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Br J Pharmacol. 2011 Sep;164(2b):499-506. doi: 10.1111/j.1476-5381.2011.01291.x.

The indolocarbazole, Gö6976, inhibits guanylyl cyclase-A and -B.

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  • 1Department of Pharmacology, University of Minnesota-Twin Cities, Minneapolis, MN, USA.

Abstract

BACKGROUND AND PURPOSE:

Atrial natriuretic peptide (ANP) and B-type natriuretic peptide (BNP) decrease vascular volume and pressure by activating guanylyl cyclase-A (GC-A). C-type natriuretic peptide (CNP) activation of guanylyl cyclase-B (GC-B) stimulates long bone growth. This study investigated the effects of the indolocarbazole, Gö6976, on the guanylyl cyclase activity of GC-A and GC-B as a first step towards developing small molecule regulators of these enzymes.

EXPERIMENTAL APPROACH:

Whole cell cGMP concentrations or ³²P-cGMP accumulation in membrane preparations measured the effects of indolocarbazoles on the enzymatic activity GC-A and GC-B from transfected 293T or endogenously expressing 3T3-L1 cells.

KEY RESULTS:

Gö6976 blocked cellular CNP-dependent cGMP elevations in 293T-GC-B cells. The t(½) for Gö6976 inhibition was 7 s and IC₅₀ was 380 nM. Gö6976 increased the EC₅₀ for CNP 4.5-fold, but increasing the CNP concentration did not overcome the inhibition. Half of the inhibition was lost 1 h after removal of Gö6976 from the medium. Cellular exposure to Gö6976 reduced basal and natriuretic peptide-dependent, but not detergent-dependent, GC-A and GC-B activity. Inhibition was also observed when Gö6976 was added directly to the cyclase assay. A constitutively phosphorylated form of GC-B was similarly inhibited.

CONCLUSIONS AND IMPLICATIONS:

These data demonstrate that Gö6976 potently, rapidly and reversibly inhibited GC-A and GC-B via a process that did not require intact cells, known phosphorylation sites or inactivation of all catalytic sites. This is the first report of an intracellular inhibitor of a transmembrane guanylyl cyclase and the first report of a non-kinase target for Gö6976.

© 2011 The Authors. British Journal of Pharmacology © 2011 The British Pharmacological Society.

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