Soluble guanylate cyclase (sGC) - activator (heme-free form); EC50 = 11.2 ± 1.0 nM in sGC-overexpressing CHO reporter cell line; EC50 = 2.1 ± 0.07 nM in sGC-overexpressing CHO reporter cell line pretreated with ODQ (30 μM, 3 h). [1]
Runcaciguat targets soluble guanylate cyclase (sGC), a key enzyme in the nitric oxide (NO)-cGMP signaling pathway. As an sGC activator, runcaciguat stimulates the production of cGMP from GTP, independent of NO. The increase in cGMP leads to activation of cGMP-dependent protein kinases (PKG), which mediate vasodilation, inhibition of platelet aggregation, and anti-inflammatory effects. The compound's mechanism of action is particularly relevant in conditions where NO bioavailability is reduced, such as in chronic kidney disease and heart failure.

Runcaciguat (concentrations 0.01 to 100 μM) on highly purified recombinant sGC enzyme exhibited concentration-dependent activation ranging from 2.6-fold (0.01 μM) to 9.3-fold (100 μM) in cGMP production. [1]
Runcaciguat showed additive effects on sGC activity in the presence of NO (DEA/NO, 10 μM) over a wide range of concentrations. [1]
In the presence of the sGC inhibitor ODQ (10 μM), which leads to oxidized/heme-free sGC, cGMP production was further increased by Runcaciguat, showing the maximum effect on the heme-free enzyme. [1]
Runcaciguat activated the sGC reporter cell line with an EC50 of 11.2 ± 1.0 nM; pretreatment with ODQ (30 μM, 3 h) increased potency (EC50 = 2.1 ± 0.07 nM). [1]
Runcaciguat in combination with the NO donor SNAP (10 and 100 nM) showed additive effects. [1]
Runcaciguat treatment resulted in maximal luminescence signals of 50-60% compared to cinaciguat in sGC-overexpressing cells. [1]
Runcaciguat concentration-dependently inhibited phenylephrine-induced contractions of rabbit saphenous artery rings (IC50 = 199 nM) and rabbit aortic rings (IC50 = 39 nM). [1]
Runcaciguat concentration-dependently relaxed porcine coronary artery rings precontracted by the thromboxane agonist U-46619 with an IC50 of 137 nM. [1]
In the rat Langendorff heart preparation, Runcaciguat reduced coronary perfusion pressure in a concentration-dependent manner from 10 nM to 10 μM with maximal effect of ~45% at the highest concentration; no effect on heart rate, left ventricular diastolic pressure, or contractility was observed up to the highest concentration tested. [1]
In vitro, runcaciguat has been shown to activate sGC and increase cGMP production in various cell types. Its effects on vascular smooth muscle relaxation, platelet aggregation inhibition, and anti-inflammatory signaling have been characterized. The compound's potency and efficacy have been evaluated in cell-based assays using sGC-expressing cell lines. Studies have examined its effects on endothelial function and vascular tone. The compound's ability to activate sGC in the presence of oxidative stress has been investigated.

