The CNS-penetrant soluble guanylate cyclase stimulator CYR119 attenuates markers of inflammation in the central nervous system.

BACKGROUND: Inflammation in the central nervous system (CNS) is observed in many neurological disorders. Nitric oxide-soluble guanylate cyclase-cyclic guanosine monophosphate (NO-sGC-cGMP) signaling plays an essential role in modulating neuroinflammation. CYR119 is a CNS-penetrant sGC stimulator that amplifies endogenous NO-sGC-cGMP signaling. We evaluated target engagement and the effects of CYR119 on markers of neuroinflammation in vitro in mouse microglial cells and in vivo in quinolinic acid (QA)-induced and high-fat diet-induced rodent neuroinflammation models. METHODS: Target engagement was verified in human embryonic kidney (HEK) cells, rat primary neurons, mouse SIM-A9 cells, and in rats by measuring changes in cGMP and downstream targets of sGC signaling [phosphorylated vasodilator-stimulated phosphoprotein (pVASP), phosphorylated cAMP-response element binding (pCREB)]. In SIM-A9 cells stimulated with lipopolysaccharides (LPS), markers of inflammation were measured when cells were treated with or without CYR119. In rats, microinjections of QA and vehicle were administered into the right and left hemispheres of striatum, respectively, and then rats were dosed daily with either CYR119 (10 mg/kg) or vehicle for 7 days. The activation of microglia [ionized calcium binding adaptor molecule 1 (Iba1)] and astrocytes [glial fibrillary acidic protein (GFAP)] was measured by immunohistochemistry. Diet-induced obese (DIO) mice were treated daily with CYR119 (10 mg/kg) for 6 weeks, after which inflammatory genetic markers were analyzed in the prefrontal cortex. RESULTS: In vitro, CYR119 synergized with exogenous NO to increase the production of cGMP in HEK cells and in primary rat neuronal cell cultures. In primary neurons, CYR119 stimulated sGC, resulting in accumulation of cGMP and phosphorylation of CREB, likely through the activation of protein kinase G (PKG). CYR119 attenuated LPS-induced elevation of interleukin 6 (IL-6) and tumor necrosis factor (TNF) in mouse microglial cells. Following oral dosing in rats, CYR119 crossed the blood-brain barrier (BBB) and stimulated an increase in cGMP levels in the cerebral spinal fluid (CSF). In addition, levels of proinflammatory markers associated with QA administration or high-fat diet feeding were lower in rodents treated with CYR119 than in those treated with vehicle. CONCLUSIONS: These data suggest that sGC stimulation could provide neuroprotective effects by attenuating inflammatory responses in nonclinical models of neuroinflammation.

The CNS-Penetrant Soluble Guanylate Cyclase Stimulator CY6463 Reveals its Therapeutic Potential in Neurodegenerative Diseases.

Effective treatments for neurodegenerative diseases remain elusive and are critically needed since the burden of these diseases increases across an aging global population. Nitric oxide (NO) is a gasotransmitter that binds to soluble guanylate cyclase (sGC) to produce cyclic guanosine monophosphate (cGMP). Impairment of this pathway has been demonstrated in neurodegenerative diseases. Normalizing deficient NO-cGMP signaling could address multiple pathophysiological features of neurodegenerative diseases. sGC stimulators are small molecules that synergize with NO, activate sGC, and increase cGMP production. Many systemic sGC stimulators have been characterized and advanced into clinical development for a variety of non-central nervous system (CNS) pathologies. Here, we disclose the discovery of CY6463, the first brain-penetrant sGC stimulator in clinical development for the treatment of neurodegenerative diseases, and demonstrate its ability to improve neuronal activity, mediate neuroprotection, and increase cognitive performance in preclinical models. In several cellular assays, CY6463 was demonstrated to be a potent stimulator of sGC. In agreement with the known effects of sGC stimulation in the vasculature, CY6463 elicits decreases in blood pressure in both rats and mice. Relative to a non-CNS penetrant sGC stimulator, rodents treated with CY6463 had higher cGMP levels in cerebrospinal fluid (CSF), functional-magnetic-resonance-imaging-blood-oxygen-level-dependent (fMRI-BOLD) signals, and cortical electroencephalographic (EEG) gamma-band oscillatory power. Additionally, CY6463 improved cognitive performance in a model of cognitive disruption induced by the administration of a noncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist. In models of neurodegeneration, CY6463 treatment increased long-term potentiation (LTP) in hippocampal slices from a Huntington's disease mouse model and decreased the loss of dendritic spines in aged and Alzheimer's disease mouse models. In a model of diet-induced obesity, CY6463 reduced markers of inflammation in the plasma. Furthermore, CY6463 elicited an additive increase in cortical gamma-band oscillatory power when co-administered with donepezil: the standard of care in Alzheimer's disease. Together, these data support the clinical development of CY6463 as a novel treatment for neurodegenerative disorders.

