of PLAP-specific inhibitors with selectivity over tissue non-specific alkaline phosphatase (TNAP) and intestinal alkaline phosphatase (IAP) will provide the necessary tools to characterize its biological role.

Identification of lipid storage modulators

The goal is to identify chemical probes affecting lipid storage regulation.

Identification of activators for the M2 isoform of human pyruvate kinase

We aimed to discover and optimize selective modulators of human pyruvate kinase M2 (hPK-M2), a splice isoform of pyruvate kinase that is specifically expressed in tumor cells.

Probe Report for RBBP9 Inhibitors

The identification of compounds that selectively inhibit RBBP9 activity may provide valuable probes for the study of apoptosis, cell cycle, and tumorigenesis.

Probe Report for P97/cdc48 Inhibitors

The identification of probes that selectively target p97 activity may provide insights into the biological roles of P97.

Identification of Functionally Selective Small Molecule Antagonists of the Neuropeptide-S Receptor: Naphthopyranopyrimidines

The goal of this project is to identify antagonists for neuropeptide S receptor as research probes to further study the functions of NPS receptor in animal models.

HTS for Inhibitors of BAP1

The long-term goal of this project is to identify a specific, selective inhibitor compound for the deubiquitinating activity of BAP1, a BRCA1 associated deubiquitinating enzyme.

Discovery of a Highly Selective KCC2 Antagonist

To identify small molecule ligands (both agonists and antagonists) of the neuronal K-Cl cotransporter KCC2.

Inhibitors of Protein Folding: DnaK

The specific aim of this project was to identify small molecule binders that would modulate/alter the function of DnaK by specifically interacting with its substrate-binding domain (SBD).

Probe Report for NPY-Y2 Receptor Antagonists

to produce brain penetrant, high affinity selective ligands for the Y2 receptor.

Discovery of a Highly Selective in vitro and in vivo M1 Allosteric Agonist Probe

To identify small molecule agonists of M1 muscarinic receptor that are cell permeable, exhibit submicromolar potency, and show greater than 10 fold selectivity over other muscarinic family members M2, M3, M4 and M5.

Tumor Hsp90 Inhibitor

To identify novel tumor Hsp90 inhibitors with potential for use in cancer therapy and in the treatment of neurodegenerative diseases.

Inhibitors of BRCT-Phosphoprotein Interaction

To identify inhibitors of the protein-protein interaction between tumor suppressor BRCT and phosphoproteins.

Splicing modulators at the Beta Globin locus

Identify small-molecule modulators of Beta-Globin splicing by high throughput screening of chemical libraries.

Agonists of the Thyroid Stimulating Hormone Receptor

Identify small-molecule agonists of the Thyroid Stimulating Hormone Receptor.

Epigenetic Modulators

Identify small-molecule modulators of cellular epigenetic pathways

Epigenetic Modulators

Identify small-molecule modulators of cellular epigenetic pathways

Epigenetic Modulators

Identify small-molecule modulators of cellular epigenetic pathways

Ikappa-B-alpha stabilizers in a human lymphoma cell line

DNA-polymerase III (polymerase core enzme) inhibitor

1. Transfer and run the HTS assay for the DNA polymerase III Holoenzyme (Pol III HE) from the representative Gram (-) model organism E. coli. 2. Prioritize compounds identified via HTS of the MLSCN library for further chemical optimization and development.

DNA Primase Inihibitor

1. Transfer and run the HTS assay for the DNA polymerase III Holoenzyme (Pol III HE) from the representative Gram (-) model organism E. coli. 2. Prioritize compounds identified via HTS of the MLSCN library for further chemical optimization and development.

qHTS Assay for Disrupters of an Hsp90 Co-Chaperone Interaction

Heat shock protein 90 (Hsp90) is the essential molecular chaperone which regulates a variety of cellular functions including development, cell cycle, and steroid hormone signaling. The disruption of interaction between the C-terminal portion of Hsp90 and TPR2A domain of HOP may regulate the Hsp90 functions.

