sGC Stimulator Research Articles

The nitric oxide-soluble guanylate cyclase-cGMP pathway in pulmonary hypertension: from PDE5 to soluble guanylate cyclase.

The nitric oxide (NO)-soluble guanylate cyclase (sGC)-cyclic guanosine monophosphate (cGMP) pathway plays a key role in the pathogenesis of pulmonary hypertension (PH). Targeted treatments include phosphodiesterase type 5 inhibitors (PDE5i) and sGC stimulators. The sGC stimulator riociguat is approved for the treatment of pulmonary arterial hypertension (PAH) and chronic thromboembolic pulmonary hypertension (CTEPH). sGC stimulators have a dual mechanism of action, enhancing the sGC response to endogenous NO and directly stimulating sGC, independent of NO. This increase in cGMP production via a dual mechanism differs from PDE5i, which protects cGMP from degradation by PDE5, rather than increasing its production. sGC stimulators may therefore have the potential to increase cGMP levels under conditions of NO depletion that could limit the effectiveness of PDE5i. Such differences in mode of action between sGC stimulators and PDE5i could lead to differences in treatment efficacy between the classes. In addition to vascular effects, sGC stimulators have the potential to reduce inflammation, angiogenesis, fibrosis and right ventricular hypertrophy and remodelling. In this review we describe the evolution of treatments targeting the NO-sGC-cGMP pathway, with a focus on PH.

Soluble guanylate cyclase stimulators for heart failure: a network meta-analysis and subgroup analyses of reduced and preserved ejection fraction

BackgroundSoluble guanylate cyclase (sGC) stimulators have been investigated for heart failure (HF) in several randomized controlled trials (RCTs). However, its place in the management guidelines of either HFrEF or HfpEF is still inconclusive.MethodsWe conducted a network meta-analysis synthesizing RCTs investigating sGC for HF management, which were retrieved by systematically searching five databases until January 24th, 2023. Dichotomous outcomes were pooled using risk ratio (RR) along with confidence interval (CI).ResultsEight RCTs with a total of 7307 patients were included. Vericiguat 10 mg significantly reduced the composite cardiovascular (CVS) mortality and HF hospitalization in HF (RR: 0.88, 95% CI [0.79; 0.98]) and in HFrEF (RR: 0.87, 95% CI [0.78; 0.97]); however, it was not effective in HFpEF (RR: 0.69, 95% CI [0.15; 3.05]). Also, vericiguat 10 mg showed no difference compared to placebo regarding the incidence of all-cause mortality (RR: 0.96, 95% CI [0.84; 1.10]), any adverse events (AEs) (RR: 0.94, 95% CI [0.83; 1.07]), any serious AEs (RR: 0.91, 95% CI [0.81; 1.01]), and any AEs leading to drug discontinuation (RR: 1.14, 95% CI [0.92; 1.40]).ConclusionVericiguat 10 mg was effective in reducing the composite CVS mortality and HF hospitalization, with an acceptable safety profile. This was only observed in HFrEF patients, but not in HFpEF patients. However, our data regarding other agents (riociguat and praliciguat) and HFpEF can be underpowered, warranting further RCTs to clarify vericiguat 10 mg place in HFrEF management guidelines and to investigate sGC stimulators for HFpEF in large-scale trials.

SOLUBLE GUANYLATE CYCLASE STIMULATORS IN HEART FAILURE WITH PRESERVED EJECTION FRACTION: A SYSTEMATIC REVIEW AND META-ANALYSIS.

Aims: We conducted a systemic review and meta-analysis to evaluate the therapeutic efficacy and safety of soluble guanylate cyclase (sGC) stimulators in patients with heart failure with preserved ejection fraction (HFpEF). Methods : We systematically searched PubMed, Embase, and Cochrane Library databases for original randomized controlled trials comparing sGC stimulators with placebo in HFpEF patients. A random-effects model was applied to evaluate the mortality, quality of life, and drug-related adverse events. This meta-analysis is registered in PROSPERO under the number CRD42023457382. Results : We included five studies involving 1,600 HFpEF patients. Comprehensively, the combined risk ratio (RR) for mortality was not significant (RR [95% CI] = 1.44 [0.71 to 2.91], P = 0.31). Furthermore, there were no statistically significant differences in the Kansas City Cardiomyopathy Questionnaire results, including the clinical summary score (weighted mean difference [WMD] [95% CI] =0.32 [-7.38 to 8.02], P = 0.94) and the overall summary score (WMD [95% CI] = -0.87 [-8.87 to 7.14], P = 0.83). Similarly, there was no significant improvement in the 6-minute walk distance (WMD [95% CI] = -6.22 [-18.56 to 6.12], P = 0.32). In addition, drug-related adverse events were more common in patients treated with sGC stimulators (RR [95% CI] = 1.63 [1.25-2.14], P < 0.05). Conclusion : Oral sGC stimulators do not significantly improve mortality outcomes, functional capacity, and quality of life in HFpEF patients but are associated with increased drug-related adverse events. Therefore, we should consider using sGC stimulators in HFpEF patients carefully.