Discovery of CYR715: A novel carboxylic acid-containing soluble guanylate cyclase stimulator.

Soluble guanylate cyclase (sGC) is a clinically validated therapeutic target in the treatment of pulmonary hypertension. Modulators of sGC have the potential to treat diseases that are affected by dysregulation of the NO-sGC-cGMP signal transduction pathway. This letter describes the SAR efforts that led to the discovery of CYR715, a novel carboxylic acid-containing sGC stimulator, with an improved metabolic profile relative to our previously described stimulator, IWP-051. CYR715 addressed potential idiosyncratic drug toxicity (IDT) liabilities associated with the formation of reactive, migrating acyl glucuronides (AG) found in related carboxylic acid-containing analogs and demonstrated high oral bioavailability in rat and dose-dependent hemodynamic pharmacology in normotensive Sprague-Dawley rats.

Pharmacokinetics, mass balance, tissue distribution, metabolism, and excretion of praliciguat, a clinical-stage soluble guanylate cyclase stimulator in rats.

The pharmacokinetics (PK), metabolism, excretion, mass balance, and tissue distribution of [14 C]praliciguat were evaluated following oral administration of a 3-mg/kg dose in Sprague-Dawley rats and in a quantitative whole-body autoradiography (QWBA) study conducted in male Long-Evans rats. Plasma Tmax was 1 hour and the t1/2 of total plasma radioactivity was 23.7 hours. Unchanged praliciguat accounted for 87.4%, and a minor metabolite (N-dealkylated-praliciguat) accounted for 7.6% of the total radioactivity in plasma through 48 hours (AUC0-48 ). Tissues with the highest exposure ratios relative to plasma were liver, intestines, adrenal gland, and adipose, and those with the lowest values were seminal vesicle, blood, CNS tissues, lens of the eye, and bone. Most of the [14 C]praliciguat-derived radioactivity was excreted within 48 hours after oral administration. Mean cumulative recovery of the administered radioactivity in urine and feces over 168 hours was 3.7% and 95.7%, respectively. Unchanged praliciguat was not quantifiable in urine or bile of cannulated rats; however, based on the total radioactivity in these fluids, a minimum of approximately 82% of the orally administered dose was absorbed. [14 C]Praliciguat was metabolized via oxidative and glucuronidation pathways and the most abundant metabolites recovered in bile were praliciguat-glucuronide and hydroxy-praliciguat-glucuronide. These results indicate that praliciguat had rapid absorption, high bioavailability, extensive tissue distribution, and elimination primarily via hepatic metabolism.

Olinciguat, an Oral sGC Stimulator, Exhibits Diverse Pharmacology Across Preclinical Models of Cardiovascular, Metabolic, Renal, and Inflammatory Disease.

