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Some bacteria survive in nutrient-poor environments and resist killing by antimicrobials by forming spores. The cortex layer of the peptidoglycan cell wall that surrounds mature spores contains a unique modification, muramic-δ-lactam, that is essential for spore germination and outgrowth. Two proteins, the amidase CwlD and the deacetylase PdaA, are required for muramic-δ-lactam synthesis in cells, but their combined ability to generate muramic-δ-lactam has not been directly demonstrated. Here we report an in vitro reconstitution of cortex peptidoglycan biosynthesis, and we show that CwlD and PdaA together are sufficient for muramic-δ-lactam formation. Our method enables characterization of the individual reaction steps, and we show for the first time that PdaA has transamidase activity, catalyzing both the deacetylation ofN-acetylmuramic acid and cyclization of the product to form muramic-δ-lactam. This activity is unique among peptidoglycan deacetylases and is notable because it may involve the direct ligation of a carboxylic acid with a primary amine. Our reconstitution products are nearly identical to the cortex peptidoglycan found in spores, and we expect that they will be useful substrates for future studies of enzymes that act on the spore cortex.

Background and objectives Impaired nitric oxide signaling through soluble guanylate cyclase has been implicated in the pathophysiology of diabetic kidney disease. Praliciguat, a soluble guanylate cyclase stimulator that amplifies nitric oxide signaling, inhibited kidney inflammation and fibrosis in animal models. Design, setting, participants, & measurements In a phase 2 trial, 156 adults with type 2 diabetes, eGFR 30–75 ml/min per 1.73 m2, and urine albumin-creatinine ratio 200–5000 mg/g treated with renin-angiotensin system inhibitors were randomly allocated 1:1:1 to placebo, 20 mg praliciguat, or 40 mg praliciguat daily for 12 weeks. The primary efficacy and safety outcomes were change from baseline to weeks 8 and 12 in urine albumin-creatinine ratio and treatment-emergent adverse events, respectively. Other outcomes assessed were 24-hour ambulatory BP and metabolic parameters. Results Of 156 participants randomized, 140 (90%) completed the study. The primary efficacy analysis demonstrated a mean change from baseline in urine albumin-creatinine ratio of −28% (90% confidence interval, −36 to −18) in the pooled praliciguat group and −15% (−28 to 0.4) in the placebo group (difference −15%; −31 to 4; P=0.17). Between-group decreases from baseline to week 12 for praliciguat versus placebo were seen in mean 24-hour systolic BP (−4 mm Hg; −8 to −1), hemoglobin A1c (−0.3%; −0.5 to −0.03), and serum cholesterol (−10 mg/dl; −19 to −1). The incidence of treatment-emergent adverse events was similar in the pooled praliciguat and placebo groups (42% and 44%, respectively). Serious adverse events, events leading to study drug discontinuation, and events potentially related to BP lowering were reported at higher frequency in the 40-mg group but were similar in 20-mg and placebo groups. Conclusions Praliciguat treatment for 12 weeks did not significantly reduce albuminuria compared with placebo in the primary efficacy analysis. Nonetheless, the observed changes in urine albumin-creatinine ratio, BP, and metabolic variables may support further investigation of praliciguat in diabetic kidney disease. Clinical Trial registry name and registration number: A Study to Evaluate the Soluble Guanylate Cyclase (sGC) Stimulator IW-1973 in Diabetic Nephropathy/Diabetic Kidney Disease as Measured by Albuminuria, NCT03217591