Stimulation of soluble guanylyl cyclase in erythrocytes induces export of cGMP and cardioprotection in type 2 diabetes

Abstract Background Red blood cells (RBCs) are known to export cardioprotective nitric oxide (NO) bioactivity. Although, RBCs contain a functional soluble guanylyl cyclase (sGC), the signalling behind the export of NO bioactivity remains unclear. Impairment of NO signalling in RBCs has been suggested to contribute to augmented cardiac ischemia-reperfusion injury in type 2 diabetes (T2D). We hypothesized that increased NO signaling by stimulation of sGC in RBCs protects from myocardial ischemia-reperfusion injury by release of a cardioprotective signal. Purpose To investigate whether stimulation of sGC, the intracellular NO receptor, in RBCs from T2D patients induces export of a cardioprotective factor and to determine its nature. Methods RBCs were collected from patients with T2D and from gender- and age-matched healthy subjects and incubated with different pharmacological compounds affecting the NO-signalling pathway. The RBCs or their supernatant were then administered to isolated Langendorff-perfused rat hearts subjected to 25 min global ischemia and 60 min reperfusion. Left ventricular developed pressure and infarct size were measured. ELISA and immunofluorescence were used to determine levels of cyclic guanosine monophosphate (cGMP) and substrate of protein kinase G activation. All animal experiments and procedures were performed according to the guidelines by the U.S National Institutes of Health (NIH publication no 85-23, revised 1996). The present study was performed following The Code of Ethics of the World Medical Association outlined in the Declaration of Helsinki of 1975 and revised in 1983 for experiments that involve human subjects. Results RBCs from T2D patients impaired post-ischemic cardiac function and increased infarct size compared to RBCs from healthy subjects, and these effects were prevented by pre-incubation of the RBCs with the sGC stimulator CYR715. Administration of the supernatant from RBCs incubated with the sGC stimulator resulted in similar cardioprotection, indicating release of a cardioprotective factor from the RBCs (Figure 1A and B). The cardioprotection was abolished by inhibition of transport of cGMP with MK571, an inhibitor of cyclic nucleotide transport. Extracellular cGMP levels in the supernatant from RBCs were increased following sGC stimulation, and administration of exogenous cGMP induced cardioprotection (Figure 1C and D). Expression of protein kinase G-dependent phosphorylated vasodilator-specific phosphoprotein in cardiomyocytes was increased following administration of supernatant from RBCs incubated with the sGC stimulator (Figure 1E). Conclusions This study shows that stimulation of sGC in RBCs leads to release of a cardioprotective mediator, identified as cGMP, that activates cardiac protein kinase G and protects from ischemia-reperfusion injury. It further identifies RBC sGC as a novel therapeutic target to prevent cardiovascular injury in T2D.Figure 1

Soluble guanylate cyclase in heart failure: A network meta-analysis and subgroup analyses of reduced and preserved ejection fraction

