Keap1-Nrf2-IN-3 , CAS No.2095066-47-8

CAS: 2095066-47-8 Cat. No.: K1447637 Summenformel: C₃₁H₃₄N₆O₃ Molekulargewicht: 538.64

Storage

Store at -20°C

Shipped In

Ice chest + Ice pads

Übersicht

Keap1-Nrf2-IN-3 is a KEAP1:NRF2 protein−protein interaction inhibitor, and with a K d value of 2.5 nM for KEAP1 protein.

Specifications

Storage

Store at -20°C

Verschickt in

Ice chest + Ice pads

Dieses Produkt erfordert Kühlkettenversand. Grundversand und andere Economy-Optionen sind nicht verfügbar.

Aktionsart

INHIBITOR

Namen und Kennungen

Molekulargewicht

538.64

Application Protocols

No item-specific, validated application protocols are provided in the Product Data for Keap1-Nrf2-IN-3. The following are general, literature-based examples for small-molecule Keap1–Nrf2 probes; adapt to your assay and verify empirically.

Example 1: FP displacement assay (general)

• Prepare 10 mM DMSO stock; serially dilute in assay buffer (HEPES 25 mM, pH 7.4; NaCl 150 mM; 0.01% Tween‑20; 0.1 mg/mL BSA).
• Mix Keap1 Kelch protein (10–50 nM) with fluorescent Nrf2 tracer at its Kd; add compound dilutions (final DMSO ≤1%).
• Incubate 30 min at RT; read polarization; fit IC50, convert to Ki with appropriate model.

Example 2: ARE-luciferase reporter (general)

• Treat cells harboring ARE-luc reporter with compound across 0.03–30 µM for 6–24 h (DMSO constant 0.5–1%).
• Measure luciferase activity; assess cytotoxicity in parallel; include Keap1 overexpression/knockdown controls.

Example 3: SPR binding (general)

• Immobilize Keap1 on CM5; inject compound series (0.1–100 µM) with matched DMSO content.
• Perform solvent correction; analyze steady-state KD.

Notes:

• Verify compound solubility/aggregation; include 0.01% detergent where compatible.
• Confirm identity/purity by LC-MS/HPLC before critical experiments.

Biological Roles

Context (literature, pathway-level; not specific to this item’s unprovided structure):

• The Keap1–Nrf2 axis regulates cellular defense against oxidative and electrophilic stress. Keap1 (Kelch-like ECH-associated protein 1) binds Nrf2, promoting its ubiquitination and proteasomal degradation under basal conditions. Disruption of this interaction stabilizes Nrf2, allowing nuclear translocation and transcriptional activation of antioxidant response element (ARE)-driven genes.

Role of small-molecule Keap1–Nrf2 modulators (general):

• Chemical ligands that engage the Kelch domain of Keap1 can competitively block Nrf2 binding. Such probes are used to interrogate redox biology, cytoprotective gene regulation, and to dissect upstream/downstream signaling nodes.
• In cells, on-target modulation is typically confirmed by measuring Nrf2 protein levels, nuclear localization, and induction of canonical gene products (e.g., NQO1, GCLM) using non-cytotoxic concentrations (experimental conditions are compound- and cell-line-dependent; verify empirically).

Assay modalities:

• Biophysical: Fluorescence polarization, SPR, MST, and ITC to quantify binding.
• Biochemical: Competitive displacement of labeled Nrf2 peptides from Keap1.
• Cellular: ARE-luciferase reporters, qPCR/Western blot for Nrf2 targets, with genetic controls to confirm specificity.

Caveats:

• Some chemotypes can be electrophilic or redox-active; include counterscreens to exclude nonspecific activation via oxidative stress rather than direct Keap1 engagement.

Buffer Applications

This product is not a classical buffering agent. However, appropriate buffer systems are critical for reliable biochemical and biophysical assays involving Keap1–Nrf2 pathway probes.