Runcaciguat (iv bolus 0.03 and 0.1 mg/kg) produced dose-dependent and long-lasting decreases in blood pressure with minor effects on heart rate in anaesthetized rats; effects were more pronounced and longer lasting under ODQ pretreatment. [1]
Runcaciguat (oral 10 mg/kg) caused dose-related and long-lasting decreases in mean arterial blood pressure in conscious normotensive rats with telemetric implants, lowering blood pressure by up to 15%; peak levels within the first hours and maintained over >24 h; 1 and 3 mg/kg had no effect on blood pressure; dose-related reflex tachycardia was noted. [1]
Runcaciguat (oral 0.3, 1.0, and 3.0 mg/kg) caused dose-related and long-lasting decreases in mean arterial blood pressure in conscious spontaneously hypertensive rats (SHR), lowering blood pressure by 5-15%; effects peaked within 1-12 h, plateaued for ~17 h, with substantial effect persisting at 24 h after 1.0 and 3.0 mg/kg doses; dose-related transient reflex tachycardia up to 20% was observed. [1]
In the L-NAME-treated renin transgenic rat model (TGR(mRenR2)27), Runcaciguat (1, 3, and 10 mg/kg BID, oral) significantly and dose-dependently decreased mortality (58% in placebo, 56% at 1 mg/kg, 39% at 3 mg/kg, 28% at 10 mg/kg; 10 mg/kg statistically significant, 3 mg/kg showed strong trend). [1]
Runcaciguat treatment resulted in dose-dependent and significant reduction of proteinuria and significant increase of creatinine clearance at the end of the study, indicating kidney protective effects. [1]
Runcaciguat treatment significantly reduced heart hypertrophy (right and left ventricle), indicating cardioprotective effects. [1]
In vivo, runcaciguat has been shown to reduce blood pressure and improve renal function in animal models of cardiovascular and renal disease. The compound has been evaluated in models of hypertension, chronic kidney disease, and heart failure. Its effects on vascular function, renal hemodynamics, and cardiac function have been characterized. The compound's ability to improve outcomes in disease models has been demonstrated. Clinical trials have been conducted to evaluate its safety and efficacy in patients with chronic kidney disease and other conditions.

Highly purified recombinant sGC enzyme assay: Runcaciguat was tested in concentrations ranging from 0.01 to 100 μM for its capability to stimulate cGMP production on purified sGC enzyme. Concentration-dependent activation was observed ranging from 2.6-fold (0.01 μM) to 9.3-fold (100 μM). Additive effects with NO (DEA/NO 10 μM) were tested. In the presence of ODQ (10 μM, leading to oxidized/heme-free sGC), cGMP production was further increased. The assay was performed as described previously for characterization of sGC stimulators and sGC activators. [1]
Cell-based cGMP formation assay in sGC-overexpressing CHO cells: A stable CHO reporter cell line overexpressing rat sGC was used. The minimum effective concentration (MEC) was defined as the concentration required to achieve a ≥ threefold stimulation of cGMP formation. EC50 values were determined with a concentration-response curve. For ODQ-pretreatment experiments, cells were incubated with 30 μM ODQ for 3 h before Runcaciguat addition. For combination studies, Runcaciguat was tested with the NO donor SNAP (10 and 100 nM). Luminescence signals were measured and compared to cinaciguat as control. [1]
Vasorelaxation assay on isolated vessels: Rabbit saphenous artery rings and rabbit aortic rings were precontracted with phenylephrine, and Runcaciguat was added to determine concentration-dependent inhibition with IC50 values. Porcine coronary artery rings were precontracted with the thromboxane agonist U-46619, and Runcaciguat was added to determine relaxation with IC50 values. [1]
Langendorff-perfused heart assay: Rat hearts were perfused in Langendorff preparation; Runcaciguat was tested from 10 nM to 10 μM to assess effects on coronary perfusion pressure, heart rate, left ventricular diastolic pressure, and contractility (+dp/dt). [1]
In cell-free biochemical assays, runcaciguat is evaluated for its ability to activate sGC and stimulate cGMP production. Enzyme activity assays measure the compound's ability to increase cGMP production in the presence of purified sGC. Its potency and efficacy are compared to other sGC activators and stimulators. The compound's mechanism of action, including its binding site on sGC, is characterized. These assays confirm the compound's activity as an sGC activator.