IMPORTANCE: Heart failure with preserved ejection fraction (HFpEF) is often characterized by nitric oxide deficiency. OBJECTIVE: To evaluate the efficacy and adverse effects of praliciguat, an oral soluble guanylate cyclase stimulator, in patients with HFpEF. DESIGN, SETTING, AND PARTICIPANTS: CAPACITY HFpEF was a randomized, double-blind, placebo-controlled, phase 2 trial. Fifty-nine sites enrolled 196 patients with heart failure and an ejection fraction of at least 40%, impaired peak rate of oxygen consumption (peak V̇o(2)), and at least 2 conditions associated with nitric oxide deficiency (diabetes, hypertension, obesity, or advanced age). The trial randomized patients to 1 of 3 praliciguat dose groups or a placebo group, but was refocused early to a comparison of the 40-mg praliciguat dose vs placebo. Participants were enrolled from November 15, 2017, to April 30, 2019, with final follow-up on August 19, 2019. INTERVENTIONS: Patients were randomized to receive 12 weeks of treatment with 40 mg of praliciguat daily (n = 91) or placebo (n = 90). MAIN OUTCOMES AND MEASURES: The primary efficacy end point was the change from baseline in peak V̇o(2) in patients who completed at least 8 weeks of assigned dosing. Secondary end points included the change from baseline in 6-minute walk test distance and in ventilatory efficiency (ventilation/carbon dioxide production slope). The primary adverse event end point was the incidence of treatment-emergent adverse events (TEAEs). RESULTS: Among 181 patients (mean [SD] age, 70 [9] years; 75 [41%] women), 155 (86%) completed the trial. In the placebo (n = 78) and praliciguat (n = 65) groups, changes in peak V̇o(2) were 0.04 mL/kg/min (95% CI, –0.49 to 0.56) and −0.26 mL/kg/min (95% CI, −0.83 to 0.31), respectively; the placebo-adjusted least-squares between-group difference in mean change from baseline was −0.30 mL/kg/min ([95% CI, −0.95 to 0.35]; P = .37). None of the 3 prespecified secondary end points were statistically significant. In the placebo and praliciguat groups, changes in 6-minute walk test distance were 58.1 m (95% CI, 26.1-90.1) and 41.4 m (95% CI, 8.2-74.5), respectively; the placebo-adjusted least-squares between-group difference in mean change from baseline was –16.7 m (95% CI, −47.4 to 13.9). In the placebo and praliciguat groups, the placebo-adjusted least-squares between-group difference in mean change in ventilation/carbon dioxide production slope was −0.3 (95% CI, −1.6 to 1.0). There were more dizziness (9.9% vs 1.1%), hypotension (8.8% vs 0%), and headache (11% vs 6.7%) TEAEs with praliciguat compared with placebo. The frequency of serious TEAEs was similar between the groups (10% in the praliciguat group and 11% in the placebo group). CONCLUSIONS AND RELEVANCE: Among patients with HFpEF, the soluble guanylate cyclase stimulator praliciguat, compared with placebo, did not significantly improve peak V̇o(2) from baseline to week 12. These findings do not support the use of praliciguat in patients with HFpEF. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT03254485

Praliciguat, a clinical-stage soluble guanylate cyclase (sGC) stimulator, increases cGMP via the nitric oxide-sGC pathway. Praliciguat has been shown to be renoprotective in rodent models of hypertensive nephropathy and renal fibrosis. In the present study, praliciguat alone and in combination with enalapril attenuated proteinuria in the obese ZSF1 rat model of diabetic nephropathy. Praliciguat monotherapy did not affect hemodynamics. In contrast, enalapril monotherapy lowered blood pressure but did not attenuate proteinuria. Renal expression of genes in pathways involved in inflammation, fibrosis, oxidative stress, and kidney injury was lower in praliciguat-treated obese ZSF1 rats than in obese control rats; fasting glucose and cholesterol were also lower with praliciguat treatment. To gain insight into how tubular mechanisms might contribute to its pharmacological effects on the kidneys, we studied the effects of praliciguat on pathological processes and signaling pathways in cultured human primary renal proximal tubular epithelial cells (RPTCs). Praliciguat inhibited the expression of proinflammatory cytokines and secretion of monocyte chemoattractant protein-1 in tumor necrosis factor-α-challenged RPTCs. Praliciguat treatment also attenuated transforming growth factor-β-mediated apoptosis, changes to a mesenchyme-like cellular phenotype, and phosphorylation of SMAD3 in RPTCs. In conclusion, praliciguat improved proteinuria in the ZSF1 rat model of diabetic nephropathy, and its actions in human RPTCs suggest that tubular effects may contribute to its renal benefits, building upon strong evidence for the role of cGMP signaling in renal health.

