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Introduction
Receptor for advanced glycation end products (RAGE) is a pattern recognition receptor that binds multiple ligands, including AGE (1), S100 proteins (2), lipopolysaccharides (3), phosphatidylserine (4), amyloid-β (Aβ) (5), and high mobility group box (HMGB)-1 (6). Interactions of these diverse ligands with RAGE result in intracellular signaling, including nuclear factor κ-B (NF-κB) activation, which results in pathogenic processes such as diabetic complications (7), inflammatory diseases, Alzheimer's disease (AD) (8) and cancer (9). Takeuchi et al (10) demonstrated that RAGE expression in HT1080 human fibrosarcoma cells induced tumor cells to proliferate, migrate, invade and metastasize. HMGB-1 was revealed to induce RAS-related C3 botulinum toxin substrate (Rac)1 and cell division control protein 42 homolog (Cdc42) functions in RAGE-expressing HT1080 fibrosarcoma cells (10). Epidemiological studies also demonstrated that RAGE expression was associated with tumor malignancies of the stomach (11), colon and rectum (12–14), prostate (15), breast (16) and bone (17). Therefore, these previous studies suggested that RAGE represents a potential therapeutic target, and that inhibiting RAGE may be useful to anticancer strategies.
Previously, Sakai et al (18) developed a novel drug design system, involving the conversion of optimized binding peptide to non-peptidic small molecules by structure-based virtual screening (SBVS), followed by optimization of the small molecules using a structure-based drug design system, namely conversion to small molecules through optimized-peptide strategy (COSMOS). Using this strategy, the most optimized binding peptide is first computationally designed on a hot spot in the target protein. Subsequently, the optimized binding peptide may be converted to small molecules by SBVS based on its pharmacophore. Then, the selected candidates are evaluated using in vitro assays. Therefore, this strategy decreases the cost and time required to search for effective lead compounds, for drug design and for optimization (18). The present study identified a RAGE inhibitor, papaverine, using this drug design system. Papaverine is an opiate alkaloid, originally isolated from the plant Papaver somniferum and now synthetically produced as a direct-acting smooth muscle relaxant. Its mechanism of action may be associated with non-selective inhibition of phosphodiesterases and direct inhibition of calcium channels (19,20).
The present study assessed whether papaverine functioned as a RAGE inhibitor using in vitro cell culture systems of RAGE- and dominant-negative (dn)RAGE-expressing HT1080 fibrosarcoma cells.
Materials and methods
Papaverine
Papaverine hydrochloride (molecular weight,...