Abstract Background Heart failure (HF), either with reduced ejection fraction (HFrEF) or preserved ejection fraction (HFpEF), is a significant healthcare burden, with about one million hospitalizations annually in the United States alone. Soluble guanylate cyclase (sGC) stimulators have been investigated for HF treatment in several randomized controlled trials (RCTs). However, its place in the management guidelines of either HFrEF or HfpEF is still inconclusive. Purpose We aim to evaluate the efficacy and safety of sGC in HF and determine the most effective agent and dosage for the management of either HFrEF or HFpEF. Methods We conducted a systematic review and network meta-analysis synthesizing RCTs investigating sGC, including vericiguat, riociguat, and praliciguat for HF management, which were retrieved by systematically searching five databases until January 24th, 2023. Dichotomous outcomes were pooled using risk ratio (RR) along with confidence interval (CI). Frequentist random-effects network meta-analysis was conducted via R software using meta and netmeta packages. Results Eight RCTs with a total of 7307 patients were included in our analysis. Four RCTs investigated vericiguat, three investigated riociguat, and another investigated praliciguat. Five RCTs investigated HFpEF patients, and three investigated HFrEF patients. Vericiguat 10 mg significantly reduced the incidence of composite cardiovascular (CVS) mortality and HF hospitalization in HF (RR: 0.88, 95% CI [0.79; 0.98], P = 0.02) and in HFrEF (RR: 0.87, 95% CI [0.78; 0.97], P = 0.02); however, it was not effective in HFpEF (RR: 0.69, 95% CI [0.15; 3.05], P = 0.62). Also, vericiguat 10 mg showed no difference compared to placebo regarding the incidence of all-cause mortality (RR: 0.96, 95% CI [0.84; 1.10], P = 0.57), any adverse events (AEs) (RR: 0.94, 95% CI [0.83; 1.07], P = 0.34), any serious AEs (RR: 0.91, 95% CI [0.81; 1.01], P = 0.08), and any AEs leading to drug discontinuation (RR: 1.14, 95% CI [0.92; 1.40], P = 0.23). However, all other comparisons of vericiguat, riociguat, and praliciguat showed no difference compared to placebo regarding the composite CVS mortality and HF hospitalization, with a similar safety profile. Conclusion Only vericiguat 10 mg was effective in reducing the incidence of the composite CVS mortality and HF hospitalization in patients with HFrEF, with an acceptable safety profile. However, this observation is mainly weighted by the VICTORIA trial, constituting 69.2% of our analyzed sample size, which investigated vericiguat 10 mg in patients with HFrEF. Therefore, our data regarding other agents (riociguat and praliciguat) and HFpEF can be underpowered. This warrants further RCTs to clarify vericiguat 10 mg place in HFrEF management guidelines and to investigate sGC stimulators for HFpEF in large-scale adequately designed trials.Figure 1Figure 2

Abstract 12557: Vericiguat Use in a Real-World HFrEF Patient Population: Two Years Post-Launch

Background: Worsening heart failure events (WHFE) affect up to one-third of patients with HFrEF. Vericiguat, the first agent indicated specifically for a WHF population, is a novel soluble guanylate cyclase (sGC) stimulator approved in 2021 to reduce the risk of cardiovascular death and HF hospitalization. Objective: To describe baseline characteristics, comorbidities, and comedication among patients on vericiguat therapy during the first two years after FDA approval. Methods: We identified adults in the TriNetX Open Claims Database who filled a prescription for vericiguat between 1 Jan 2021 and 24 April 2023. Summary statistics were computed describing patient and treatment patterns including those of other guideline-directed medical therapy (GDMT), stratified by record of achieving 10mg target dose. Categorical variables compared across groups using Pearson Chi-Square tests and continuous variables were compared across groups using non-parametric Kruskal-Wallis tests. Results: Among 5527 patients treated with vericiguat, mean age was 68 years, most were men (67%), and insured by Medicare (47%). Just over one third had a record of achieving the 10mg target dose before data cutoff. Other common GDMT used included beta-blockers (71%), ARNi (47%), MRA (38%) and SGLT2i (39%). Although comorbidities were similar between groups, use of these GDMT classes was statistically significantly higher among patients with a record of achieving target dose of vericiguat (P &lt; 0.05). Similar trends were observed among use of GDMT combinations based on record of achieving target dose (P &lt;0.002). Conclusions: Patients with a record of achieving vericiguat target dose were significantly more likely to be on other recommended GDMT and on GDMT combinations despite a similar comorbidity burden. Additional analyses are needed to better understand factors associated with patients reaching classes and target doses of GDMT, including vericiguat.

Impact of vericiguat on baroreflex-mediated sympathetic circulatory regulation: An open-loop analysis.