Background: Impaired nitric oxide (NO) signaling has been implicated in the progression of diabetic kidney disease (DKD). Praliciguat (PRL) is a soluble guanylate cyclase stimulator that augments NO signaling in vitro and reduces proteinuria and fasting plasma glucose in the ZSF1 rat model of DKD. Methods: We evaluated the safety and efficacy of PRL in adults (25-75 y) with type 2 diabetes, eGFR 30-75 mL/min/1.73 m2, urine albumin creatinine ratio (UACR) 200-5000 mg/g, hemoglobin A1c (HbA1c) Results: A total of 156 participants enrolled and 140 completed treatment. Model estimates of mean UACR CFB [90% CI] [intent-to-treat (ITT)] were -28% [-36, -18] for pooled PRL and -15% [-27, 0] for PBO; PBO-adjusted UACR CFB was -15% [-31, 4] (p=0.17). Quality checks identified a site with data inconsistent with that from the overall study population. In a post-hoc sensitivity analysis excluding this site, PBO-adjusted UACR CFB for PRL was -20% [-33, 5], nominal p=0.03. PBO-adjusted CFB for other variables at week 12 (ITT) were 24 h systolic BP -4.2 mmHg [-7.5, -0.8], 24 h heart rate 3.4 bpm [1.6, 5.2], HbA1c -0.27% [-0.50, 0.03], and cholesterol 10.1 mg/dL [-19.2, -1.0]. Both praliciguat doses were well tolerated. Conclusion: PRL did not significantly reduce UACR in the primary ITT analysis, but positive trends in UACR, BP, and metabolic variables warrant further clinical study of PRL in DKD. Disclosure J.P. Hanrahan: Employee; Self; Cyclerion Therapeutics. I. de Boer: Advisory Panel; Self; Boehringer Ingelheim International GmbH. Consultant; Self; George Clinical, Goldfinch Bio, Ironwood Pharmaceuticals. G. Bakris: Consultant; Self; Alnylam, Merck & Co., Inc., Relypsa, Inc., Teijin Pharma Limited. Other Relationship; Self; Bayer AG, Novo Nordisk Inc., Vascular Dynamics. P. Wilson: Employee; Self; Cyclerion Therapeutics. J.D. Wakefield: Stock/Shareholder; Self; Cyclerion Therapeutics. J.P. Seferovic: Employee; Self; Cyclerion Therapeutics. J. Chickering: None. M. Cressman: Employee; Self; Covance, LabCorp. M. Currie: Employee; Self; Cyclerion Therapeutics. Stock/Shareholder; Self; Cyclerion Therapeutics. G.T. Milne: None. A.T. Profy: Employee; Self; Cyclerion Therapeutics, Ironwood Pharmaceuticals. Employee; Spouse/Partner; UpToDate. Stock/Shareholder; Self; Cyclerion Therapeutics, Ironwood Pharmaceuticals. Funding Cyclerion Therapeutics

The pharmacokinetics (PK), metabolism, excretion, mass balance, and tissue distribution of [14C]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 T max was 1 hour and the t 1/2 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 [14C]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. [14C]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.

Nitric oxide (NO)-sensitive soluble guanylyl cyclase (sGC), an enzyme that catalyzes the conversion of guanosine-5'-triphosphate (GTP) to cyclic guanosine-3',5'-monophophate (cGMP), transduces many of the physiological effects of the gasotransmitter NO. Upon binding of NO to the prosthetic heme group of sGC, a conformational change occurs, resulting in enzymatic activation and increased production of cGMP. cGMP modulates several downstream cellular and physiological responses, including but not limited to vasodilation. Impairment of this signaling system and altered NO-cGMP homeostasis have been implicated in cardiovascular, pulmonary, renal, gastrointestinal, central nervous system, and hepatic pathologies. sGC stimulators, small molecule drugs that synergistically increase sGC enzyme activity with NO, have shown great potential to treat a variety of diseases via modulation of NO-sGC-cGMP signaling. Here, we give an overview of novel, orally available sGC stimulators that Ironwood Pharmaceuticals is developing. We outline the non-clinical and clinical studies, highlighting pharmacological and pharmacokinetic (PK) profiles, including pharmacodynamic (PD) effects, and efficacy in a variety of disease models.