Nitric oxide (NO)-soluble guanylate cyclase (sGC)-cyclic 3',5' GMP (cGMP) signaling plays a central role in regulation of diverse processes including smooth muscle relaxation, inflammation, and fibrosis. sGC is activated by the short-lived physiologic mediator NO. sGC stimulators are small-molecule compounds that directly bind to sGC to enhance NO-mediated cGMP signaling. Olinciguat, (R)-3,3,3-trifluoro-2-(((5-fluoro-2-(1-(2-fluorobenzyl)-5-(isoxazol-3-yl)-1H-pyrazol-3-yl)pyrimidin-4-yl)amino)methyl)-2-hydroxypropanamide, is a new sGC stimulator currently in Phase 2 clinical development. To understand the potential clinical utility of olinciguat, we studied its pharmacokinetics, tissue distribution, and pharmacologic effects in preclinical models. Olinciguat relaxed human vascular smooth muscle and was a potent inhibitor of vascular smooth muscle proliferation in vitro. These antiproliferative effects were potentiated by the phosphodiesterase 5 inhibitor tadalafil, which did not inhibit vascular smooth muscle proliferation on its own. Olinciguat was orally bioavailable and predominantly cleared by the liver in rats. In a rat whole body autoradiography study, olinciguat-derived radioactivity in most tissues was comparable to plasma levels, indicating a balanced distribution between vascular and extravascular compartments. Olinciguat was explored in rodent models to study its effects on the vasculature, the heart, the kidneys, metabolism, and inflammation. Olinciguat reduced blood pressure in normotensive and hypertensive rats. Olinciguat was cardioprotective in the Dahl rat salt-sensitive hypertensive heart failure model. In the rat ZSF1 model of diabetic nephropathy and metabolic syndrome, olinciguat was renoprotective and associated with lower circulating glucose, cholesterol, and triglycerides. In a mouse TNFα-induced inflammation model, olinciguat treatment was associated with lower levels of endothelial and leukocyte-derived soluble adhesion molecules. The pharmacological features of olinciguat suggest that it may have broad therapeutic potential and that it may be suited for diseases that have both vascular and extravascular pathologies.

Discovery of IWP-051, a Novel Orally Bioavailable sGC Stimulator with Once-Daily Dosing Potential in Humans.

In recent years, soluble guanylate cyclase (sGC, EC 4.6.1.2) has emerged as an attractive therapeutic target for treating cardiovascular diseases and diseases associated with fibrosis and end-organ failure. Herein, we describe our design and synthesis of a series of 4-hydroxypyrimidine sGC stimulators starting with an internally discovered lead. Our efforts have led to the discovery of IWP-051, a molecule that achieves good alignment of potency, stability, selectivity, and pharmacodynamic effects while maintaining favorable pharmacokinetic properties with once-daily dosing potential in humans.

The relationship between pregnancy and oxidative stress markers on patients undergoing ovarian stimulations.

PURPOSE: We investigated the activities and relevance of a validated panel of antioxidant enzymes, cytokines, specific lipid peroxidation end products and six fatty acids by correlational analyses with peak E(2) levels and pregnancy outcome after ovarian stimulation for IVF or IUI. METHODS: Blood samples obtained from 15 patients undergoing ovarian stimulation with rFSH or hMG were divided into two groups. Group-1 was baseline blood collected on day-2-3 of women cycle. Group-2 is blood collected at the end of FSH/hMG injection. Serum was collected and stored in liquid nitrogen at -196 °C until analysis. Standard IVF and IUI procedures were followed. The serum levels of Paraoxonase (PON1), Superoxide Dismutases (SOD), Interleukin-6 (IL-6), Glutathione Peroxidase (GPx), 8-Isoprostane, and fatty acids Arachidic, Palmitic, Stearic, Oleic, Linoleic & Linolenic were measured. RESULTS: With the exception of 8-Isoprostane, results showed a positive correlation between baseline and peak levels of E(2) and that of SOD, GPx, PON1, and IL-6. The PON1, IL-6 and SOD were significantly (p < 0.05) higher in pregnant than non-pregnant group. Fatty acid levels at baseline and peak E(2) were not different but pregnancy rates were found to be decreasing with higher palmitic, and stearic acid levels. CONCLUSIONS: Ovarian stimulation causes a significant increase in serum PON1, SOD, GPx and IL-6 activity in women undergoing IVF or IUI. The high levels of IL-6, SOD, and PON1 and lower levels of palmitic, and stearic acids in the pregnancy positive group indicate that these oxidative stress and nutritional factors may be used as a predictive marker in controlled ovarian stimulation success.