General context

The VIB Screening Core provides support in the development of miniaturized test systems/assays, which are relevant for a biological process-of-interest. In interaction with the scientists, the team of the VIB Screening Core enhances the technical reproducibility and robustness of an assay and makes it compatible with liquid handling and automation. Subsequently, a screening experiment is executed to identify substances that interfere with the assay. The aim is to accelerate the discovery of biologically active substances that can be developed into chemical probes to study biological processes and/or starting points to develop novel products for medical or agricultural applications.

The VIB Screening Core embeds the Centre for Bioassay Development and Screening (C-BIOS), a Ghent University Expertise Centre co-financed via the Special Research Fund (Bijzonder Onderzoeksfonds, BOF). C-BIOS unites the expertise and research infrastructure of VIB, Ghent University and the Cancer Research Institute Ghent (CRIG) and provides a centralized platform to efficiently support assay development and screening for the broad Life Sciences research community.

    Services

The VIB Screening Core provides services at the different levels of a screening project:

Assay development

• Assay transfer
• Assay optimization and miniaturization
• Evaluation of assay robustness
• Automation of assay protocols

Screening

• Primary screening
• Confirmation screening
• Data analysis and visualization
• Hit clustering and selection

Hit validation

• Compound reacquisition
• Dose-response analysis
• Dose-toxicity analysis
• Secondary screening
• Counter screening

Compound collection

The compound collection of the VIB Screening Core amounts to a total of almost 50,000 compounds, acquired from Enamine and MicroSource Discovery Systems. The core also manages a diverse subset of the collection of CBI (Chemistry to foster Biology Innovation), a chemistry cluster that brings together chemical expertise present in the universities of Antwerp, Leuven, Ghent, Hasselt and Brussels. The total collection comprises several subcollections, from which researchers can select to screen in their project:

• Drug-like structural diversity set: 20,160 compounds
• Pharmacological diversity set: 10,240 compounds
• Agro-like structural diversity set: 15,000 compounds
• Drug repurposing set (Spectrum): 2,400 compounds
• CBI set: 400 compounds

siRNA Library

The VIB Screening Core has acquired the Human siGENOME/ON-TARGETplus SMARTpool siRNA library (Dharmacon, Thermo Fisher Scientific) for siRNA screening. The library comprises nine distinct subsets, and each subset is available for screening separately. The different subsets include:

• GPCRs (390 genes)
• Ion Channels (352 genes)
• Kinases (720 genes)
• Phosphatases (257 genes)
• Proteases (480 genes)
• Ubiquitin Conjugation 1 (89 genes)
• Ubiquitin Conjugation 2 (115 genes)
• Ubiquitin Conjugation 3 (396 genes)
• Drug Targets (4,795 genes)

a set that contains additional genes of potential therapeutic interest, including genes involved in apoptosis, senescence, nucleic acid binding, autophagy, DNA repair and characterized nuclear receptors.

Instrumentation

Standalone devices

• Two Freedom EVO200 (Tecan) stations for versatile liquid handling applications in a broad range of microtiter plates and labware
• BD Pathway 855 (BD Biosciences) high-content analysis microscope (Methusalem grant Peter Vandenabeele) for low-throughput imaging of fixed or live cells

Integrated screening and detection platform

The platform is a joint investment between VIB, CRIG and Ghent University. To maximize capacity and to enable assay and screening automation, the following devices are integrated in the platform:

• Operetta CLS (PerkinElmer) high-content analysis confocal microscope for medium- to high-throughput acquisition and analysis of fluorescence, brightfield and digital phase contrast images. The device is equipped with environmental control for live cell imaging, laser-based autofocus, high power LED excitation and a highly sensitive 4.7 MP sCMOS camera.
• EnVision Xcite Multimode Plate Reader (PerkinElmer) for homogenous read-outs across a range of detection technologies including absorbance, fluorescence and luminescence.
• Expression 11000XL (Epson) scanner for low-resolution brightfield imaging.
• JANUS Mini VariSpan 4Tip (PerkinElmer) station for liquid handling in a broad range of microtiter plates and labware.
• JANUS Mini MDT (PerkinElmer) 96-channel head station for automated sample preparation in 96- and 384-well plates.
• Multidrop Combi (Thermo) for bulk dispensing of reagents and cells.
• HJ@SealPeelStation (HJ-Bioanalytik) for automated sealing and peeling of microtiter plates.
• LPX220 (LiCONiC) storage hotel for storing plates and tip boxes at ambient temperature.
• STX44 (LiCONiC) cell incubator with temperature and CO2 control.
• Laser operated scanner for barcode reading of microtiter plates.
• plate::handler II 6-axis robot (PerkinElmer) for integrating all platform devices.
• plate::works software (PerkinElmer) for scheduling automation protocols.

Software solutions

• CDD Vault (Collaborative Drug Discovery) for compound registry and data management.
• Columbus (PerkinElmer) for data analysis and data storage.
• Spotfire (TIBCO) for data visualization.
• Canvas (Schrödinger) for cheminformatic analysis.

    Publications

Hu Z, Vanderhaeghen R, Cools T, Wang Y, De Clercq I, Leroux O, Nguyen L, Belt K, Millar AH, Audenaert D, Hilson P, Small ID, Mouille G, Vernhettes S, Van Breusegem F, Whelan J, Höfte H, De Veylder L. Mitochondrial Defects Confer Tolerance against Cellulose Deficiency. Plant Cell. 2016 Aug 19;28(9):2276-90. [pmid]

Xuan W*, Audenaert D*, Parizot B, Möller BK, Njo MF, De Rybel B, De Rop G, Van Isterdael G, Mähönen AP, Vanneste S, Beeckman T. Root Cap-Derived Auxin Pre-patterns the Longitudinal Axis of the Arabidopsis Root. Curr Biol. 2015 May 18;25(10):1381-8. doi: 10.1016/j.cub.2015.03.046. Epub 2015 May 7. * equal contribution [pmid]

Van de Wouwer D, Vanholme R, Decou R, Goeminne G, Audenaert D, Nguyen L, Höfer R, Pesquet E, Vanholme B, Boerjan W. Chemical Genetics Uncovers Novel Inhibitors of Lignification, Including p-Iodobenzoic Acid Targeting CINNAMATE-4-HYDROXYLASE. Plant Physiol. 2016 Sep;172(1):198-220. [pmid]

Dejonghe W, Kuenen S, Mylle E, Vasileva M, Keech O, Viotti C, Swerts J, Fendrych M, Ortiz-Morea FA, Mishev K, Delang S, Scholl S, Zarza X, Heilmann M, Kourelis J, Kasprowicz J, Nguyen le SL, Drozdzecki A, Van Houtte I, Szatmári AM, Majda M, Baisa G, Bednarek SY, Robert S, Audenaert D, Testerink C, Munnik T, Van Damme D, Heilmann I, Schumacher K, Winne J, Friml J, Verstreken P, Russinova E. Mitochondrial uncouplers inhibit clathrin-mediated endocytosis largely through cytoplasmic acidification. Nat Commun. 2016 Jun 8;7:11710. [pmid]

Xuan W, Band LR, Kumpf RP, Van Damme D, Parizot B, De Rop G, Opdenacker D, Möller BK, Skorzinski N, Njo MF, De Rybel B, Audenaert D, Nowack MK, Vanneste S, Beeckman T. Cyclic programmed cell death stimulates hormone signaling and root development in Arabidopsis. Science. 2016 Jan 22;351(6271):384-7. [pmid]

Kerchev P, Mühlenbock P, Denecker J, Morreel K, Hoeberichts FA, Van Der Kelen K, Vandorpe M, Nguyen L, Audenaert D, Van Breusegem F. Activation of auxin signalling counteracts photorespiratory H2O2-dependent cell death. Plant Cell Environ. 2015 Feb;38(2):253-65. [pmid]