Background Heart failure with preserved ejection fraction (HFpEF) is a significant cause of morbidity and mortality worldwide. Exercise intolerance is the main symptom of HFpEF and is associated with a poor quality of life and increased mortality. Currently, there are no approved medications for the treatment of HFpEF. Praliciguat (IW-1973), a novel soluble guanylate cyclase stimulator that may help restore deficient nitric oxide–soluble guanylate cyclase–cyclic guanosine 3′,5′-monophosphate signaling, is being investigated for the treatment of patients with HFpEF. Methods CAPACITY HFpEF is a phase 2, multicenter, randomized, double-blind, placebo-controlled, parallel-group trial designed to evaluate the safety and efficacy of praliciguat over 12 weeks in approximately 184 patients with HFpEF. Eligible patients must have evidence supporting clinical HFpEF and at least 2 of the following 4 conditions associated with NO deficiency: diabetes/prediabetes, hypertension, obesity, and age >70 years. The primary efficacy end point is the change from baseline in peak VO2 by cardiopulmonary exercise test (CPET). Secondary end points include the change from baseline in 6-minute walk test distance and the change in ventilatory efficiency on CPET, as well as number of CPET responders. Other exploratory end points include changes in echocardiographic parameters, New York Heart Association functional classification, cardiac events, blood and urine biomarkers pathophysiologically relevant to heart failure, and patient-reported outcomes including Kansas City Cardiomyopathy Questionnaire. Conclusions The CAPACITY HFpEF trial will provide data on short-term safety and efficacy of praliciguat on peak exercise capacity, as well as multiple secondary end points of submaximal functional capacity, patient-reported outcomes, and biomarkers.

Aims/hypothesis Praliciguat (IW-1973), a soluble guanylate cyclase stimulator, amplifies nitric oxide signalling. This exploratory trial investigated the safety, tolerability, pharmacokinetic profile and pharmacodynamic effects of praliciguat in individuals with type 2 diabetes and hypertension. Methods This Phase IIA, double-blind, placebo-controlled trial investigated praliciguat in 26 participants with type 2 diabetes and hypertension on stable glucose- and BP-lowering therapies. Participants were randomly allocated in a 3:5:5 ratio to three groups: placebo (n = 6), praliciguat 40 mg once daily for days 1–14 (n = 10), or praliciguat 20 mg twice daily for days 1–7 then 40 mg once daily for days 8–14 (n = 10). Assessments were made in clinic and included treatment-emergent adverse events, pharmacokinetics, metabolic variables, 24 h BP and heart rate, platelet function, reactive hyperaemia index (RHI) and plasma biomarkers. Participants, the sponsor, the investigator and clinic study staff (except designated pharmacy personnel) were blinded to group assignment. Results Participants treated for 14 days with praliciguat had least-square mean change-from-baseline differences vs placebo (95% CI) of −0.7 (−1.8, 0.4) mmol/l for fasting plasma glucose, −0.7 (−1.1, −0.2) mmol/l for total cholesterol, −0.5 (−1.0, −0.1) mmol/l for LDL-cholesterol, −23 (−56, 9) for HOMA-IR in those not being treated with insulin, and −5 (−10, 1) mmHg and 3 (−1, 6) beats/min for average 24 h mean arterial pressure and heart rate, respectively. Apart from one serious adverse event (SAE; upper gastrointestinal haemorrhage), praliciguat was well tolerated. Praliciguat did not affect platelet function or RHI. Among exploratory biomarkers, plasma levels of asymmetric dimethylarginine decreased in praliciguat vs placebo recipients. Conclusions/interpretation In participants with type 2 diabetes and hypertension on standard therapies, over 14 days praliciguat was well tolerated, except for a single SAE, and showed positive trends in metabolic and BP variables. These results support further clinical investigation of praliciguat. Trial registration ClinicalTrials.gov NCT03091920. Funding This trial was funded by Cyclerion Therapeutics.

Introduction Heart failure with preserved ejection fraction (HFpEF) is associated with poor quality of life and increased mortality with exercise intolerance being the main symptom. Hypothesis The soluble guanylate cyclase (sGC) stimulator praliciguat (PRL) may help restore deficient NO-sGC-cGMP signaling and improve exercise capacity. Methods CAPACITY HFpEF was a phase 2, multicenter, randomized, double-blind, placebo-controlled trial to evaluate the safety and efficacy of PRL over 12 weeks in patients with HFpEF and at least 2 of 4 conditions potentially associated with NO deficiency (diabetes, hypertension, obesity, age >70 years). Patients were randomized to PRL or placebo. The primary endpoints were change from baseline to week 12 in peak oxygen consumption (pVO2) by cardiopulmonary exercise testing and treatment-emergent adverse events (TEAEs). Secondary endpoints included the change in 6-minute walk distance (6MWD), ventilatory efficiency (VE/VCO2 slope), and the proportion of pVO2 responders (change ≥1.5 mL/kg/min). Health status was assessed using the Kansas City Cardiomyopathy Questionnaire (KCCQ). Results Among 196 patients randomized (mean age 70.3 years, 57% men), 169 completed. For the evaluable population (completed 8 weeks of dosing with no dose reduction), mean baseline pVO2 was 13.3 and 12.8 mL O2/kg/min in PRL 40 mg (N=65) and placebo (N=78) groups, respectively. Placebo-adjusted least squares mean changes from baseline to week 12 [95% CI] were pVO2 (-0.30 mL O2/kg/min [-0.95, 0.35] p=0.37), 6MWD (-16.7 m [-47.4, 13.9]) and VE/VCO2 slope (-0.30 [-1.59, 1.00]). The odds ratio for pVO2 response [95% CI] was 0.91 [0.40, 2.1]. KCCQ overall and domain scores did not improve in the PRL group versus the placebo group. PRL was generally well tolerated; headache, dizziness, and hypotension TEAEs were reported more frequently in the PRL group. The frequency of serious TEAEs was similar between groups. Conclusion In a population selected to enrich for the metabolic/NO-deficient profile of HFpEF, PRL was well tolerated but did not affect pVO2, 6MWD, VE/VCO2 slope, proportion of pVO2 responders or KCCQ scores over 12 weeks compared to placebo.