To quantify in vivo the effects of the soluble guanylate cyclase (sGC) stimulator, vericiguat, on autonomic cardiovascular regulation in comparison with the nitric oxide (NO) donor, sodium nitroprusside. In anesthetized Wistar-Kyoto rats, baroreflex-mediated changes in sympathetic nerve activity (SNA), arterial pressure (AP), central venous pressure (CVP), and aortic flow (AoF) were examined before and during the intravenous continuous administration (10 μg·kg-1·min-1) of vericiguat or sodium nitroprusside (n = 8 each). Systemic vascular resistance (SVR) was calculated as SVR = (AP-CVP) / AoF. Neither vericiguat nor sodium nitroprusside affected fitted parameters of the baroreflex-mediated SNA response. Both vericiguat and sodium nitroprusside decreased the AP mainly through their peripheral effects. Vericiguat halved the slope of the SNA-SVR relationship from 0.012 ± 0.002 to 0.006 ± 0.002 mmHg·min·mL-1·%-1 (P = 0.008), whereas sodium nitroprusside caused a near parallel downward shift in the SNA-SVR relationship with a reduction of the SVR intercept from 1.235 ± 0.187 to 0.851 ± 0.123 mmHg·min/mL (P = 0.008). Neither vericiguat nor sodium nitroprusside significantly affected the baroreflex-mediated SNA response. The vasodilative effect of vericiguat became greater toward high levels of SNA and AP, possibly reflecting the increased sGC sensitivity to endogenous NO. By contrast, the effect of sodium nitroprusside was more uniform over the range of SNA. These results help better understand cardiovascular effects of vericiguat.

Stimulation of Erythrocyte Soluble Guanylyl Cyclase Induces cGMP Export and Cardioprotection in Type 2 Diabetes

Reduced nitric oxide (NO) bioactivity in red blood cells (RBCs) is critical for augmented myocardial ischemia-reperfusion injury in type 2 diabetes. This study identified the nature of "NO bioactivity" by stimulating the intracellular NO receptor soluble guanylyl cyclase (sGC) in RBCs. sGC stimulation in RBCs from patients with type 2 diabetes increased export of cyclic guanosine monophosphate from RBCs and activated cardiac protein kinase G, thereby attenuating ischemia-reperfusion injury. These results provide novel insight into RBC signaling by identifying cyclic guanosine monophosphate from RBC as a mediator of protection against cardiac ischemia-reperfusion injury induced by sGC stimulation in RBCs.

Targeting Soluble Guanylyl Cyclase during Ischemia and Reperfusion.

Ischemia and reperfusion (IR) damage organs and contribute to many disease states. Few effective treatments exist that attenuate IR injury. The augmentation of nitric oxide (NO) signaling remains a promising therapeutic target for IR injury. NO binds to soluble guanylyl cyclase (sGC) to regulate vasodilation, maintain endothelial barrier integrity, and modulate inflammation through the production of cyclic-GMP in vascular smooth muscle. Pharmacologic sGC stimulators and activators have recently been developed. In preclinical studies, sGC stimulators, which augment the reduced form of sGC, and activators, which activate the oxidized non-NO binding form of sGC, increase vasodilation and decrease cardiac, cerebral, renal, pulmonary, and hepatic injury following IR. These effects may be a result of the improved regulation of perfusion and decreased oxidative injury during IR. sGC stimulators are now used clinically to treat some chronic conditions such as heart failure and pulmonary hypertension. Clinical trials of sGC activators have been terminated secondary to adverse side effects including hypotension. Additional clinical studies to investigate the effects of sGC stimulation and activation during acute conditions, such as IR, are warranted.

Ameliorating diabetes-associated atherosclerosis and diabetic nephropathy through modulation of soluble guanylate cyclase.

Diabetes mellitus (DM) is an independent risk factor for micro- and macrovascular complications such as nephropathy and atherosclerosis respectively, which are the major causes of premature morbidity and mortality in Type 1 and Type 2 diabetic patients. Endothelial dysfunction is the critical first step of vascular disease and is characterized by reduced bioavailability of the essential endothelial vasodilator, nitric oxide (NO), coupled with an elevation in inflammation and oxidative stress. A novel pathway to bolster NO activity is to upregulate soluble guanylate cyclase (sGC), an enzyme responsible for mediating the protective actions of NO. Two classes of sGC modulators exist, activators and stimulators, with differing sensitivity to oxidative stress. In this study, we investigated the therapeutic effects of the sGC stimulator BAY 41-2272 (Bay 41) and the sGC activator BAY 60-2770 (Bay 60) on endpoints of atherosclerosis and renal disease as well as inflammation and oxidative stress in diabetic Apolipoprotein E knockout (ApoE-/-) mice. We hypothesized that under oxidative conditions known to accompany diabetes, sGC activation might be more efficacious than sGC stimulation in limiting diabetic vascular complications. We demonstrate that Bay 60 not only significantly decreased nitrotyrosine staining (P < 0.01) and F4/80 positive cells by 75% (P < 0.05), but it also significantly reduced total plaque area (P < 0.05) and improved endothelial function (P < 0.01). Our data suggest an important anti-atherogenic role for Bay 60 accompanied by reduced oxidative stress and inflammation under diabetic settings. Treatment with the stimulator Bay 41, on the other hand, had minimal effects or caused no changes with respect to cardiovascular or renal pathology. In the kidneys, treatment with Bay 60 significantly lessened urinary albuminuria, mesangial expansion and nitrotyrosine staining under diabetic conditions. In summary, our head-to-head comparator is the first preclinical study to show that a sGC activator is more efficacious than a sGC stimulator for the treatment of diabetes-associated vascular and renal complications.

