title: Screening Pools of Compounds against Multiple Endogenously Expressed Targets in a Chemoproteomics Binding Assay.
abstract: Chemoproteomics-based competition-binding assays allow the screening of compounds against endogenous proteins in cell or tissue extracts, but these assays are hampered by low throughput and high cost. Using compound pools rather than single compounds in a screening campaign holds the promise of increased efficiency and substantial cost reduction. Previous attempts to screen compounds in pools often fell short due to complex data tracking, deconvolution issues, compound interferences, and automation problems. The desire to screen compounds in a high-throughput chemoproteomics format sparked a reassessment of compound pooling. Through the integration of acoustic dispensing, we enabled a flexible pooling process, allowing mixture creation by combining randomized or specific samples to create defined pools. Automation enabled end-to-end tracking, using barcode scan check points and output files to track data and ensure integrity during the mixture creation process. The compound pooling approach proved to be highly compatible with the chemoproteomics assay technology. Pools of 10 compounds in a single well did not show compound interference effects or increased false-positive/negative rates. In the present study, four targets, TBK1, PI3Kδ, PI3Kγ, and mTOR, were screened using a chemoproteomics approach against pools of 10 compounds per well, resulting in robust hit identification.
authors.affiliation: Cellzome GmbH, Molecular Discovery Research, GlaxoSmithKline, Heidelberg, Germany.
authors.forename: Elsa
authors.initials: E
authors.lastname: Salzer
authors.affiliation: GlaxoSmithKline, SMTech, Collegeville, PA, USA.
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Nixon
authors.affiliation: Cellzome GmbH, Molecular Discovery Research, GlaxoSmithKline, Heidelberg, Germany.
authors.forename: Gerard
authors.initials: G
authors.lastname: Drewes
authors.affiliation: Cellzome GmbH, Molecular Discovery Research, GlaxoSmithKline, Heidelberg, Germany.
authors.forename: Friedrich
authors.initials: F
authors.lastname: Reinhard
authors.affiliation: Cellzome GmbH, Molecular Discovery Research, GlaxoSmithKline, Heidelberg, Germany.
authors.forename: Giovanna
authors.initials: G
authors.lastname: Bergamini
authors.affiliation: Cellzome GmbH, Molecular Discovery Research, GlaxoSmithKline, Heidelberg, Germany Christina.e.rau@gsk.com.
authors.forename: Christina
authors.initials: C
authors.lastname: Rau
mesh_terms: D000162:Acoustics; D057205:Automation, Laboratory / Q000379:methods; D020811:Biomedical Technology / Q000379:methods*; D002457:Cell Extracts / Q000737:chemistry*; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D057166:High-Throughput Screening Assays / Q000379:methods*; D011485:Protein Binding; D020543:Proteome / Q000032:analysis*; D012996:Solutions
chemical_list: D002457:Cell Extracts; D020543:Proteome; D012996:Solutions
keywords: acoustic dispenser; chemoproteomics; compound pooling; high-throughput screening (HTS)