Clin Res Cardiol 107, Suppl 1, April 2018

The Non-steroidal MR-Antagonist Finerenone Induces Differential Cardiomyocytic Gene Regulation and Nuclear Cofactor Binding

J. Grune1, N. Beyhoff1, E. Smeir1, R. Chudek1, A. Blumrich1, Z. Ban1, S. Brix1, I. Betz1, M. Schupp1, A. Foryst-Ludwig1, R. Klopfleisch2, P. Stawowy3, W. Kübler4, R. Houtman5, P. Kolkhof6, U. Kintscher1

1Center for Cardiovasc. Research, Inst. f. Pharmakologie, Charité - Universitätsmedizin Berlin, Berlin; 2Department of Veterinary Medicine, Institute of Veterinary Pathology, Berlin; 3Klinik für Innere Medizin - Kardiologie, Deutsches Herzzentrum Berlin, Berlin; 4Institut für Physiologie, Deutsches Herzzentrum Berlin, Berlin; 5Pamgene International B.V, s-Hertogenbosch, NL; 6Bayer Vital GmbH, Leverkusen;

Background: Ligand-mediated activation of mineralocorticoid receptors (MR) induces deleterious cardiovascular processes like fibrosis. Vice versa, therapy with MR antagonists (MRA) is highly efficient in chronic heart failure. Recently, highly selective, non-steroidal MRAs such as finerenone (FIN) have been developed. We could show in previous studies that FIN mediates anti-fibrotic actions in a mouse model of isolated subendocardial fibrosis. To further understand the molecular mechanism of FIN´s cardiac actions, we compared FIN and the steroidal MRA eplerenone (EPL) with regards to selective MR-cofactor modulation, and subsequent selective transcriptional inhibition of pro-fibrotic target genes. Methods: Cell culture experiments were carried out, using H9C2/MR+ cells, a rat cardiomyocyte cell line stably expressing the full-length rat MR. Cells were treated for 24h +/- aldosterone (100nM). Pre-incubation +/- distinct MRA concentrations started 30 min prior to aldosterone treatment. After stimulation, cells were harvested for further gene expression analysis. Direct binding between the MR ligand-binding domain (LBD) and co-regulator nuclear receptor boxes were determined by MARCoNI (Microarray Assay for Real-time Coregulator-Nuclear receptor Interaction). A concentration series containing 7 dilutions (3-fold) of FIN (-8.3 to -5 logM) and EPL (-7.3 to 4 logM) was tested in agonist mode (vs. solvent, i.e. 2% DMSO) and antagonist mode (vs. -8.2 logM aldosterone). Results: Stimulation of H9C2/MR+ with ALDO for 24h resulted in increased tenascin X (TNX) gene expression, a known MR-target gene and matricellular protein involved in collagen deposition. ALDO-MR-mediated TNX-regulation in H9C2/MR+ cells was exclusively inhibited by FIN, in a dose-dependent manner, and not by EPL, even not at 500-fold higher concentrations than the chosen ALDO-concentration. In the presence of ALDO (antagonistic mode) both MRAs dose-dependently inhibited the binding of the coactivators SRC1, TRAP220, TIF1α, PGC1α, and ASC2 using the MARCoNI assay. The dose response curves for the inhibition of ALDO-induced MR-coactivator binding are shifted to the left in the presence of FIN by at least one log step compared to EPL indicating a higher antagonistic potency of FIN. Furthermore, FIN induced a complete inhibition of ALDO-mediated MR-coactivator binding, whereas inhibition by EPL was only partially active at the respective maximum inhibition. Conclusions: These data identify important differences of EPL and FIN regarding their ability to modulate MR-cofactor binding and downstream gene expression. These processes were associated with ligand-specific MR-dependent cardiac gene regulation. Thus, we could identify a new molecular mechanism of differential MR-cofactor-modulation, differential cardiac gene expression of a pro-fibrotic target gene potentially leading to a more pronounced anti-fibrotic efficacy of non-steroidal versus steroidal MR blockade in a preclinical model of cardiac fibrosis.

http://www.abstractserver.de/dgk2018/jt/abstracts//P396.htm