Vericiguat preserved cardiac function and mitochondrial quality in a rat model of mitral regurgitation

AimsNew drugs for heart failure (HF) that target restoring the impaired NO-sGC-cGMP pathway are being developed. We aimed to investigate the effects of vericiguat, an sGC stimulator, on cardiac function, blood pressure (BP), cardiac mitochondrial quality, and cardiac fibrosis in rat models of chronic mitral regurgitation (MR). Materials and methodsWe surgically induced MR in 20 Sprague-Dawley rats and performed sham procedures on 10 rats (negative control). Four weeks post-surgery, we randomly divided the MR rats into two groups: MR group and MR + vericiguat group. Vericiguat (0.5 mg/kg, PO) was administered once a day via oral gavage for 8 weeks, while the sham and MR groups received equivalent volumes of drinking water instead. We took echocardiography and BP measurements at baseline (4 weeks post-surgery) and at the end of study (8 weeks after treatment). At the study end, all rats were euthanized and their hearts were immediately collected, weighed, and used for histopathology and mitochondrial quality assessments. Key findingsVericiguat preserved cardiac functions and structural remodeling in the MR rats, with significantly lower systolic BPs than baseline values (P < 0.05). Additionally, vericiguat significantly improved the mitochondrial quality by attenuating ROS production, depolarization and swelling when comparing the values in both groups (P < 0.05). The fibrosis area also significantly decreased in the MR + vericiguat group (P < 0.05). SignificanceVericiguat demonstrated cardioprotective effects on cardiac function, BP, and fibrosis by preserving mitochondrial quality in rats with HF due to MR.

Identification and characterization of the new generation soluble guanylate cyclase stimulator BAY-747 designed for the treatment of resistant hypertension.

First-generation soluble guanylate cyclase (sGC) stimulators have shown clinical benefit in pulmonary hypertension (riociguat) and chronic heart failure (vericiguat). However, given the broad therapeutic opportunities for sGC stimulators, tailored molecules for distinct indications are required. We report the high-throughput screening (HTS)-based discovery of a second generation of sGC stimulators from a novel imidazo[1,2-a]pyridine lead series. An intense medicinal chemistry programme resulted in the discovery of the sGC stimulator BAY 1165747 (BAY-747). The pharmacokinetic profile of BAY-747 was determined in different species, and it was broadly characterized in pharmacological model systems relevant for vasodilatation and hypertension. BAY-747 is a highly potent sGC stimulator in vitro. In addition, BAY-747 showed an excellent pharmacokinetic profile with long half-life and low peak-to-trough ratio. BAY-747 was investigated in experimental in vivo models of malignant and resistant hypertension (rHT). In spontaneously hypertensive (SH) rats, BAY-747 caused a dose-related and long-lasting decrease in mean arterial blood pressure (MAP). Oral treatment over 12 days resulted in a persistent decrease. BAY-747 provided additional benefit when dosed on top of losartan, amlodipine or spironolactone and even on top of triple combinations of frequently used antihypertensive drugs. In a new canine model of rHT, BAY-747 caused a dose-related and long-lasting (>6 h) MAP decrease. BAY-747 is a potent, orally available sGC stimulator. BAY-747 shows long-acting pharmacodynamic effects with a very low peak-to-trough ratio. BAY-747 could be a treatment alternative for patients with hypertension, especially those not responding to standard-of-care therapy.

