![]() performed cell-based DEL selections against the NK3 tachykinin GPCR. In 2011, the Bradley group selected a peptide nucleic acid (DNA mimetic)-encoded peptide library against cell surface receptors. Similar to prior work with cell-based selection of aptamers and phage-displayed peptides against cell surface target proteins, (21) there are a few examples of DEL selections which involve direct binding to live cells followed by repeated centrifugation and washing. (20)ĭNA-encoded library (DEL) technology is changing the landscape of drug discovery research, but its utility in developing lead compounds for membrane proteins remains underexplored. With this system, a G-protein-biased agonist of the glucagon-like peptide-1 receptor, PL5, was identified. Besides structure-based strategies, Lerner and co-workers developed an autocrine-based signaling system that, when paired with fluorescence-based cell sorting, identified biased agonists of GPCRs from genetically expressed peptide libraries. (19) Before the implementation of such campaigns, however, extensive structural information of the target is required. (3,17) Recently, a structure-based evolution approach explored the signaling profiles of PZM21 derivatives, which led to the discovery of FH210, an MOR agonist with improved selectivity and more pronounced G protein bias. Alternatively, researchers have employed computational and structure-based discovery approaches to develop biased agonists at GPCRs. While several biased ligands of ORs (e.g., PZM21, (17) TRV130 (18)) have been developed, the discovery of biased agonists remains challenging using traditional screening approaches. (15) While the current situation of biased agonists of OR targets is muddled, biased agonism has become a key component of drug development for many GPCR targets. Results with such biased agonists have been mixed, however, and many subsequent studies have directly contradicted the hypothesis that arrestin signaling is solely responsible for negative side effects. (14) This launched enthusiasm for development of G-protein-biased agonists of the MOR as safer analgesics. (13,14) A 2005 report of morphine treatment of β-arrestin 2 knockout mice showed both enhanced analgesia and dramatically reduced respiratory suppression and constipation. (12) For the MOR, some studies have indicated that the analgesic effects of agonists occur largely via G-protein signaling, while unwanted side effects occur through arrestin signaling. (11) ORs include the δ-, μ-, and κ-ORs (DOR, MOR, and KOR, respectively) and the nonclassical nociception OR (NOP). Opioid receptors (ORs) are among the class A subgroup of GPCRs. ![]() This approach should be generally applicable for the direct selection of chemical inducers of dimerization from DELs and expand the utility of DELs to enrich molecules with a specific and desired biochemical function. This approach was applied to δ-, μ-, and κ-opioid receptors and enabled the discovery of compound, a selective, G-protein-biased agonist of the κ-opioid receptor (EC 50 = 100 nM, E max = 82%, G i bias factor = 6.6). We used this selection with a DEL against opioid receptor GPCRs on living cells for the identification of small molecules that possess the specific function of activation of either β-arrestin or G protein signaling pathways. Herein, we report a split protein complementation approach that allows direct identification of DNA-linked molecules that induce the dimerization of two proteins. The use of DELs has been largely limited to affinity-based selections against purified protein targets, which identify binders only. DNA-encoded chemical libraries (DELs) dramatically improve the throughput of drug discovery by allowing a collective selection, rather than discrete screening, of large compound libraries. Challenges exist, however, in the development of new biased activators due, in part, to the low throughput of traditional screening approaches. Among these activities, biased agonists have potential to serve as both chemical probes to understand specific aspects of receptor signaling and therapeutic leads with more specific, desired activity. GPCR ligands can have a complex array of pharmacological activities. G protein-coupled receptors (GPCRs) are the largest superfamily of human membrane target proteins for approved drugs.
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