title: Stimulators of the soluble guanylyl cyclase: promising functional insights from rare coding atherosclerosis-related GUCY1A3 variants.
abstract: Stimulators of the soluble guanylyl cyclase (sGC) are emerging therapeutic agents in cardiovascular diseases. Genetic alterations of the GUCY1A3 gene, which encodes the α1 subunit of the sGC, are associated with coronary artery disease. Studies investigating sGC stimulators in subjects with CAD and carrying risk-related variants in sGC are, however, lacking. Here, we functionally investigate the impact of coding GUCY1A3 variants on sGC activity and the therapeutic potential of sGC stimulators in vitro. In addition to a known loss-of-function variant, eight coding variants in GUCY1A3 were cloned and expressed in HEK 293 cells. Protein levels and dimerization capability with the β1 subunit were analysed by immunoblotting and co-immunoprecipitation, respectively. All α1 variants found in MI patients dimerized with the β1 subunit. Protein levels were reduced by 72 % in one variant (p < 0.01). Enzymatic activity was analysed using cGMP radioimmunoassay after stimulation with a nitric oxide (NO) donor. Five variants displayed decreased cGMP production upon NO stimulation (p < 0.001). The addition of the sGC stimulator BAY 41-2272 increased cGMP formation in all of these variants (p < 0.01). Except for the variant leading to decreased protein level, cGMP amounts reached the wildtype NO-induced level after addition of BAY 41-2272. In conclusion, rare coding variants in GUCY1A3 lead to reduced cGMP formation which can be rescued by a sGC stimulator in vitro. These results might therefore represent the starting point for discovery of novel treatment strategies for patients at risk with coding GUCY1A3 variants.
authors.affiliation: Klinik für Herz- und Kreislauferkrankungen, Deutsches Herzzentrum München, Technische Universität München, Lazarettstr. 36, 80636, Munich, Germany.
authors.forename: Jana
authors.initials: J
authors.lastname: Wobst
authors.affiliation: Klinik für Herz- und Kreislauferkrankungen, Deutsches Herzzentrum München, Technische Universität München, Lazarettstr. 36, 80636, Munich, Germany.
authors.forename: Simon
authors.initials: S
authors.lastname: von Ameln
authors.affiliation: Klinik für Herz- und Kreislauferkrankungen, Deutsches Herzzentrum München, Technische Universität München, Lazarettstr. 36, 80636, Munich, Germany.
authors.forename: Bernhard
authors.initials: B
authors.lastname: Wolf
authors.affiliation: Department of Proteomics and Signal Transduction, Max-Planck-Institut für Biochemie, Am Klopferspitz 18, 82152, Martinsried, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Wierer
authors.affiliation: Klinik für Herz- und Kreislauferkrankungen, Deutsches Herzzentrum München, Technische Universität München, Lazarettstr. 36, 80636, Munich, Germany.
authors.forename: Tan An
authors.initials: TA
authors.lastname: Dang
authors.affiliation: Klinik für Herz- und Kreislauferkrankungen, Deutsches Herzzentrum München, Technische Universität München, Lazarettstr. 36, 80636, Munich, Germany.
authors.forename: Hendrik B
authors.initials: HB
authors.lastname: Sager
authors.affiliation: Institut für Integrative und Experimentelle Genomik, Universität zu Lübeck, Maria-Goeppert-Str. 1, 23562, Lübeck, Germany.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Tennstedt
authors.affiliation: Klinik für Herz- und Kreislauferkrankungen, Deutsches Herzzentrum München, Technische Universität München, Lazarettstr. 36, 80636, Munich, Germany.|Deutsches Zentrum für Herz-Kreislauf-Forschung (DZHK) e.V., Partner Site Munich Heart Alliance, 80636, Munich, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Hengstenberg
authors.affiliation: Institut für Pharmakologie und Toxikologie, Ruhr-Universität Bochum, Medizinische Fakultät MA N1/39, 44780, Bochum, Germany.
authors.forename: Doris
authors.initials: D
authors.lastname: Koesling
authors.affiliation: Physiologisches Institut, Julius-Maximilians-Universität Würzburg, Röntgenring 9, 97070, Würzburg, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Friebe
authors.affiliation: Institut für Laboratoriumsmedizin, Deutsches Herzzentrum München, Technische Universität München, Lazarettstr. 36, 80636, Munich, Germany.
authors.forename: Siegmund L
authors.initials: SL
authors.lastname: Braun
authors.affiliation: Institut für Integrative und Experimentelle Genomik, Universität zu Lübeck, Maria-Goeppert-Str. 1, 23562, Lübeck, Germany.|Deutsches Zentrum für Herz-Kreislauf-Forschung (DZHK) e.V., Partner Site Hamburg/Kiel/Lübeck, 23562, Lübeck, Germany.
authors.forename: Jeanette
authors.initials: J
authors.lastname: Erdmann
authors.affiliation: Klinik für Herz- und Kreislauferkrankungen, Deutsches Herzzentrum München, Technische Universität München, Lazarettstr. 36, 80636, Munich, Germany. schunkert@dhm.mhn.de.|Deutsches Zentrum für Herz-Kreislauf-Forschung (DZHK) e.V., Partner Site Munich Heart Alliance, 80636, Munich, Germany. schunkert@dhm.mhn.de.
authors.forename: Heribert
authors.initials: H
authors.lastname: Schunkert
authors.affiliation: Klinik für Herz- und Kreislauferkrankungen, Deutsches Herzzentrum München, Technische Universität München, Lazarettstr. 36, 80636, Munich, Germany. thorsten.kessler@tum.de.
authors.forename: Thorsten
authors.initials: T
authors.lastname: Kessler
mesh_terms: D000328:Adult; D000818:Animals; D003324:Coronary Artery Disease / Q000235:genetics*; D006152:Cyclic GMP / Q000096:biosynthesis* / Q000235:genetics; D020022:Genetic Predisposition to Disease / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D051379:Mice; D008875:Middle Aged; D020125:Mutation, Missense; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide; D011720:Pyrazoles / Q000494:pharmacology; D011725:Pyridines / Q000494:pharmacology; D011863:Radioimmunoassay; D000071756:Soluble Guanylyl Cyclase / Q000235:genetics* / Q000378:metabolism; D055815:Young Adult
chemical_list: C423582:3-(4-Amino-5-cyclopropylpyrimidine-2-yl)-1-(2-fluorobenzyl)-1H-pyrazolo(3,4-b)pyridine; C000605661:GUCY1A1 protein, human; D011720:Pyrazoles; D011725:Pyridines; D000071756:Soluble Guanylyl Cyclase; D006152:Cyclic GMP
keywords: Coronary artery disease; Myocardial infarction; Pharmacogenetics; Soluble guanylyl cyclase