The treatment with sGC stimulator improves survival of hypertensive rats in response to volume-overload induced by aorto-caval fistula

Heart failure (HF) has been declared as global pandemic and current therapies are still ineffective, especially in patients that develop concurrent cardio-renal syndrome. Considerable attention has been focused on the nitric oxide (NO)/soluble guanylyl cyclase (sGC)/cyclic guanosine monophosphate (cGMP) pathway. In the current study, we aimed to investigate the effectiveness of sGC stimulator (BAY41-8543) with the same mode of action as vericiguat, for the treatment of heart failure (HF) with cardio-renal syndrome. As a model, we chose heterozygous Ren-2 transgenic rats (TGR), with high-output heart failure, induced by aorto-caval fistula (ACF). The rats were subjected into three experimental protocols to evaluate short-term effects of the treatment, impact on blood pressure, and finally the long-term survival lasting 210 days. As control groups, we used hypertensive sham TGR and normotensive sham HanSD rats. We have shown that the sGC stimulator effectively increased the survival of rats with HF in comparison to untreated animals. After 60 days of sGC stimulator treatment, the survival was still 50% compared to 8% in the untreated rats. One-week treatment with sGC stimulator increased the excretion of cGMP in ACF TGR (109 ± 28 nnmol/12 h), but the ACE inhibitor decreased it (-63 ± 21 nnmol/12 h). Moreover, sGC stimulator caused a decrease in SBP, but this effect was only temporary (day 0: 117 ± 3; day 2: 108 ± 1; day 14: 124 ± 2 mmHg). These results support the concept that sGC stimulators might represent a valuable class of drugs to battle heart failure especially with cardio-renal syndrome, but further studies are necessary.

Drug repurposing-a promising approach for patients with angina but non-obstructive coronary artery disease (ANOCA).

In today's era of individualized precision medicine drug repurposing represents a promising approach to offer patients fast access to novel treatments. Apart from drug repurposing in cancer treatments, cardiovascular pharmacology is another attractive field for this approach. Patients with angina pectoris without obstructive coronary artery disease (ANOCA) report refractory angina despite standard medications in up to 40% of cases. Drug repurposing also appears to be an auspicious option for this indication. From a pathophysiological point of view ANOCA patients frequently suffer from vasomotor disorders such as coronary spasm and/or impaired microvascular vasodilatation. Consequently, we carefully screened the literature and identified two potential therapeutic targets: the blockade of the endothelin-1 (ET-1) receptor and the stimulation of soluble guanylate cyclase (sGC). Genetically increased endothelin expression results in elevated levels of ET-1, justifying ET-1 receptor blockers as drug candidates to treat coronary spasm. sGC stimulators may be beneficial as they stimulate the NO-sGC-cGMP pathway leading to GMP-mediated vasodilatation.

RIOCIGUAT-INDUCED DOWNREGULATION OF GENES RESPONSIBLE FOR SODIUM TRANSPORT THROUGH THE ALVEOLAR EPITHELIUM IN THE MONOCROTALINE MODEL OF PULMONARY HYPERTENSION

Objective: Pleural effusions (PLEF) in patients with pulmonary arterial hypertension (PAH) co-diagnosed with isolated right-sided heart failure (RHF) have been reported to be associated with increased mortality. We hypothesized that alterations of transporters responsible for alveolar fluid clearance contributes to fluid accumulation in lungs and secondary formation of PLEF. This was examined in a model of PAH including administration of riociguat (RIO) that as a stimulator of sGC may be responsible for cGMP-dependent inhibition of epithelial sodium channel (ENaC) and might affect alveolar epithelial sodium transport. Design and method: Wistar rats were injected with monocrotaline (MCT; 60 mg/kg, s.c.) or vehicle (CON). 2 weeks after MCT administration, RIO treatment (10 mg/kg/d, p.o.) was started (CON+RIO, MCT+RIO). MCT groups were subdivided: MCT4W - sacrificed 4 weeks after MCT application and prematurely terminated MCT (ptMCT) - sacrificed after PAH deterioration. Expressions of ENaC alpha, beta, gama subunits, Na-K-Cl cotransporter (NKCC1), sodium-glucose linked transporter (SGLT1), sodium-coupled neutral amino acid transporters (SNAT2, SNAT5), aquaporin 3 (AQP3) were determined by RT-qPCR in lungs. Lung samples were subjected to histological analysis. Results: Increased lung weight was observed in MCT4W (2.331g vs.CON, p&lt;0.05) and ptMCT (2.778g, vs.MCT4W, p&lt;0.05). Similar situation was observed in RIO treated groups, however, more profound compared to MCT4W (ptMCT+RIO = 3.086g, p&lt;0.05). Expression of ENaC, SNAT5 was decreased in ptMCT and ptMCT+RIO. Expression of NKCC1 decreased in ptMCT+RIO (by 30%) and simultaneously expression of SNAT2 in MCT4W+RIO, ptMCT+RIO (by 58%), both p&lt;0.05 vs.CON, without any changes in MCT4W and ptMCT. 15-fold increase in AQP3 expression was observed in ptMCT, ptMCT+RIO (p&lt;0.05 vs.CON). The presence of fluid in alveoli was observed in MCT4W and ptMCT with incidence of PLEF 20% and 30% respectively (ns). Incidence of PLEF in MCT4W+RIO and ptMCT+RIO was 22% and 44% (p&lt;0.05), however, with interstitial swelling present. Conclusions: Downregulation of ENaC in MCT-induced PAH may represent a predisposing factor for alveolar fluid clearance impairment contributing to accumulation of fluid in lungs manifested as lung weight increase. Incidence of PLEF is probably related to lung weight increase. Riociguat rather worsened the incidence of PLEF and downregulation of ENaC.

