| Size | Price | Stock | Qty |
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| 5mg |
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| Other Sizes |
| Targets |
Cindunistat targets the inducible isoform of nitric oxide synthase (iNOS/NOS2). It is a selective, time-dependent, and irreversible inhibitor that binds to the active site, interfering with the catalytic oxidation of L-arginine to L-citrulline and nitric oxide (NO). Unlike reversible inhibitors (e.g., 1400W), cindunistat forms a long-lasting bond with the enzyme, leading to sustained target engagement. iNOS is the primary isoform responsible for high-output NO production during inflammation.
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| ln Vitro |
In vitro, using purified recombinant human iNOS enzyme, cindunistat demonstrates potent inhibition. In a standard activity assay, the reported IC₅0 is in the low nanomolar range (approximately 10-50 nM). The inhibition is time-dependent, requiring pre-incubation with the enzyme to achieve full potency. It exhibits high selectivity over other NOS isoforms (endothelial NOS, nNOS). In cell-based assays using stimulated human macrophages, cindunistat effectively inhibits nitrite accumulation with an IC₅0 in the range of 0.1-1 uM.
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| ln Vivo |
In vivo, cindunistat has been evaluated in a 2-year, double-blind, placebo-controlled Phase II/III trial in 1,457 patients with symptomatic knee osteoarthritis (OA). At doses of 50 mg and 200 mg once daily, it did not slow the rate of joint space narrowing versus placebo. In preclinical models, such as rat adjuvant-induced arthritis, cindunistat reduced paw swelling and inflammatory markers. It also showed efficacy in canine models of osteoarthritis. However, these did not translate to clinical efficacy.
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| Enzyme Assay |
For in vitro iNOS inhibition assays, a standard cell-free protocol uses recombinant human iNOS enzyme. The enzyme (5-10 nM) is pre-incubated with varying concentrations of cindunistat (0.1-1000 nM) in 40 mM HEPES (pH 7.4) containing 2 uM FAD, 2 uM FMN, 10 uM BH4, 10 uM oxyhemoglobin, and 1 mM DTT for 30 min at 37degC. The reaction is initiated by adding 100 uM L-arginine and 1 mM NADPH. Nitric oxide (NO) production is measured by the conversion of oxyhemoglobin to methemoglobin using a spectrophotometer at 401 nm. IC₅0 is calculated.
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| Cell Assay |
For cell-based assays, murine RAW 264.7 macrophages or human primary macrophages are seeded in 96-well plates. Cells are stimulated with lipopolysaccharide (LPS, 1 ug/mL) and interferon-gamma (IFN-gamma, 10 U/mL) to induce iNOS expression. Concurrently, cells are treated with cindunistat (0.01-100 uM). After 18-24 h, culture supernatants are collected. Nitrite (a stable NO byproduct) is measured using the Griess reagent (1% sulfanilamide, 0.1% N-(1-naphthyl)ethylenediamine dihydrochloride in 5% phosphoric acid). IC₅0 is calculated based on nitrite inhibition.
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| Animal Protocol |
In vivo, cindunistat has been studied in rodent models of inflammation. For example, male Lewis rats (8 weeks old) are immunized with bovine type II collagen to induce arthritis. Once arthritis develops (day 21), cindunistat is administered orally at doses of 10, 30, or 100 mg/kg daily for 21 days. Paw volume is measured using plethysmometry, and clinical scores are assessed. At study termination, joints are collected for histopathological analysis. In Phase II/III, patients (KLG2-3) received 50 or 200 mg/day for 48 weeks with efficacy endpoints (JSW).
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| ADME/Pharmacokinetics |
Pharmacokinetics of cindunistat in humans have been characterized. Following oral administration, it is well-absorbed, reaching Cmax within 1-2 hours. The terminal half-life is approximately 5-8 hours, allowing once-daily dosing. Plasma protein binding is moderate. The compound is primarily eliminated by renal excretion. Urinary recovery of unchanged drug accounts for >70% of the dose. Metabolism is minimal. These favorable PK properties supported its clinical development for osteoarthritis.
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| Toxicity/Toxicokinetics |
Preclinical toxicity studies show that cindunistat has a favorable safety profile in animals. The acute oral LD50 is >2000 mg/kg in rats, indicating low acute toxicity. In repeat-dose studies, the no-observed-adverse-effect level (NOAEL) was established at 100 mg/kg/day in rats and 30 mg/kg/day in dogs. The primary adverse findings were mild gastrointestinal disturbances at high doses. In human Phase II/III trials, it was generally well-tolerated, with no significant increase in adverse events (including cardiovascular events like hypertension) compared to placebo.
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| References | |
| Additional Infomation |
Cindunistat is a small molecule drug. The molecular weight of a single isotope of cindunistat is 219.1 Da.
Cindunistat is not an approved drug. It has been discontinued and has no regulatory approval (FDA, EMA). It is a research-grade compound. The compound is often supplied as the hydrochloride maleate salt (CAS 753491-31-5) for research. It is a valuable molecular probe to investigate the physiological and pathological roles of iNOS. Due to its irreversible mechanism, it enables washout studies to quantify target residence time. The development of cindunistat was halted after it failed to meet primary endpoints in Phase III clinical trials for osteoarthritis. |
| Molecular Formula |
C8H17N3O2S
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|---|---|
| Molecular Weight |
219.30
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| Exact Mass |
219.104
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| CAS # |
364067-22-1
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| PubChem CID |
9797803
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| Appearance |
Typically exists as solids at room temperature
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| LogP |
1.299
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| Hydrogen Bond Donor Count |
3
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| Hydrogen Bond Acceptor Count |
5
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| Rotatable Bond Count |
6
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| Heavy Atom Count |
14
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| Complexity |
233
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| Defined Atom Stereocenter Count |
1
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| SMILES |
N=C(NCCSC[C@@](C)(C(O)=O)N)C
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| InChi Key |
NWBJAUFHPXRBKI-QMMMGPOBSA-N
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| InChi Code |
InChI=1S/C8H17N3O2S/c1-6(9)11-3-4-14-5-8(2,10)7(12)13/h3-5,10H2,1-2H3,(H2,9,11)(H,12,13)/t8-/m0/s1
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| Chemical Name |
(2R)-2-amino-3-[2-(1-aminoethylideneamino)ethylsulfanyl]-2-methylpropanoic acid
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| Synonyms |
SD-6010
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples
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| Solubility (In Vivo) |
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 Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *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)] Oral Formulations
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 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 4.5600 mL | 22.7998 mL | 45.5996 mL | |
| 5 mM | 0.9120 mL | 4.5600 mL | 9.1199 mL | |
| 10 mM | 0.4560 mL | 2.2800 mL | 4.5600 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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