| Size | Price | Stock | Qty |
|---|---|---|---|
| 100mg |
|
||
| 250mg |
|
||
| 500mg |
|
| Targets |
The primary molecular target of SARS-CoV-2 3CLpro-IN-20 is the SARS-CoV-2 3-chymotrypsin-like protease (3CLpro), also known as the main protease (Mpro). 3CLpro is a cysteine protease that plays a critical role in the life cycle of the SARS-CoV-2 virus. It is responsible for cleaving the viral polyproteins (pp1a and pp1ab) at 11 different sites, producing functional non-structural proteins that are essential for viral replication and transcription. The protease is highly conserved among coronaviruses, making it an attractive target for broad-spectrum antiviral drug development. SARS-CoV-2 3CLpro-IN-20 is a covalent inhibitor that binds irreversibly or reversibly to the active site cysteine residue (Cys145) of the protease, forming a covalent bond that inhibits the enzyme's catalytic activity. By inhibiting 3CLpro, the compound prevents the processing of viral polyproteins and blocks viral replication.
|
|---|---|
| ln Vitro |
In vitro activity of SARS-CoV-2 3CLpro-IN-20 is characterized by its potent inhibition of SARS-CoV-2 3CLpro. In enzyme inhibition assays, the compound has an IC50 of 0.43 μM and a Ki value of approximately 0.33 μM. These values indicate that the compound is a potent inhibitor of the protease, with nanomolar to sub-micromolar potency. The covalent binding mechanism contributes to the compound's potency, as it allows for efficient inhibition even at relatively low concentrations. In cell-based antiviral assays, the compound is expected to inhibit SARS-CoV-2 replication in infected cells, with EC50 values likely in the same range as the biochemical IC50. The compound may also show activity against other coronaviruses that have similar 3CL proteases due to the conservation of the active site. The compound's selectivity for the viral protease over human proteases is an important consideration for its therapeutic potential, though specific selectivity data are not provided in the available literature.
|
| ln Vivo |
In vivo activity of SARS-CoV-2 3CLpro-IN-20 has not been extensively reported in the available literature, as the compound is primarily a research tool for protease targeting studies. However, based on its in vitro potency as a 3CLpro inhibitor, the compound would be expected to show antiviral efficacy in animal models of SARS-CoV-2 infection if it has suitable pharmacokinetic properties. In mouse models of SARS-CoV-2 infection (e.g., hACE2 transgenic mice or mouse-adapted SARS-CoV-2), administration of the compound at appropriate doses would be expected to reduce viral loads in the lungs, attenuate lung pathology, and improve survival. The covalent binding mechanism may provide prolonged target engagement and potentially allow for less frequent dosing. However, the compound's in vivo efficacy would depend on its pharmacokinetic properties, particularly its oral bioavailability, tissue distribution, and metabolic stability. Comprehensive in vivo studies would be needed to evaluate the compound's therapeutic potential.
|
| Enzyme Assay |
For in vitro 3CLpro inhibition assays with SARS-CoV-2 3CLpro-IN-20, the following protocol is used: recombinant SARS-CoV-2 3CLpro is expressed in E. coli and purified by affinity chromatography. The protease activity is measured using a fluorogenic substrate, typically a peptide substrate (e.g., Dabcyl-KTSAVLQSGFRKME-Edans) that is cleaved by the protease to produce a fluorescence signal. The assay is performed in 50 mM Tris-HCl buffer (pH 7.3) containing 1 mM EDTA and 2 mM DTT at 25-37°C. The test compound is dissolved in DMSO and serially diluted in assay buffer to final concentrations ranging from 0.001 to 100 μM. The enzyme (10-50 nM) is pre-incubated with the compound for 10-30 minutes to allow for covalent binding. The substrate (10-50 μM) is then added, and the fluorescence (excitation 320 nm, emission 420 nm) is monitored continuously for 10-60 minutes. The initial velocity of the reaction is calculated from the linear portion of the progress curve. IC50 values are calculated from dose-response curves using nonlinear regression. The Ki value is determined using the Morrison equation for tight-binding inhibitors or by varying substrate and inhibitor concentrations.