Tau Filament Binding

Identify ligands capable of binding amyloidgenic tau conformations with high affinity.

Inhibitor of the Mevalonate Pathway in Streptococcus pneumoniae 1: Inhibitors of Mevalonate Kinase (MK)

To identify specific probes that inhibit the MK enzyme, the first of three enzymes in the mevalonate pathway of S. pneumoniae.

Pantothenate Synthetase (PanC) Inhibitor

To identify specific probes that inhibit the Mycobacterium tuberculosis pantothenate synthetase enzyme for X-ray crystallographic studies

S1P1 Antagonist

Identify compounds which provide insight into the molecular mechanism of S1P biological function.

High Throughput Fluorescence Polarization Screen for Bcl-B Phenotypic Converters

To identify small molecule chemical probes that bind to Bcl-B within its TR3-binding site.

Autoinducing pheromone (AIP)-dependent bacterial quorum sensing: AIP binding target

1. To screen libraries of small molecules in a high throughput fluorescence-based screening assay to identify compounds capable of suppressing pheromone-dependent activation of the promoter for a global regulator of Staphylococcus aureus virulence, RNAIII. 2. To confirm that the compounds that inhibit RNAIII promoter activation also inhibit expression of the virulence genes that are regulated by RNAIII.

Autoinducing pheromone (AIP)-dependent bacterial quorum sensing of S. aureus Agr3 agr locus genotype: target downstream of AIP binding

1. To screen libraries of small molecules in a high throughput fluorescence-based screening assay to identify compounds capable of suppressing pheromone-dependent activation of the promoter for a global regulator of Staphylococcus aureus virulence, RNAIII. 2. To confirm that the compounds that inhibit RNAIII promoter activation also inhibit expression of the virulence genes that are regulated by RNAIII.

Activator of Prostate Cell Differentiation

1. To screen for small molecules that modulate prostate cell differentiation, using HyperCyt® high throughput (HT) flow cytometry. 2. To counter-screen active compounds identified in the primary and confirmatory screens in LNCaP cells in a similar dose response assay using PC-3 cells to identify small molecules that induce intracellular granularity in an androgen-independent manner.

Competitive inhibitor of lingand binding for G protein-coupled receptor 30 (GPR30)

To identify small molecule ligands specific for GPR30 or the classical estrogen receptors (ER? and ER?).

Competitive inhibitor of lingand binding for G protein-coupled receptor 30 (GPR30)

To identify small molecule ligands specific for GPR30 or the classical estrogen receptors (ER? and ER?).

Formylpeptide Receptor (FPR) Antagonist

Identify small-molecule probes directed against the human formylpeptide receptor (FPR), a G-protein coupled receptor implicated in anti-bacterial inflammatory responses and malignant glioma metastasis.

Formylpeptide Receptor-Like 1 (FPRL-1) Antagonist

Identify small-molecule probes directed against the human formylpeptide receptor-like-1 (FPRL1), a G-protein coupled receptor implicated in anti-bacterial inflammatory responses and malignant glioma metastasis.

Ruthenium-based TrkA Inhibitor

To design potent, specific, and novel Ruthenium-based small-molecule inhibitors of the protein kinase TrkA.

Aggregation and Clearance of Huntingtin Inclusions: Huntington’s Disease Assay

To discover modifiers of pathways responsible for clearance of mutant Huntingtin protein.

Protein Kinase D Inhibitor

To identify small molecule inhibitors of PKD

Inhibitors of the Polo box domain of Human Polo-like kinase 1

1. Screen MLSCN compound libraries for inhibitors of the polo box domain of polo-like kinase-1 (Plk-1-PBD) using a high throughput-ready fluorescence polarization (FP) assay. 2. Follow up primary screen of PBD inhibitors with secondary biochemical and cell based assays to confirm hits.