General buffer guidance (literature best practices):

• FP/fluorescence assays: 20–50 mM HEPES, pH 7.4–7.5; 100–150 mM NaCl; 0.005–0.02% Tween‑20; 0.5–1 mM TCEP or DTT as needed to maintain protein stability; 0.1–1 mg/mL BSA to reduce nonspecific binding. Keep DMSO constant across wells (e.g., 1%).
• SPR: 10 mM HEPES, pH 7.4; 150 mM NaCl; 0.05% surfactant (e.g., P20) as recommended by instrument manufacturer; include 1–2% DMSO for solvent correction if analyte is in DMSO.
• ITC: 20–50 mM HEPES or phosphate, pH 7.2–7.4; match buffer in both syringe and cell; keep DMSO content identical to avoid heat-of-dilution artifacts.

Practical notes:

• Filter (0.22 µm) and degas buffers for sensitive optical or calorimetric readouts.
• Validate reducing agent compatibility with both protein and compound; some small molecules are unstable in strong reducing conditions.
• Avoid phosphate buffers in certain fluorescence assays where background or complexation may interfere; empirically assess.

Green Alternatives

Because Keap1-Nrf2-IN-3 is a specialized research probe rather than a process solvent or bulk reagent, green chemistry considerations focus on solvent choice and assay format rather than substituting the active itself.

Greener practice suggestions (general):

• Prefer aqueous assay buffers with minimal DMSO (≤1% v/v) where feasible.
• Use microvolume, high-density assay plates to reduce solvent consumption and waste.
• Choose lower-toxicity co-solvents (ethanol) over acetonitrile when assay-compatible.
• Implement plate sealing to minimize evaporative losses and VOC exposure.

Small comparison (general solvent context):

| Aspect | DMSO | Ethanol | |---|---|---| | Solubilizing power | Very high | Moderate | | Assay tolerance | Commonly ≤1–2% | Commonly ≤1% | | Toxicity/biodegradability | Moderate concern; not readily biodegradable | Lower acute toxicity; biodegradable | | Protein stability impact | Generally compatible; validate | Often compatible; validate |

Additional considerations:

• Consolidate deliveries and store at −20°C to extend shelf life and reduce waste from expired materials (item-specific storage).
• Adopt green analytical methods (UPLC, lower-flow LC-MS) for QC to reduce solvent use.

Note: No greener “substitute molecule” is proposed here because the compound’s function is target-specific in discovery research.

Pharmaceutical Uses

No pharmacopeial or excipient status is provided for this item. This product is supplied strictly for research use only and is not intended for human or veterinary use.

General context (non-clinical, discovery-stage only):

• Small-molecule Keap1–Nrf2 ligands are commonly employed as chemical probes in target validation, hit-to-lead optimization, and ADME/tox profiling workflows within discovery research. Any data generated inform structure–activity relationships and mechanism but are not indicative of suitability for therapeutic administration.
• Formulation for in vivo studies, if attempted in preclinical research settings, typically requires bespoke vehicles (e.g., PEG/solvent/surfactant systems) and rigorous characterization of stability, solubility, and exposure; such procedures are outside the scope of this product listing and must be developed and validated by the end user.

Compliance note:

• Do not use in diagnostic procedures. Not GMP. No claims are made regarding clinical safety, efficacy, or regulatory status.
• For translational or GLP/GMP needs, source material manufactured under appropriate quality systems and obtain full regulatory documentation.

Physical Properties

Item-specific physico-chemical data are not provided in the Product Data. Refer to the product CoA/Spec Sheet and SDS for authoritative specifications.

Item-specific values:

• Appearance: Not specified for this item; refer to CoA/Spec Sheet.
• Molecular Weight: Not specified for this item; refer to CoA/Spec Sheet.
• Molecular Formula: Not specified for this item; refer to CoA/Spec Sheet.
• Melting/Boiling Point: Not specified for this item; refer to CoA/Spec Sheet.
• Density, Refractive Index, pKa, logP/logD: Not specified for this item; refer to CoA/Spec Sheet.
• Solubility: Not specified for this item; refer to CoA/Spec Sheet.

General expectations for small-molecule Keap1–Nrf2 probes (literature, not specific to this item):

• Many reported inhibitors are solids at ambient temperature, with moderate molecular weight (ca. 300–600 Da) and polar surface area sufficient for aqueous assay compatibility via DMSO stocks diluted into buffer.
• Stock solutions are commonly prepared in DMSO (e.g., 10–50 mM) and diluted into aqueous assay buffers to reach sub-micromolar to low-micromolar working concentrations (literature practice).

