The receptor for advanced glycation end-products (RAGE) and its ligands are important players in pathological conditions such as diabetes, neurodegenerative diseases, and cancers. RAGE is a cell surface molecule of the immunoglobulin superfamily. Alternatively spliced variants lacking either only the intracellular domain or both the intracellular and the transmembrane domains are also expressed in some tissues. RAGE functions in development and inflammation. It interacts with multiple ligands including members of the S100 protein family, high mobility group box-1 protein (HMGB1), and advanced glycation end products. It has been shown that the activation of RAGE triggers a positive feedback loop that stimulates the expression of both the receptor and some of its ligands, thus amplifying their deleterious effects (1-3). Therefore, the targeting of the specific interactions of RAGE with its ligands is a potential method to control these diseases. Toward this aim the project proposes an in vitro assay to screen for new molecules that block the interaction of the receptor with its pathological ligands.
The project aims to deliver purified unlabeled and fluorescently-labeled soluble RAGE and several of its ligands, a protocol for competitive binding assays based on the obtained products, and furthermore, an automated and validated technology which provides a rapid and easy way for screening of libraries of compounds.
References:
CSIII
1998 BSc degree in Biochemistry, Faculty of Biology- University of Bucharest, 2000 MSc degree in Molecular Biology, Faculty of Biology- University of Bucharest 2000-2001 Research assistant- Institute of Biochemistry PhD 2007, Department of Cell Biology, University Medical Center Utrecht, The Netherlands, “Rab4 and its interacting proteins in endosome function” 2007-present Researcher- Institute of Biochemistry Grants received: 1. Intracellular trafficking and signaling properties of the receptor for advanced glycation end produscts (RAGE) under pathological conditions, 2007-2009, Reintegration grant RP-11, contract nr 14/1.10.2007 2. Development of More...
| Project Team: |
Carmen Tanase, PhD Florin Pastrama, PhD |
| Project Leader: | Dr. Ioana Popa Institute of Biochemistry Molecular Cell Biology Department Splaiul Independentei 296, Bucuresti Email: ipopa@biochim.ro |
Scientific Results:
Our project aimed at the development of a method for rapid and sensitive detection of RAGE-ligand interactions. We obtained various recombinant variants of soluble RAGE and some of its ligands. Using these proteins we developed, optimized and validated two fluorescence based methods suitable for high-throughput of compounds. One method relies on the direct measurement of the fluorescent ligand binding to pre-adsorbed sRAGE in microplates, whereas in the other method a FRET signal is generated in solution upon RAGE, indirectly fluorescently labeled, interacted with a fluorescent ligand. The methods were further used in a screen of a small library of natural compounds, which led to the identification of three potential RAGE-ligand binding inhibitors.
Dissemination:
Poster
Title:"New Tools for Discovering Inhibitors of the Receptor for Advanced Glycation End Products"
Authors: Carmen A. Tanase, Florin Pastrama, and Ioana L. Popa* (*corresponding author)
Meeting: International Annual Meeting of the Romanian Society of Biochemistry and Molecular Biology, June 8-9 2017, Timisoara, Romania
Abstract published in New Front. Chem. (2017) Vol.26, Iss.2, S2_P11
Poster
Title:"High-throughput assay for discovery of new inhibitors of S100-RAGE binding"
Authors: Ioana L. Popa*, Carmen A. Tanase, Florin Pastrama (*corresponding author)
Meeting:International Annual Meeting of the Romanian Society of Biochemistry and Molecular Biology, September 5-7 2018, Bucharest, Romania
Poster
Title:"Homogenous TR-FRET-based high-throughput screening assay to identify inhibitors of RAGE-ligand interaction"
Authors: Carmen A. Tanase, Florin Pastrama, and Ioana L. Popa* (*corresponding author)
Meeting: International Annual Meeting of the Romanian Society of Biochemistry and Molecular Biology, September 5-7 2018, Bucharest, Romania
Oral presentation
Title: " High-throughput screening to identify RAGE inhibitors for cancer therapy"
Authors: Ioana Popa, Carmen Tanase, Florin Pastrama,
Meeting: International Annual Meeting of the Romanian Society of Biochemistry and Molecular Biology, September 26-27 2019, Iasi, Romania
Romanian Academy