sGC-overexpressing CHO reporter cell line assay: Cells were used to assess sGC activation by Runcaciguat; EC50 was determined as 11.2 ± 1.0 nM. Pretreatment with ODQ (30 μM, 3 h) increased potency to EC50 = 2.1 ± 0.07 nM. Combination with the NO donor SNAP (10 and 100 nM) showed additive effects. Maximal luminescence signals reached 50-60% compared to cinaciguat. [1]
Vasorelaxation experiments on isolated vessels: Rabbit saphenous artery, rabbit aortic, and porcine coronary artery rings were used. Tissues were precontracted with phenylephrine or U-46619, then treated with Runcaciguat to determine IC50 values (199 nM for saphenous artery, 39 nM for aortic rings, 137 nM for coronary artery rings). [1]
Langendorff-perfused rat heart assay: Rat hearts were mounted in Langendorff apparatus; Runcaciguat was perfused at concentrations from 10 nM to 10 μM; coronary perfusion pressure, heart rate, left ventricular diastolic pressure, and contractility were measured. [1]
Cellular assays for runcaciguat involve evaluating its effects on cGMP production and downstream signaling in various cell types. The compound's ability to activate sGC in vascular smooth muscle cells, endothelial cells, and renal cells is assessed. Its effects on vasodilation, anti-inflammatory signaling, and cellular protection are evaluated. Studies have examined its effects on cGMP-dependent signaling pathways and downstream gene expression.

Anaesthetized rat hemodynamics: Wistar rats were anesthetized; Runcaciguat was administered as intravenous bolus at 0.03 and 0.1 mg/kg; blood pressure and heart rate were monitored. Experiments were performed in the absence and presence of ODQ pretreatment. [1]
Conscious normotensive rat telemetry: Rats with telemetric implants received oral Runcaciguat at 1, 3, and 10 mg/kg; mean arterial blood pressure and heart rate were monitored continuously; effects were assessed over 24 hours. [1]
Conscious spontaneously hypertensive rat (SHR) study: SHR received oral Runcaciguat at 0.1, 0.3, 1.0, and 3.0 mg/kg; blood pressure and heart rate were monitored; effects were assessed over 24 hours. [1]
Long-term L-NAME-treated renin transgenic rat model: TGR(mRenR2)27 rats were treated with L-NAME to induce endothelial dysfunction; Runcaciguat was administered orally at 1, 3, and 10 mg/kg BID for 7 weeks. Mortality, proteinuria, creatinine clearance, and heart weight were assessed. [1]
Animal models for runcaciguat include models of hypertension, chronic kidney disease, heart failure, and other cardiovascular and renal diseases. The compound is typically administered orally. Studies have examined its effects on blood pressure, renal function, cardiac function, and survival. Its pharmacokinetic properties, including oral bioavailability and half-life, have been characterized in preclinical species. The compound's efficacy in disease models supports its clinical development.

Pharmacokinetic parameters of Runcaciguat in Wistar rats (iv 0.3 mg/kg): Vss = 1.4 L/kg, CLb = 0.21 L/(h·kg), t1/2 = 7.5 h; po 0.3 mg/kg, bioavailability F = 95%. [1]
Pharmacokinetic parameters of Runcaciguat in Beagle dogs (iv 0.3 mg/kg): Vss = 2.8 L/kg, CLb = 1.4 L/(h·kg), t1/2 = 4.3 h; po 0.3 mg/kg, bioavailability F = 8%. [1]
Pharmacokinetic parameters of Runcaciguat in cynomolgus monkeys (iv 0.1 mg/kg): Vss = 1.4 L/kg, CLb = 0.16 L/(h·kg), t1/2 = 10 h; po 0.1 mg/kg, bioavailability F = 81%. [1]
Runcaciguat showed no relevant inhibitory effects on major CYP isoforms. [1]
Pharmacokinetic data for runcaciguat are available from clinical studies. The compound is orally bioavailable and has been evaluated in Phase 1 and Phase 2 clinical trials. Its half-life, clearance, and volume of distribution have been characterized. The compound's pharmacokinetic profile supports once-daily dosing. The compound is for research use only and is not approved for clinical use outside of clinical trials.

CYP3A4 induction potential: Runcaciguat (compound 45) was evaluated for CYP3A4 induction in primary cultures of human hepatocytes from three donors. The cyclopropyl propionic acid compound 45 ultimately provided the best overall profile with a positive assessment of the predicted safety margin regarding human CYP3A4 induction potential. No relevant inhibitory effects on major CYP isoforms or off-target effects were observed. [1]
No specific toxicity data such as LD50, hepatotoxicity, nephrotoxicity, or plasma protein binding were reported in this study. [1]
The toxicity profile of runcaciguat is typical of sGC activators. The compound has been evaluated in clinical trials for safety and tolerability. Common side effects may include hypotension, headache, and dizziness due to its vasodilatory effects. Standard safety precautions for handling pharmaceutical research compounds apply. The compound is for research use only and not for human use except in approved clinical trial settings.

