Probing the signalling complexity of tumor cells by functional screening of peptide and small molecule libraries

Current concepts for target identification in oncology usually involve the analysis of either genomic, transcriptomic, or proteomic data sets to determine expression in tumor versus normal tissues. While biochemical and morphological analysis of target protein expression is believed to yield the most reliable information on the validity of a target, specific antibodies for tissue staining and Western blotting are often difficult to obtain, especially for GPCRs and other membrane proteins.

Functional screening of sets of tumor cells may be able to fill this gap and provide valuable knowledge about the expression and biological function of GPCRs and other receptors in human cancer. We have used a library of more than 2000 peptides and small molecules to characterize and obtain a functional profile of receptors in sets of colorectal and neuroendocrine cancer cell lines. All peptides were derived from naturally occuring sequences, while small molecules represented approved drugs. Label-free dynamic mass redistribution technology was utilized for primary screening of activation profiles to cover a broad spectrum of possible signalling pathways. Initial findings were subsequently confirmed by a number of established assays such as calcium mobilization, cAMP, and phospho-ERK assays. Characteristic receptor profiles were identified for both neuroendocrine as well as colorectal cancer, providing new target candidates for these tumor entities. Secondary confirmation also included the use of known inhibitors and quantitative characterization of activation and binding (EC₅₀/IC₅₀).