Notes for practical use (general):

• If solubility in pure aqueous buffer is limited, use co-solvents (DMSO final ≤1–2% v/v) and include nonionic surfactant (e.g., 0.01% Tween-20) to reduce aggregation artifacts in biophysical assays (literature guidance).
• Filter or centrifuge working solutions to remove particulates before sensitive readouts (FP, SPR, ITC).

Quality and Grades

Item-specific quality information:

• Grade/Purity: Not specified for this item; refer to CoA/Spec Sheet.

How to interpret grades (general guidance for small-molecule research probes):

• Research-grade: Suitable for discovery biology, target validation, and mechanistic studies. Purity is typically verified by HPLC/UPLC and identity by MS/NMR. Low levels of residual solvents or inorganic salts may be present within accepted research limits (consult CoA).
• Screening-library grade: Emphasizes identity and chromatographic purity for high-throughput screening (HTS) and biochemical/biophysical assays. UV trace and impurity profiles are often provided upon request.

What to look for on the CoA/Spec Sheet:

• Purity method and threshold (e.g., HPLC area% at a specified wavelength); residual solvent profile if available.
• Identity confirmation (MS [M+H]+, 1H NMR) and water content (Karl Fischer) where applicable.
• Batch-specific storage advice (e.g., hygroscopicity, light sensitivity) and any stabilizers used (if any; none specified for this item).

Practical tips:

• For quantitative biology, confirm purity by orthogonal methods (e.g., UPLC-UV and qNMR) when needed.
• If working near solubility limits, assess for colloidal aggregation (dynamic light scattering or detergent-sensitivity controls) to ensure clean SAR/assay data.

Reaction and Applications

Chemical reaction applications: This product is a biological small‑molecule probe rather than a synthetic reagent; it is not typically used as a substrate, catalyst, or solvent in chemical transformations. See Biological Roles, Target Specificity, and Application Protocols for relevant use cases.

Biological/assay applications (general, literature-based for Keap1–Nrf2 pathway tools; not specific to this item):

• Probe for the Keap1–Nrf2 protein–protein interaction: Used to study ligand engagement of the Keap1 Kelch domain, often by fluorescence polarization (FP), surface plasmon resonance (SPR), ITC, or AlphaScreen/HTRF competitive assays.
• Cellular pathway studies: Employed as a chemical tool to modulate Nrf2 stabilization by interfering with Keap1 binding, enabling downstream readouts of antioxidant response element (ARE) reporter activity. Assay conditions and potencies vary by compound; verify with primary literature and internal controls.
• Structural biology: Can support fragment growth/lead optimization campaigns and co-crystallography or cryo-EM of Keap1–ligand complexes (where solubility and stability permit).

Good assay practice:

• Include counter-screens (e.g., redox cycling, general reactivity, and PAINS filters) and detergent controls to rule out assay artifacts.
• Titrate DMSO-matched vehicle controls; monitor protein stability in the presence of the compound.
• Confirm on-target activity via orthogonal assays (e.g., FP vs. SPR) and, in cells, by genetic controls (Keap1 KO/overexpression) to validate mechanism of action.

Reaction Conditions

Not applicable in the classical synthetic sense: Keap1-Nrf2-IN-3 is supplied as a target-engagement probe, not as a reagent for performing chemical reactions.

Assay-condition guidance (general, literature-based for Keap1–Nrf2 PPI studies; not specific to this item):

• FP displacement assay: Typical protein (Keap1 Kelch domain) at 5–100 nM; fluorescent Nrf2 peptide tracer at its Kd; incubate 15–60 min at room temperature in HEPES buffer with 0.01% Tween‑20; read polarization and fit IC50/Ki using appropriate models.
• SPR binding: Immobilize Keap1 on CM5 or similar; inject compound in 2‑fold dilutions (e.g., 0.1–100 µM) with constant DMSO; use multiple reference surfaces and DMSO calibration; analyze with steady-state or kinetic models.
• Cellular ARE-luciferase: Seed cells to 70–80% confluence; treat with compound dilutions (e.g., 0.03–30 µM) at constant DMSO (≤0.5–1%); measure reporter after 6–24 h; include cytotoxicity counterscreen and Nrf2/Keap1 genetic controls.

