| Targets |
NP3-253 targets the NLRP3 (NOD-like receptor family pyrin domain containing 3) inflammasome, a key component of the innate immune system. The NLRP3 inflammasome is a multiprotein complex that is activated by various danger signals (PAMPs and DAMPs), leading to the activation of caspase-1, which cleaves pro-IL-1beta and pro-IL-18 into their mature forms, and induces pyroptosis (inflammatory cell death). Dysregulation of the NLRP3 inflammasome is implicated in numerous inflammatory and autoimmune diseases, including gout, type 2 diabetes, atherosclerosis, Alzheimer‘s disease, Parkinson's disease, and multiple sclerosis. NP3-253 is an NLRP3 inhibitor that blocks inflammasome activation, reducing IL-1beta and IL-18 production and pyroptosis.
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| ln Vitro |
In vitro, NP3-253 is an NLRP3 inhibitor. It is used to study the biological properties of NLRP3 in peripheral and neuroinflammatory models. The compound is orally active and can cross the blood-brain barrier. Specific IC₅0 values for NLRP3 inhibition are not provided. NP3-253 is a research-grade tool for exploring the role of NLRP3 in inflammation and neuroinflammation.
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| ln Vivo |
In vivo, NP3-253 is an orally active NLRP3 inhibitor that can cross the blood-brain barrier. It can be used as a tool to further explore the biological properties of NLRP3 in peripheral and neuroinflammatory models. The compound is for research use only and is not an approved drug. No specific in vivo efficacy data is provided.
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| Enzyme Assay |
The in vitro NLRP3 inflammasome activity can be measured in LPS-primed, ATP-stimulated macrophages. Human THP-1 monocytes are differentiated into macrophages using PMA (50 ng/mL) for 48-72 h. Cells are seeded in 96-well plates (1×10⁵ cells/well) and primed with LPS (100 ng/mL) for 4 h. NP3-253 (0.1-100 uM) is added for 30 min, then ATP (5 mM) is added for 30-60 min to activate the NLRP3 inflammasome. The supernatant is collected, and IL-1beta and IL-18 levels are measured by ELISA. Caspase-1 activity is measured using a fluorogenic substrate (YVAD-AMC). For pyroptosis, LDH release is measured.
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| Cell Assay |
For cell-based studies, THP-1 macrophages or primary mouse macrophages are used for IL-1beta secretion assays as described. Cell viability is assessed by MTT assay. NLRP3 inflammasome activation is confirmed by Western blotting for caspase-1 p20 and IL-1beta p17 in the supernatant. For neuroinflammation studies, microglial cells (BV-2 or primary microglia) are used. Cells are primed with LPS (100 ng/mL) and then treated with nigericin (10 uM) or ATP (5 mM) to activate NLRP3. NP3-253 inhibits IL-1beta secretion in a concentration-dependent manner.
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| Animal Protocol |
In vivo efficacy can be evaluated in a mouse model of acute peritonitis. Female C57BL/6 mice (6-8 wk, n=10/group) are injected intraperitoneally (IP) with LPS (10 mg/kg) to induce systemic inflammation. NP3-253 is formulated in 10% DMSO + 5% Tween 80 + 85% saline and administered orally (PO) at doses of 10-100 mg/kg 1 h before LPS injection. After 6 h, mice are euthanized, and peritoneal lavage fluid is collected. IL-1beta and IL-18 levels in the lavage fluid are measured by ELISA. For a neuroinflammation model (e.g., LPS-induced neuroinflammation), C57BL/6 mice are injected intraperitoneally with LPS (5 mg/kg), and NP3-253 is administered orally daily for 3-7 days. Brain tissues are harvested for cytokine analysis (IL-1beta, IL-18) and immunohistochemistry (Iba-1 for microglia).
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| ADME/Pharmacokinetics |
No specific PK data for NP3-253 is available. As a small molecule (MW 380.41), it is expected to have good oral bioavailability. It is blood-brain barrier-permeable. Solubility: May dissolve in DMSO. For in vivo studies, it can be formulated in 10% DMSO + 5% Tween 80 + 85% saline or 10% DMSO + 40% PEG300 + 5% Tween 80 + 45% saline for IP administration, or in 0.5% CMC-Na for oral administration. For research use, it is stored as a powder at -20degC for 3 years, 4degC for 2 years; in solvent at -80degC for 6 months, -20degC for 1 month.
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| Toxicity/Toxicokinetics |
For NP3-253, hazard statements: H315 (Causes skin irritation), H319 (Causes serious eye irritation), H335 (May cause respiratory irritation). Signal word: Warning. Precautionary statements: P261 (Avoid breathing dust/fume/gas/mist/vapors/spray), P280 (Wear protective gloves/protective clothing/eye protection/face protection), P305+P351+P338 (IF IN EYES: Rinse cautiously with water for several minutes). Storage: at -20degC, sealed, protect from light.
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| References | |
| Additional Infomation |
NP3-253 (CAS# 2557349-50-3) is a research-grade, orally active, and blood-brain-barrier-permeable NLRP3 inflammasome inhibitor. It is not an FDA-approved drug. It is used as a tool to study NLRP3 biology in peripheral and neuroinflammatory models. For research use only, not for diagnostic or therapeutic applications.
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| CAS # |
2557349-50-3
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| Appearance |
Typically exists as solids at room temperature
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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.) |
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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