Background: Impaired nitric oxide (NO) signaling through soluble guanylate cyclase (sGC) has been associated with microvascular complications of diabetes. Praliciguat (IW-1973), an sGC stimulator that enhances NO signaling, reduced fasting plasma glucose and proteinuria in an animal model of diabetic nephropathy. In a Phase 1 clinical study in healthy adults, doses of 15-40 mg were tolerated and lowered blood pressure (BP). Methods: This Phase 2a open-label in-clinic study assessed the safety, tolerability, pharmacokinetics (PK) and pharmacodynamics of oral praliciguat in 11 patients with type 2 diabetes mellitus (T2DM) and hypertension on stable glycemic and BP therapy. Patients received sequential 3-day dosing cycles escalating through placebo (PBO), 10, 20, 30, 40 and 50 mg praliciguat QD. Assessments included adverse events (AEs), blood chemistry, ambulatory BP and heart rate, platelet function, and reactive hyperemia index (RHI). Results: The escalating dose regimen was generally well tolerated; there were no serious or severe AEs. Escalation in 2 patients was stopped due to asymptomatic BP lowering. Overall PK showed a median Tmax of 1-3 h, low to moderate variability, and ∼2-fold accumulation supporting QD dosing. Average 12-h daytime mean arterial BP decreased by 2 to 9 mmHg relative to time-matched PBO baseline across praliciguat dose levels, while heart rate increased 4 to 7 bpm. Notable changes from PBO baseline [mean (95% CI)] at the end of treatment were seen for fasting serum glucose [-26.5 mg/dL (-43.4, -9.5], HbA1c [-0.4% (-0.8, -0.04)], cholesterol [-43 mg/dL (-67, -20)], and body weight [-3.5 kg (-4.9, -2.0)]. Praliciguat had no effect on RHI or platelet function. Conclusion: T2DM patients with hypertension on standard therapy showed improvements in hemodynamics and metabolic parameters in this small in-clinic study. These results support further evaluation of praliciguat in T2DM patients including those with complications. Disclosure J.P. Hanrahan: Employee; Self; Ironwood. J.D. Wakefield: Employee; Self; Ironwood Pharmaceuticals, Inc.. Stock/Shareholder; Self; Ironwood Pharmaceuticals, Inc.. P.J. Wilson: None. D.P. Zimmer: Employee; Self; Ironwood Pharmaceuticals, Inc.. Stock/Shareholder; Self; Ironwood Pharmaceuticals, Inc.. M. Mihova: None. J. Chickering: None. D. Ruff: None. M.L. Hall: Employee; Self; Ironwood Pharmaceuticals, Inc.. Stock/Shareholder; Self; Ironwood Pharmaceuticals, Inc. M. Currie: Employee; Self; Ironwood Pharmaceuticals, Inc. G. Milne: Employee; Self; Ironwood Pharmaceuticals, Inc.. Stock/Shareholder; Self; Ironwood Pharmaceuticals, Inc.. Employee; Spouse/Partner; Catabasis Pharmaceuticals. Stock/Shareholder; Spouse/Partner; Catabasis Pharmaceuticals. A.T. Profy: Other Relationship; Self; Ironwood Pharmaceuticals.

