High Throughput and High Content Screening Capabilities of the University of Cincinnati Drug Discovery Center.

The Drug Discovery Center collaborates with a wide range of academic and industrial research centers to facilitate the identification of active small molecules with high potential for use as biological probes or as starting points for drug discovery programs. The DDC operates state-of-the-art high throughput and high content screening instrumentation and a diverse 350,000 compound library. The center's personnel provide collaborators with advice in assay design, analytical technology selection, and library design via cheminformatics and/or structure-based approaches. Typical programs are exemplified by an HTS program targeting the identification of novel atypical PKC inhibitors for potential use in cancer and an HCS program targeting the identification of stimulators of the differentiation of oligodendrocyte progenitor cells to oligodendrocytes for potential use in multiple sclerosis. Activities of atypical PKCs, PKCiota and PKCzeta, were measured using both fluorescent detection of ADP production and MALDI-TOF detection of substrate phosphorylation. 30,000 compounds were screened for their effect on PKC activity using these orthogonal detection methods to identify potential inhibitors. Oligodendrocyte differentiation was measured in a High Content Screen by pairing a selective marker of oligodendrocytes with Alexa Fluor 488 secondary antibody for the detection of mature ODs. DAPI stain was used for nucleus detection. Confocal microscope images were acquired and analyzed using an algorithm for neurite outgrowth adapted for the characterization of oligodendrocyte processes. Three measurements: Mean Maximum Process Length per Cell, Mean Process Signal Intensity per Cell, and Percentage Differentiated Cells per Well, were quantified by the image analysis script and afforded a statistically significant separation of promoter controls and inhibitor controls from non-treated (neutral) controls with Mean Maximum Process Length as measurement.