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An RNA molecule that specifically inhibits G-protein-coupled receptor kinase 2 in vitro

¹ Life and Medical Sciences Bonn, Program Unit Chemical Biology, c/o Kekulé-Institute for Organic Chemistry and Biochemistry, University of Bonn, 53121 Bonn, Germany
² NascaCell Technologies AG, 81377 Munich, Germany
³ Institute of Pharmacology and Toxicology, University of Würzburg, 97078 Würzburg, Germany
⁴ GPC Biotech, 82152 Martinsried, Germany
⁵ School of Pharmacy, University of Reading, Whiteknights, Reading RG6 6AJ, United Kingdom

G-protein-coupled receptors are desensitized by a two-step process. In a first step, G-protein-coupled receptor kinases (GRKs) phosphorylate agonist-activated receptors that subsequently bind to a second class of proteins, the arrestins. GRKs can be classified into three subfamilies, which have been implicated in various diseases. The physiological role(s) of GRKs have been difficult to study as selective inhibitors are not available. We have used SELEX (systematic evolution of ligands by exponential enrichment) to develop RNA aptamers that potently and selectively inhibit GRK2. This process has yielded an aptamer, C13, which bound to GRK2 with a high affinity and inhibited GRK2-catalyzed rhodopsin phosphorylation with an IC₅₀ of 4.1 nM. Phosphorylation of rhodopsin catalyzed by GRK5 was also inhibited, albeit with 20-fold lower potency (IC₅₀ of 79 nM). Furthermore, C13 reveals significant specificity, since almost no inhibitory activity was detectable testing it against a panel of 14 other kinases. The aptamer is two orders of magnitude more potent than the best GRK2 inhibitors described previously and shows high selectivity for the GRK family of protein kinases.

Keywords: in vitro selection; SELEX; aptamer; GRK2; kinase inhibitor

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