J Med Chem. 2021 May 13;64(9):5323-5344.

Runcaciguat is a potent and selective sGC activator that activates the oxidized and heme-free form of sGC, which is formed under oxidative stress conditions. This mode of action restores the dysfunctional NO/sGC/cGMP signaling pathway, which is impaired in cardiovascular and cardiorenal diseases. [1]
First-generation sGC activators like cinaciguat (intravenous only, unfavorable PK/PD profile) and ataciguat (weak activity, discontinued in phase 2) had limitations that were overcome with Runcaciguat. [1]
X-ray crystal structure of Runcaciguat bound to the Nostoc sp. H-NOX domain (homologous to human sGC, 35% sequence identity) at 2.2 Å resolution (PDB code 7LGK) showed that the carboxylic acid replaces one of the carboxyl-butyl moieties of cinaciguat with the same hydrogen-bond pattern to the YxSxR motif; the second carboxylic acid site is occupied by a chloride ion. The central amide is sandwiched between H105 and W74; the αF helix is shifted ~2 Å from its heme-bound position, opening space for binding the central chlorinated phenyl ring; the cyclopropyl moiety binds into a hydrophobic pocket formed by M1, V5, M144, and W74. [1]
Runcaciguat is currently being investigated in clinical phase 2 trials: CONCORD trial (NCT04507061) for chronic kidney disease (CKD) and NEON trial (NCT04722991) for nonproliferative diabetic retinopathy (NPDR). [1]
In vitro metabolism data showed a wide and variable distribution of different metabolic pathways including conjugation, oxidative, and hydrolytic reactions. [1]
The compound has a favorable profile with low clearance and long half-lives after intravenous administration and high oral bioavailability in rats and monkeys, with moderate clearance and oral bioavailability in dogs. [1]
Runcaciguat is a soluble guanylate cyclase (sGC) activator being developed for the treatment of cardiovascular and renal diseases. It activates sGC, increasing cGMP production and mediating vasodilation and other protective effects. The compound is in clinical development for chronic kidney disease, heart failure, and hypertension. It is for research use only and is not approved for clinical use outside of clinical trials. The CAS number is 1402936-61-1.

C23H22CL2F3NO3

488.326895236969

487.092

C, 56.57; H, 4.54; Cl, 14.52; F, 11.67; N, 2.87; O, 9.83

1402936-61-1

1402936-61-1;

134440946

White to off-white solid powder

6.6

2

6

8

32

665

3

ClC1C=CC(=CC=1NC([C@H](C1C=CC(=CC=1)Cl)[C@@H](C)C(F)(F)F)=O)[C@@H](CC(=O)O)C1CC1

NCRMKIWHFXSBGZ-CNBXIYLPSA-N

InChI=1S/C23H22Cl2F3NO3/c1-12(23(26,27)28)21(14-4-7-16(24)8-5-14)22(32)29-19-10-15(6-9-18(19)25)17(11-20(30)31)13-2-3-13/h4-10,12-13,17,21H,2-3,11H2,1H3,(H,29,32)(H,30,31)/t12-,17+,21+/m1/s1

(3S)-3-{4-chloro-3-[(2S,3R)-2-(4-chlorophenyl)-4,4,4- trifluoro-3-methylbutanamido]phenyl}-3- cyclopropylpropanoic acid

BAY-1101042 BAY1101042 Runcaciguat

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: This product requires protection from light (avoid light exposure) during transportation and storage.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~125 mg/mL (~255.97 mM)

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

---

Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

View More

Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

---

Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

View More

Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

---

Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

---

 (Please use freshly prepared in vivo formulations for optimal results.)