Background: Praliciguat (IW-1973), a soluble guanylate cyclase stimulator, has been shown to augment nitric oxide (NO) signaling and reduce fasting plasma glucose and proteinuria in an animal model of diabetic nephropathy. In healthy adults, repeated oral doses of 15-40 mg were tolerated and lowered blood pressure (BP). Methods: This Phase 2a, double-blind, placebo (PBO)-controlled study assessed the effects of oral praliciguat in 26 patients with type 2 diabetes mellitus (T2DM) and hypertension on stable glycemic and BP-lowering therapy. Most (69%) were also on statins. Two praliciguat regimens were tested: (1) 40 mg QD for days 1-14 (N=10), and (2) 20 mg BID for days 1-7 then 40 mg QD for days 8-14 (N=10). Assessments included serum chemistry, fasting plasma glucose, BP and heart rate, platelet function, reactive hyperemia index (RHI), plasma biomarkers, and adverse events (AEs). Results: Results were similar for both praliciguat regimens and are here combined. After 14 days of treatment, praliciguat-treated patients had notable differences in least squares mean change from baseline (95% CI) relative to PBO in 24-hour mean arterial pressure [-4.7 mmHg (-10, 0.8)], heart rate [2.9 bpm (-0.7, 6.5)], fasting glucose [-13 mg/dL (-32, 7)], HOMA-IR [-26 (-58, 7)], total serum cholesterol [-26 mg/dL (-44, -7)], and LDL cholesterol [-20 mg/dL (-37, -3)]. Praliciguat had no effect on platelet function or RHI. Among the exploratory biomarkers tested, levels of the endogenous NO synthase inhibitor asymmetric dimethylarginine were lower in praliciguat vs. PBO recipients. Both praliciguat dose regimens were generally well tolerated. Except for one serious AE of upper gastrointestinal hemorrhage, AEs in praliciguat-treated patients were mild. Conclusion: In this initial trial, praliciguat had beneficial effects on BP and metabolic parameters in T2DM patients with hypertension on standard therapies. Further clinical investigation of praliciguat in T2DM patients is warranted. Disclosure J.P. Hanrahan: Employee; Self; Ironwood. J.D. Wakefield: Employee; Self; Ironwood Pharmaceuticals, Inc.. Stock/Shareholder; Self; Ironwood Pharmaceuticals, Inc.. P.J. Wilson: None. P. Miller: None. J. Chickering: None. L. Morrow: Stock/Shareholder; Self; ProSciento. Employee; Spouse/Partner; Eli Lilly and Company. M.L. Hall: Employee; Self; Ironwood Pharmaceuticals, Inc.. Stock/Shareholder; Self; Ironwood Pharmaceuticals, Inc. M. Currie: Employee; Self; Ironwood Pharmaceuticals, Inc. G. Milne: Employee; Self; Ironwood Pharmaceuticals, Inc.. Stock/Shareholder; Self; Ironwood Pharmaceuticals, Inc.. Employee; Spouse/Partner; Catabasis Pharmaceuticals. Stock/Shareholder; Spouse/Partner; Catabasis Pharmaceuticals. A.T. Profy: Other Relationship; Self; Ironwood Pharmaceuticals.

Nitric oxide (NO)-soluble guanylate cyclase (sGC)-cyclic guanosine monophosphate (cGMP) signaling is central to the regulation of several physiological processes, including blood flow and inflammation. Deficient NO signaling is implicated in multiple diseases. sGC stimulators are small molecules that enhance sGC activity, particularly in combination with NO. In a randomized, placebo-controlled phase 1 study, the safety, tolerability, pharmacokinetics, and pharmacodynamics of multiple ascending doses of the sGC stimulator praliciguat were assessed in 44 healthy adults. Four cohorts of 11 subjects (8 praliciguat, 3 placebo) received once-daily praliciguat for 14 days before up-titrating for 7 days (treatment sequences: 15/30 mg, 20/40 mg, 30/40 mg, and weight-based). All doses were tolerated. No serious or severe adverse events (AEs) were reported. The most common AEs in praliciguat recipients were headache and symptoms consistent with blood pressure (BP) lowering/vasodilation. There were no laboratory, vital sign, electrocardiographic, or platelet function findings indicative of a safety concern. Pharmacokinetics were dose proportional, with an effective half-life of 24-37 hours, supporting once-daily dosing. Praliciguat produced dose-related increases in plasma cGMP consistent with stimulation of sGC. Repeated once-daily dosing showed sustained decreases in BP. Results support evaluation of praliciguat for the treatment of conditions associated with deficient NO signaling.