Inhibitor of anti-apoptotic protein Bfl-1

To identify chemical probes of Bfl-1 through a fluorescence polarization assay (FPA) using FITC-Bid BH3 peptide.

Inhibitor of anti-apoptotic protein Bfl-1

To identify chemical probes of Bfl-1 through a fluorescence polarization assay (FPA) using FITC-Bid BH3 peptide.

EphA4 Receptor Antagonists for Nervous System Repair

To identify small molecule antagonists that inhibit the binding of EphA4 to the KYL peptide which interacts with high affinity to the ephrin-binding site of EphA4.

Tissue-nonspecific Alkaline Phosphatase (TNAP)

To identify novel highly potent and selective inhibitors of TNAP which to date have not been available.

Intestinal alkaline phosphatase (IAP-L) Inhibitor

To identify inhibitors of intestinal alkaline phosphatase (IAP) that show selectivity for the target over the other alkaline phosphatase isozymes (PLAP, TNAP).

Chemical Inhibitors of ER Stress

To identify compounds that selectively inhibit cell death induced by endoplasmic reticulum (ER) stress in mammalian cells.

Human Cathepsin L Inhibitor

To perform a high throughput screen of the NIH Molecular Libraries Small Molecule Repository (MLSMR) to identify novel, potent, and selective inhibitors of human cathepsin L.

N-(quinolin-8-yl)benzenesulfonamides as Agents Capable of Down-Regulating NF?B Activity within Two Separate High-Throughput Screens of NF?B Activation

To identify specific compounds that inhibit signaling mediated by NF?B after stimulation of human umbilical vein endothelial cells (HUVEC) by TNFalpha

Matrix Metalloproteinase-8 (MMP-8) Inhibitor

To identify selective chemical inhibitors of MMP-8

Matrix Metalloproteinase-13 (MMP-13) Inhibitor

To identify selective chemical inhibitors of MMP-13

Matrix Metalloproteinase-13 (MMP-13) Inhibitor

To identify selective chemical inhibitors of MMP-13

Sphingosine-1-phosphate receptor 2 (S1P2) Agonist

To identify specific agonists of the S1P2 receptor

Phosphodiesterase 4 (PDE4) Inhibitor in the cyclic response element-binding proteins (CREB) pathway

Identify small molecules modulators of PDEs (specifically PDE4) which are likely to be strong regulators of CREB signaling.

Chemical inhibitors of antigen receptor-induced NF-?B

To identify compounds that selectively inhibit one of the several known pathways that lead to NF-kB activation in mammalian cells.

Tissue-nonspecific Alkaline Phosphatase (TNAP)

To identify novel highly potent and selective inhibitors of TNAP which to date have not been available.

High Throughput Screening Campaign to Identify Inhibitors of MMP-8

To identify selective exosite/active site inhibitors of A Disintegrin and metalloproteinase with thrombospondin motifs-4 (ADAMTS-4)

AmpC beta-lactamase Non-Covalent Inhibitor

To identify novel inhibitors (both covalent and non-covalent) of the AmpC B-lactamase enzyme class.

AmpC beta-lactamase Covalent Inhibitor

To identify novel inhibitors (both covalent and non-covalent) of the AmpC B-lactamase enzyme class.

Inhibitors of the NS2B-NS3 Proteinase of West Nile Virus

Identify drug-like small molecule inhibitors of the NS3 West Nile virus protease by high throughput screening of chemical libraries.

Human Cathepsin L Inhibitor

To perform a high throughput screen of the NIH Molecular Libraries Small Molecule Repository (MLSMR) to identify novel, potent, and selective inhibitors of human cathepsin L.