|
| Cell Assay |
For in vitro cell-based antiviral assays with SARS-CoV-2 3CLpro-IN-20, the following typical protocol is used: Vero E6 or A549-ACE2 cells are cultured in DMEM with 10% FBS and antibiotics at 37°C in 5% CO₂. Cells are seeded in 96-well plates at 2-4 × 10⁴ cells per well and allowed to adhere overnight. The test compound is dissolved in DMSO and serially diluted in culture medium to final concentrations ranging from 0.01 to 100 μM. SARS-CoV-2 (e.g., isolate USA-WA1/2020) is diluted to 100 TCID₅₀ and added to the cells along with the compound. After 1-2 hours of adsorption, the virus-compound mixture is removed and replaced with fresh medium containing the compound at the same concentrations. Cells are incubated for 48-72 hours, and viral replication is assessed by measuring viral RNA by qRT-PCR, by immunofluorescence staining for viral nucleocapsid protein, or by cytopathic effect (CPE) reduction assay. The EC50 (effective concentration for 50% inhibition of viral replication) is calculated from dose-response curves. Cytotoxicity is assessed in parallel using the MTT assay to calculate CC50 and selectivity index (SI = CC50/EC50). For time-of-addition studies, the compound is added at various time points relative to virus infection to determine the stage of the viral life cycle that is inhibited.
|
| Animal Protocol |
For in vivo animal studies with SARS-CoV-2 3CLpro inhibitors, the following general protocol is used: K18-hACE2 transgenic mice (6-8 weeks old, female) are infected intranasally with 10⁴-10⁵ PFU of SARS-CoV-2. The test compound is formulated in a suitable vehicle (e.g., 10% DMSO, 40% PEG400, and 50% saline or 0.5% methylcellulose) and administered orally or intraperitoneally at doses of 10, 30, and 100 mg/kg, typically twice daily for 3-5 days. Control groups receive vehicle only. Body weight is monitored daily, and clinical scores (based on appearance, activity, respiration) are recorded. At various time points post-infection, mice are euthanized, and lungs are collected for viral load determination by plaque assay or qRT-PCR. Lung tissue is also processed for histopathological examination (H&E staining) to assess inflammation, edema, and lung injury. Cytokine and chemokine levels in lung homogenates are measured by ELISA or multiplex assay. For pharmacokinetic studies, blood and lung tissue samples are collected at various time points after dosing, and compound concentrations are analyzed by LC-MS/MS.
|
| ADME/Pharmacokinetics |
SARS-CoV-2 3CLpro-IN-20 (CAS# 878985-00-3) is a covalent SARS-CoV-2 3CLpro inhibitor (Compound 5g) with an IC50 of 0.43 μM and a Ki of approximately 0.33 μM. It has a molecular formula of C19H12BrNO2 and a molecular weight of 366.21 g/mol. It is used for protease targeting research to study viral replication and maturation. Future research could focus on optimizing the compound's pharmacokinetic properties for in vivo use, evaluating its efficacy in animal models of SARS-CoV-2 infection, investigating its activity against other coronaviruses, and developing it as a potential therapeutic for COVID-19 and other coronavirus diseases.
|
| Molecular Formula |
C19H12BRNO2
|
|---|---|
| Molecular Weight |
366.208084106445
|
| Exact Mass |
365.005
|
| CAS # |
878985-00-3
|
| PubChem CID |
4035834
|
| Appearance |
Orange to red solid powder
|
| LogP |
4.3
|
| Hydrogen Bond Donor Count |
0
|
| Hydrogen Bond Acceptor Count |
2
|
| Rotatable Bond Count |
2
|
| Heavy Atom Count |
23
|
| Complexity |
494
|
| Defined Atom Stereocenter Count |
0
|
| SMILES |
N1(CC2=CC=C3C(=C2)C=CC=C3)C2=C(C=C(Br)C=C2)C(=O)C1=O
|
| InChi Key |
VTPWMSNOJBWKQC-UHFFFAOYSA-N
|
| InChi Code |
InChI=1S/C19H12BrNO2/c20-15-7-8-17-16(10-15)18(22)19(23)21(17)11-12-5-6-13-3-1-2-4-14(13)9-12/h1-10H,11H2
|
| Chemical Name |
5-bromo-1-(naphthalen-2-ylmethyl)indole-2,3-dione
|
| Synonyms |
5-bromo-1-(naphthalen-2-ylmethyl)-1H-indole-2,3-dione
|
| HS Tariff Code |
2934.99.9001
|
| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Solubility (In Vitro) |
DMSO : ~4.76 mg/mL (~13.00 mM)
|
|---|---|
| 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 | 2.7307 mL | 13.6534 mL | 27.3067 mL | |
| 5 mM | 0.5461 mL | 2.7307 mL | 5.4613 mL | |
| 10 mM | 0.2731 mL | 1.3653 mL | 2.7307 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.
Products are for research use only; We do not sell to patients
Copyright 2020 InvivoChem LLC | All Rights Reserved
