Biochemical and Structural Characterization of a Peptidic Inhibitor of the YAP:TEAD Interaction That Binds to the α-Helix Pocket on TEAD.

The TEAD transcription factors are the most distal elements of the Hippo pathway, and their transcriptional activity is regulated by several proteins, including YAP. In some cancers, the Hippo pathway is deregulated and inhibitors of the YAP:TEAD interaction are foreseen as new anticancer drugs. The binding of YAP to TEAD is driven by the interaction of an α-helix and an Ω-loop present in its TEAD-binding domain with two distinct pockets at the TEAD surface.

Gene-signature-derived ICs/ECs reflect the potency of causative upstream targets and downstream phenotypes.

Multiplexed gene-signature-based phenotypic assays are increasingly used for the identification and profiling of small molecule-tool compounds and drugs. Here we introduce a method (provided as R-package) for the quantification of the dose-response potency of a gene-signature as EC and IC values. Two signaling pathways were used as models to validate our methods: beta-adrenergic agonistic activity on cAMP generation (dedicated dataset generated for this study) and EGFR inhibitory effect on cancer cell viability.

Helios: History and Anatomy of a Successful In-House Enterprise High-Throughput Screening and Profiling Data Analysis System.

We describe the main characteristics of the Novartis Helios data analysis software system (Novartis, Basel, Switzerland) for plate-based screening and profiling assays, which was designed and built about 11 years ago. It has been in productive use for more than 10 years and is one of the important standard software applications running for a large user community at all Novartis Institutes for BioMedical Research sites globally. A high degree of automation is reached by embedding the data analysis capabilities into a software ecosystem that deals with the management of samples, plates, and result data files, including automated data loading.

Theoretical and experimental relationships between percent inhibition and IC50 data observed in high-throughput screening.

The four-parameter logistic Hill equation models the theoretical relationship between inhibitor concentration and response and is used to derive IC(50) values as a measure of compound potency. This relationship is the basis for screening strategies that first measure percent inhibition at a single, uniform concentration and then determine IC(50) values for compounds above a threshold. In screening practice, however, a "good" correlation between percent inhibition values and IC(50) values is not always observed, and in the literature, there seems confusion about what correlation even to expect.

Comparison of multivariate data analysis strategies for high-content screening.

High-content screening (HCS) is increasingly used in biomedical research generating multivariate, single-cell data sets. Before scoring a treatment, the complex data sets are processed (e.g.

Compound set enrichment: a novel approach to analysis of primary HTS data.

The main goal of high-throughput screening (HTS) is to identify active chemical series rather than just individual active compounds. In light of this goal, a new method (called compound set enrichment) to identify active chemical series from primary screening data is proposed. The method employs the scaffold tree compound classification in conjunction with the Kolmogorov-Smirnov statistic to assess the overall activity of a compound scaffold.

Assay data quality assessment.

An overview of the characteristics of classical and outlier-resistant data summaries is provided. The latter are important because outlier data can skew results and decisions based on them. The simple data summaries are the basis for all composite assay and screening data quality measures, for example, the signal-to-noise ratio, signal-to-background ratio, assay and screening window coefficients Z ' and Z, or strictly standardized mean difference (SSMD).

Comparison of variability and sensitivity between nuclear translocation and luciferase reporter gene assays.

High-content screening (HCS), a technology based on subcellular imaging by automated microscopy and sophisticated image analysis, has emerged as an important platform in small-molecule screening for early drug discovery. To validate a subcellular imaging assay for primary screening campaigns, an HCS assay was compared with a non-image-based readout in terms of variability and sensitivity. A study was performed monitoring the accumulation of the forkhead transcription factor of the O subfamily (FOXO3a) coupled with green fluorescent protein in the nucleus of human osteosarcoma (U-2 OS) cells.

Recommendations for the reduction of compound artifacts in time-resolved fluorescence resonance energy transfer assays.

Time-resolved (TR) fluorescence resonance energy transfer (FRET) is a widely accepted technology for high throughput screening (HTS), being able to detect and quantify the interactions of specific biomolecules in a homogeneous format. TR-FRET has several advantages for HTS applications that reduce assay artifacts such as compound interference. However, in some cases artifacts due to compound autofluorescence, color quenching, or signal stability are still observed.

A multiparameter screening assay to assess the cytokine-induced expression of endothelial cell adhesion molecules.

Compounds which inhibit endothelial cell inflammatory responses are believed to be of therapeutic value. The cell adhesion molecules E-selectin, ICAM-1, and VCAM-1 play important roles in inflammatory reactions by mediating leukocyte-endothelial interactions. To identify compounds which inhibit the expression of these adhesion molecules following cytokine stimulation we developed an assay which measures E-selectin, ICAM-1, and VCAM-1 in the same experiment.