Effects of Soluble Guanylate Cyclase Stimulators and Activators on Anti-Aggregatory Signalling in Patients with Coronary Artery Spasm.

Impairment of the nitric oxide/soluble guanylate cyclase (NO)/sGC) signalling cascade is associated with many forms of cardiovascular disease, resulting not only in compromised vasodilatation but also loss of anti-aggregatory homeostasis. Myocardial ischaemia, heart failure, and atrial fibrillation are associated with moderate impairment of NO/sGC signalling, and we have recently demonstrated that coronary artery spasm (CAS) is engendered by severe impairment of platelet NO/sGC activity resulting in combined platelet and vascular endothelial damage. We therefore sought to determine whether sGC stimulators or activators might normalise NO/sGC homeostasis in platelets. ADP-induced platelet aggregation and its inhibition by the NO donor sodium nitroprusside (SNP), the sGC stimulator riociguat (RIO), and the sCG activator cinaciguat (CINA) alone or in addition to SNP were quantitated. Three groups of individuals were compared: normal subjects (n = 9), patients (Group 1) with myocardial ischaemia, heart failure and/or atrial fibrillation (n = 30), and patients (Group 2) in the chronic stage of CAS (n = 16). As expected, responses to SNP were impaired (p = 0.02) in patients versus normal subjects, with Group 2 patients most severely affected (p = 0.005). RIO alone exerted no anti-aggregatory effects but potentiated responses to SNP to a similar extent irrespective of baseline SNP response. CINA exerted only intrinsic anti-aggregatory effects, but the extent of these varied directly (r = 0.54; p = 0.0009) with individual responses to SNP. Thus, both RIO and CINA tend to normalise anti-aggregatory function in patients in whom NO/sGC signalling is impaired. The anti-aggregatory effects of RIO consist entirely of potentiation of NO, which is not selective of platelet NO resistance. However, the intrinsic anti-aggregatory effects of CINA are most marked in individuals with initially normal NO/sGC signalling, and thus their magnitude is at variance with extent of physiological impairment. These data suggest that RIO and other sGC stimulators should be evaluated for clinical utility in both prophylaxis and treatment of CAS.

INOCA/ANOCA: Mechanisms and novel treatments

Angina or ischemia with no obstructive coronary disease (ANOCA/INOCA) is a common but under-treated condition due to poorly understood pathophysiologic mechanisms, limited diagnostic tools, and lack of proven targeted therapy. Coronary microvascular dysfunction (CMD) occurs when the microvasculature inadequately perfuses the myocardium under stress, or at rest in the case of microvascular spasm resulting in ANOCA/INOCA. Coronary functional angiography (CFA) measures endothelial independent microvascular dysfunction (coronary flow reduction <2.5) in response to adenosine and endothelial dependent microvascular dysfunction (lack of dilation and/or constriction) to acetylcholine testing as well as epicardial and microvascular spasm. Current treatment for coronary microvascular dysfunction is limited to renin-angiotensin system (RAS) inhibitors and statins as well as antianginal medications. Novel therapies targeting the underlying pathology are under development and include the coronary sinus reducer, CD34+ stem cell therapy, and novel pharmacologic agents such as sGC stimulators or endothelin-receptor blockers. We review the current understanding of pathophysiology, diagnostic tools, and novel therapies for coronary microvascular dysfunction in ANOCA/INOCA.

Oxidized soluble guanylyl cyclase causes erectile dysfunction in alcoholic mice.

