1. NF-κB
  2. Keap1-Nrf2

KI696 

Cat. No.: HY-101140 Purity: 99.25%
Handling Instructions

KI696 is a high affinity probe that disrupts the Keap1/NRF2 interaction.

For research use only. We do not sell to patients.

KI696 Chemical Structure

KI696 Chemical Structure

CAS No. : 1799974-70-1

Size Price Stock Quantity
10 mM * 1 mL in DMSO USD 715 In-stock
Estimated Time of Arrival: December 31
1 mg USD 190 In-stock
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5 mg USD 590 In-stock
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10 mg USD 990 In-stock
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25 mg USD 1990 In-stock
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50 mg USD 3500 In-stock
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100 mg USD 5500 In-stock
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Customer Review

  • Biological Activity

  • Protocol

  • Technical Information

  • Purity & Documentation

  • References

Description

KI696 is a high affinity probe that disrupts the Keap1/NRF2 interaction.

IC50 & Target

Target: Keap1-NRF2[1]

In Vitro

KI696 (compound 7) exhibits very high affinity for the KEAP1 Kelch domain (ITC Kd=1.3 nM with the exception of the organic anion transporting polypeptide 1B1 (OATP1B1) (IC50=2.5 µM), the bile salt export pump BSEP (IC50=4.0 µM), and the phosphodiesterase PDE3A (IC50=10 µM), no significant cross-reactivity is observed. No cytotoxicity is observed towards BEAS-2B cells with KI696 at concentrations up to 10 µM. KI696 increases NRF2 Nuclear Translocation in Normal Human Bronchial Epithelial cells. KI696 increases mRNA expression of the NRF2-dependent genes NQO1 and GCLM in NHBE cells transfected with the non-targeting siRNA, while NRF2 gene silencing significantly decreases compound activity. KI696 increases NQO1 Activity in an NRF2-Dependent Manner. Treatment with tBHP clearly has a detrimental effect on cell health and appearance while pre-treatment of cells with 1 µM KI696 before the exposure to tBHP maintained cell morphology consistent with the DMSO control. KI696 Induces the Expression of NRF2-Regulated Genes in COPD patient-derived bronchial epithelial cells[1].

In Vivo

KI696 induces the expression of each of the Nqo1, Ho-1, Txnrd1, Srxn1, Gsta3, Gclc genes in a dose-dependent manner, with maximum increases over vehicle controls of 37-(Nqo1), 17-(Ho-1), 9-(Txnrd1), 28-(Srxn1), 15-(Gsta3) and 13-fold (Gclc) occurring at the 50 µmol/kg dose. EC50 values are 44.0, 25.7, 42.6, 33.8, 28.4, and 44.1 µmol/kg, respectively, giving an average EC50 value of 36.4±3.4 µmol/kg. KI696 attenuates ozone-Induced pulmonary inflammation. KI696 restores ozone-induced depletion of lung GSH levels. KI696 is administered to rats at 10, 35 and 50 µmol/kg by IV infusion, resulting in steady state compound concentrations in the blood of 407±44 nM, 946±50 nM and 1437±186 nM, respectively, over the 6 hour infusion period. Exposure to ozone 24 hours post-dose produces a significant depletion in lung levels of the anti-oxidant molecule, GSH, which is restored by KI696 in a dose-dependent manner[1].

Solvent & Solubility
In Vitro: 

10 mM in DMSO

Preparing
Stock Solutions
Concentration Solvent Mass 1 mg 5 mg 10 mg
1 mM 1.8161 mL 9.0805 mL 18.1610 mL
5 mM 0.3632 mL 1.8161 mL 3.6322 mL
10 mM 0.1816 mL 0.9081 mL 1.8161 mL
*Please refer to the solubility information to select the appropriate solvent.
References
Kinase Assay
[1]

Inhibition of the Kelch domain-NRF2 interaction is determined using a fluorescence polarisation-based competition assay in a black 384-well microplate. Each well contained 2 nM 5’-TAMRA-NRF2 peptide (AFFAQLQLDEETGEFL) and 7 nM human KEAP1 (residues 321-609) in 50 µL of assay buffer (50 mM Tris-HCl pH 8.0, 100 mM NaCl, 5 mM MgCl2, 2 mM 3-[(3-Cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS), 1 mM DTT, 0.005% BSA, 1% DMSO). After 1 hour at room temperature, fluorescence polarisation (excitation 485 nm/emission 520 nm) is measured using a BMG Pherastar FS plate reader. IC50 values are determined by fitting the data to a four parameter logistic fit[1].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

Cell Assay
[1]

BEAS-2B cells are plated in 384 well black clear-bottomed plates and are incubated overnight (37°C, 5% CO2). On day 2, the plates are centrifuged and 50 nL of compound (KI696) or controls are added to the cells for 48 hours. On day 4, the medium is aspirated from the plate and crude cell lysates are made by using 1X lysis buffer with Complete, Mini, EDTA-free Protease Inhibitor. After lysis, the plates are incubated for 20 minutes at room temperature and the MTT cocktail is prepared for measurement of NQO1 activity. The samples are analyzed on an Envision plate reader, reading absorbance at 570 nm for five individual readings with 10 minute intervals. Product formation is measured kinetically and the pEC50 of NQO1 specific activity induction is calculated by plotting the change in absorbance versus log [compound] followed by 4-parameter fitting[1].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

Animal Administration
[1]

Rats[1]

To examine the effect of KI696 under conditions of oxidative stress, rats are administered KI696 at 35 µmol/kg (the approximate average EC50 value for gene expression) by IV infusion over 6 hours, and after 24 hours are exposed to ozone (1 ppm for 3 hours). Fifteen minutes following the termination of ozone exposure, the numbers of total cells, neutrophils and mononuclear cells in the bronchoalveolar lavage (BAL) fluid are measured[1].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

References
Molecular Weight

550.63

Formula

C₂₈H₃₀N₄O₆S

CAS No.

1799974-70-1

SMILES

COC1=C(N2C)C(N=N2)=CC([[email protected]](C3=CC(CN(C[[email protected]]4C)S(=O)(C5=CC=CC=C5O4)=O)=C(C)C=C3)CC(O)=O)=C1

Storage
Powder -20°C 3 years
  4°C 2 years
In solvent -80°C 6 months
  -20°C 1 month
Shipping

Room temperature in continental US; may vary elsewhere

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Product Name:
KI696
Cat. No.:
HY-101140
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KI696

Cat. No.: HY-101140