12-kDa FK506-binding protein (FKBP12) Activator

Identify novel chemical probes that target FKBP12

Toll-Like Receptor 4-MyD88 Signaling Inhibitor

To identify inhibitors of TLR4 signaling

Measles Virus Inhibitor

1. To identify novel MV-specific probes through high throughput screening (HTS) of the MLSCN library. 2. To counterscreen putative probes in independent assays and rank confirmed hits by active concentration and cytotoxicit. 3. To determine the specificity of selected probes and assess their mechanisms of antiviral activity

High-Throughput Screening Campaign to Identify Inhibitors of SF-1

To identify selective cell-permeable inhibitors of the steroidogenic factor 1

High-Throughput Screening Campaign to Identify Inhibitors of SF-1

To identify modular, chemically tractable, selective, and cell-permeable inverse agonists of the nuclear receptor RAR.

High-Throughput Screening Campaign to Identify Inhibitors of SF-1

To identify selective cell-permeable inhibitors of the steroidogenic factor 1 (orphan nuclear receptor NR5A1)

HIV-1 Reverse Transcriptase Associated Ribonuclease H (Rnase H) Inhibitor

To identify small-molecule inhibitors of HIV reverse transcriptase associated ribonuclease H (HIV RT RNH).

Zebrafish Angiogenesis Inhibitor

Specific Aim of Probe Project

Rho Kinase II Inhibitor

To find potent inhibitors for Rho-Kinase. “Kinase-Glo”, an ATP depletion assay was used to find inhibitors that are specific to the ATP binding site.

CREB signaling pathway: CRE potentiator

Identify small molecular potentiators of the CREB pathway for memory and cognitive disorders using quantitative high throughput screening (qHTS).

Allosteric Modulators of the M1 Muscarinic Receptor - Antagonist

To identify small molecule modulators of M1 muscarinic receptor that are cell permeable, exhibit micromolar potency, and show greater than 10 fold selectivity over other muscarinic family members M2, M3, M4 and M5.

Molecule Modulators: Voltage-Dependent Potassium (RMI)

To identify small molecule modulators of KvBeta that are cell permeable and have a Km sufficient for in vitro cellular physiology assays.

Glucocerebrosidase Inhibitor (Chemotype 3)

Identification of inhibitors of glucocerebrosidase that may function as molecular chaperones to restore glucocerebrosidase activity.

Glucocerebrosidase Inhibitor (Chemotype 2)

Identification of inhibitors of glucocerebrosidase that may function as molecular chaperones to restore glucocerebrosidase activity.

Glucocerebrosidase Inhibitor (Chemotype 1)

Identification of inhibitors of glucocerebrosidase that may function as molecular chaperones to restore glucocerebrosidase activity.

Sphingosine-1-phosphate receptor 1 (S1P1) Agonist

Identify compounds which provide insight into the molecular mechanism of S1P biological function.

Sphingosine-1-phosphate receptor 3 (S1P3) Agonist

Identify compounds which provide insight into the molecular mechanism of S1P biological function.

Sphingosine-1-phosphate receptor 3 (S1P3) Agonist

Identify compounds which provide insight into the molecular mechanism of S1P biological function.

Sphingosine-1-phosphate receptor 3 (S1P3) Agonist

Identify compounds which provide insight into the molecular mechanism of S1P biological function.

Sphingosine-1-phosphate receptor 3 (S1P3) Agonist

Identify compounds which provide insight into the molecular mechanism of S1P biological function.

Inhibitor of Schistosoma Mansoni Redox Cascade

Since no parasite-specific inhibitors of Prx are currently available, our overall goals can be summarized as: The identification of inhibitors of Schistosoma mansoni peroxiredoxins by conducting a high throughput screen of the Small Molecule Repository of the Molecular Libraries Screening Centers Network.

Inhibitor of Bacillus stearothermophilus Pyruvate Kinase

The research has two specific aims: 1) to engineer inhibitors for bacterial pyruvate kinases as leads for anti-infective agents. 2) To use structure-based, analogue synthesis and medicinal chemical principles to identify inhibitors of therapeutically useful PKs from infectious pathogens and other species including human where inhibitors have been considerably more difficult to obtain.