Always optimize conditions for your specific protein construct, tracer, cell line, and detection platform. Validate compound integrity (LC-MS) pre- and post-assay where possible.

Safety and Handling

Safety overview for Keap1-Nrf2-IN-3 (research chemical):

Item-specific hazard data from Product Data:

• GHS Classification: Not specified for this item; refer to SDS.
• Signal Word: Not specified for this item; refer to SDS.
• H-Statements: Not specified for this item; refer to SDS.
• Pictograms: Not specified for this item; refer to SDS.

General laboratory precautions (supplemental, not a substitute for SDS):

• Handle in a fume hood; avoid inhalation of dusts/aerosols and contact with skin/eyes.
• Wear appropriate PPE: lab coat, safety glasses, and suitable chemically resistant gloves (e.g., nitrile). For weighing powders, consider a particulate mask or balance enclosure.
• Avoid ingestion; do not pipette by mouth. Wash hands and contaminated skin thoroughly after handling.
• Prevent environmental release; collect waste in appropriate organic/halogenated waste streams per institutional policy.

Storage and incompatibilities:

• Storage condition (item-specific): Store at −20°C (provided).
• Ship conditions (item-specific): Shipped in ice chest + ice pads (provided).
• Keep container tightly closed, protected from moisture and light. Allow vials to equilibrate to room temperature in a desiccator before opening to minimize condensation.
• General incompatibilities (typical for organic small molecules): strong oxidizers, strong acids/bases may cause decomposition; avoid prolonged exposure to elevated temperature.

First-aid overview (general):

• Skin/eye contact: Rinse with water for ≥15 minutes; remove contaminated clothing. Seek medical attention if irritation persists.
• Inhalation: Move to fresh air; seek medical attention if symptoms develop.
• Ingestion: Rinse mouth; seek medical attention. Provide SDS to healthcare professionals.

Solvent Selection

Item-specific solvent and solubility data are not provided; consult the CoA/Spec Sheet. The following is general guidance for small-molecule Keap1–Nrf2 pathway probes in biochemical assays.

• Primary stock solvent: DMSO is typically preferred due to broad solubilizing power and assay compatibility at low final percentages.
• Aqueous buffers: Working solutions are typically prepared by diluting DMSO stocks into assay buffers (e.g., HEPES, PBS, Tris). Aim for a final DMSO concentration ≤1–2% v/v to minimize assay interference (literature practice).
• Co-solvents: If precipitation occurs, consider small fractions of ethanol or acetonitrile; verify compatibility with the assay modality (FP, SPR, ITC, enzymatic readouts).
• Detergents: 0.005–0.02% nonionic surfactants (Tween-20 or Triton X‑100) can mitigate nonspecific adsorption and aggregation in PPI assays (literature guidance).

Comparison (general):

• DMSO: Maximum solubilization; may affect certain fluorescence readouts at >2%.
• Ethanol: More volatile; generally tolerated up to ~1% v/v in many biochemical assays.
• Acetonitrile: Useful for rapid mixing; can destabilize some proteins—validate experimentally.

Practical tips:

• Pre-wet plasticware with buffer containing surfactant to reduce compound loss to surfaces.
• Prepare serial dilutions immediately before use to limit freeze–thaw/oxidation and maintain accuracy at low micromolar to nanomolar ranges.

Storage and Reconstitution

Item-specific conditions (from Product Data):

• Storage: Store at −20°C.
• Shipping: Shipped in ice chest + ice pads.

General best practices for small-molecule research probes:

• Upon receipt, allow the sealed container to warm to room temperature before opening to prevent condensation. Reseal promptly under dry atmosphere if hygroscopic.
• Solid material: For long-term storage, keep desiccated and protected from light. If subdividing, aliquot into inert, amber vials.
• Solution stocks: Prepare concentrated stocks (e.g., 10–50 mM) in anhydrous DMSO. Dispense single-use aliquots and store at −20°C or −80°C to minimize freeze–thaw cycles.
• Freeze–thaw guidance: Avoid repeated cycles; thaw on ice, mix gently, and refreeze promptly. Discard aliquots showing precipitation or discoloration.
• Stability checks: Periodically verify integrity by LC‑MS/HPLC, especially after extended storage or multiple thaws.

Reconstitution (general):

• Solvent choice: DMSO is typically preferred. If aqueous reconstitution is required, dilute DMSO stocks into buffer with gentle mixing; maintain final DMSO ≤1–2% v/v as assay permits.
• Filtration: If particulates are observed, centrifuge briefly or filter through 0.22 μm PTFE/RC filters compatible with organic solvents.

Note: In the absence of item-specific solubility and stability data, perform a small-scale solubility/stability scout and consult the CoA/Spec Sheet for lot-specific recommendations.

Structure and Identity

Brief overview: This listing is for a small‑molecule research probe designated Keap1‑Nrf2‑IN‑3 (SKU K1447637), commonly used in studies of the Keap1–Nrf2 pathway.

• SKU: K1447637
• Product Name: Keap1-Nrf2-IN-3
• CAS: 2095066-47-8
• Category: Small molecule/compound library (per catalog path)
• Research Use: For research use only

Item-specific identifiers (from Product Data):

• Molecular Formula: Not specified for this item; refer to CoA/Spec Sheet.
• Molecular Weight: Not specified for this item; refer to CoA/Spec Sheet.
• SMILES: Not specified for this item; refer to CoA/Spec Sheet.
• InChIKey: Not specified for this item; refer to CoA/Spec Sheet.

Structural features (general, literature-level context for Keap1–Nrf2 PPI modulators):

• Many reported Keap1–Nrf2 PPI inhibitors/ligands are heteroaromatic, often bearing acidic or phenolic motifs to engage the Keap1 Kelch domain hot-spot residues (literature, not specific to this item).
• Typical functional groups may include amides, carboxylates, phenols, or sulfonamides to provide hydrogen-bond donors/acceptors and anionic character at physiological pH (literature, not specific to this item).

2D description (general, not specific to this item):

• Ligands for the Keap1 Kelch domain frequently feature a polyfunctional scaffold with an anionic/phenolic terminus, an amide linker, and a heteroaryl ring system to complement the shallow PPI pocket (literature). For exact atom connectivity of Keap1‑Nrf2‑IN‑3, consult the CoA/Spec Sheet.

Synthetic Utility

This product is provided as a finished small‑molecule probe for biological research and is not typically used as a synthetic building block.

Accordingly, classic synthetic-chemistry applications (e.g., use as a reagent, catalyst, monomer, or protecting group) are not applicable. If your intent is to derivatize or immobilize the molecule for probe development (e.g., for pull‑down or tracer synthesis), please consult the full structural information (SMILES, functional groups) on the CoA/Spec Sheet to assess feasible points of modification.

General considerations for probe derivatization (literature-level, not specific to this item):

• Attachment handles: Amides, phenols, or carboxylates (when present in a given chemotype) can serve as conjugation points to linkers for affinity resins or fluorophores; prioritize positions distal to key binding pharmacophores.
• Bioorthogonal tags: Azide/alkyne handles enable click chemistry for tracer or imaging agent assembly; ensure tag addition does not disrupt Keap1 binding.
• SAR maintenance: Maintain ionization state and hydrogen-bonding features essential for Kelch pocket recognition; verify binding post-modification by FP/SPR.

Target Specificity

Item-specific target details (antigen/clone/isotype, etc.) are not applicable because this product is a small-molecule probe, not an antibody. No additional target-binding constants or selectivity data are provided in the Product Data for this specific item.

General context (literature-level for Keap1–Nrf2 pathway tools; not specific to this item):

• Intended molecular target: Keap1 Kelch domain at the Nrf2-binding interface (ETGE/DLG motifs). Small molecules are designed to mimic critical interactions to displace Nrf2 from Keap1.
• Selectivity considerations: Evaluate off-target effects on other BTB/Kelch proteins and assess redox reactivity to exclude nonspecific activation via oxidative stress. