Irreversible Activation and Stabilization of Soluble Guanylate Cyclase by the Protoporphyrin IX Mimetic Cinaciguat

Belonging to the class of so-called soluble guanylate cyclase (sGC) activators, cinaciguat and BAY 60-2770 are interesting therapeutic tools for the treatment of various cardiovascular pathologies. The drugs are supposed to preferentially stimulate oxidized or heme-depleted, but not native sGC. Since this concept has been challenged by studies demonstrating complete relaxation of nondiseased vessels, this study was designed to reinvestigate the mode of action in greater detail. To this purpose, the effect of cinaciguat was studied on vessel tone of porcine coronary arteries and rat thoracic aortas. Organ bath studies showed that the compound caused time- and concentration-dependent relaxation of precontracted vessels with a maximal effect observed at 90 minutes. The dilatory response was not affected by extensive washout of the drug. Cinaciguat-induced vasodilation was associated with a time- and concentration-dependent increase of cGMP levels. Experiments with purified sGC in the presence of Tween 20 showed that cinaciguat activates the heme-free enzyme in a concentration-dependent manner with an EC(50) value of ~0.2 μM and maximal cGMP formation at 10 μM. By contrast, the effect of cinaciguat on 1H-[1,2,4]oxadiazolo-[4,3-a]quinoxalin-1-one–oxidized (ferric) sGC was moderate, reaching ~10%–15% of maximal activity. Dilution experiments of cinaciguat/Tween 20–preincubated sGC revealed the irreversible character of the drug. Assuming a sensitive balance between heme-free, ferric, and nitric oxide–sensitive ferrous sGC in cells and tissues, we propose that cinaciguat by virtue of its irreversible mode of action is capable of shifting this equilibrium toward the heme-free apo-sGC species.