Alcohol abuse has been associated with erectile dysfunction (ED) but the molecular mechanisms implicated are unknown. This study analyses the role of alterations in soluble guanylyl cyclase (sGC) in ED. ED was analysed in adult male C57BL/6J mice subjected to the Chronic Intermittent Ethanol (CIE) paradigm. Erectile function was assessed in anesthetized mice in vivo by evaluating intracavernosal pressure (ICP) and in vitro in isolated mice corpora cavernosa (CC) mounted in a myograph. Protein expression and reactive oxygen species were analysed by Western blot and dihydroethidium staining, respectively. In CIE mice, we observed a significant decrease in the relaxant response of the CC to stimulation of NO release from nitrergic nerves by electrical field stimulation, to NO release from endothelial cells by acetylcholine, to the PDE5 inhibitor sildenafil, and to the sGC stimulator riociguat. Conversely, the response to the sGC activator cinaciguat, whose action is independent on the oxidation state of sGC, was significantly enhanced in these CC. The responses to the adenylyl cyclase stimulation with forskolin were unchanged. We found an increase in reactive oxygen species in the CC from CIE mice as well as an increase in CYP2E1 and NOX2 protein expression. In vivo pre-treatment with tempol prevented alcohol-induced erectile dysfunction. Our results demonstrate that alcoholic mice show ED in vitro and in vivo due to an alteration of the redox state of sGC and suggest that sGC activators may be effective in ED associated with alcoholism.

First-in-human trial to assess safety, tolerability, pharmacokinetics, and pharmacodynamics of zagociguat (CY6463), a CNS-penetrant soluble guanylyl cyclase stimulator.

Soluble guanylate cyclase (sGC) and its product, cyclic guanosine monophosphate, play a role in learning and memory formation. Zagociguat (CY6463) is a novel stimulator of sGC being developed for the treatment of neurodegenerative disease. Single zagociguat doses of 0.3, 1, 3, 10, 20, 30, and 50 mg were administered once to healthy participants in a single-ascending-dose phase; then zagociguat 2, 5, 10, and 15 mg was administered q.d. for 14 days in a multiple-ascending-dose phase; and, finally, zagociguat 10 mg was administered once in both fed and fasted state in a food-interaction phase. Safety of zagociguat was evaluated by monitoring treatment-emergent adverse events, suicide risk, vital signs, electrocardiography, and laboratory tests. Pharmacokinetics of zagociguat were assessed through blood, urine, and cerebrospinal fluid sampling. Pharmacodynamic effects of zagociguat were evaluated with central nervous system (CNS) tests and pharmaco-electroencephalography. Zagociguat was well-tolerated across all doses evaluated. Zagociguat exposures increased in a dose-proportional manner. Median time to maximum concentration ranged from 0.8 to 5 h and mean terminal half-life from 52.8 to 67.1 h. CNS penetration of the compound was confirmed by cerebrospinal fluid sampling. Zagociguat induced up to 6.1 mmHg reduction in mean systolic and up to 7.5 mmHg reduction in mean diastolic blood pressure. No consistent pharmacodynamic (PD) effects on neurocognitive function were observed. Zagociguat was well-tolerated, CNS-penetrant, and demonstrated PD activity consistent with other sGC stimulators. The results of this study support further development of zagociguat.

Effect of Different Extraction Methods on Anthocyanin Content in Hibiscus sabdariffa L. and their Antiplatelet and Vasorelaxant Properties.

Hibiscus sabdariffa L. is a worldwide component for tea and beverages, being a natural source of anthocyanins, which are associated with cardiovascular activities. To investigate this relationship, we explored different methods of aqueous extraction on the anthocyanin content and antioxidant activity of H. sabdariffa L. calyx extract (HSCE). Pharmacological effects via platelet aggregation, calcium mobilization, cyclic nucleotide levels, vasodilator-stimulated phosphoprotein Ser157 and Ser239, and on the vasomotor response of aortic rings isolated from mice are studied herewith. We found that the application of ultrasonic turbolization, 20min, combined with acidified water was significantly more effective in the extraction process, providing extracts with the highest levels of anthocyanins (8.73 and 9.63mg/100g) and higher antioxidant activity (6.66 and 6.78μM trolox/g of sample). HSCE significantly inhibited (100-1000μg/mL) arachidonic acid-induced platelet aggregation, reduced calcium mobilization, and increased cAMP and cGMP levels with VASPSer157 and VASPSer239 phosphorylation. Vasorelaxation reduction was confirmed by the aortic rings and endothelium assays treated with nitric oxide synthase inhibitors, soluble guanylyl cyclase (sGC) oxidizing agent, or Ca2+-activated K+ channel inhibitor. The increasing of cGMP levels could be understood considering the sGC stimulation by HSCE compounds in the specific stimulus domain, which allows an understanding of the observed antiplatelet and vasorelaxant properties of H. sabdariffa L. calyx extract.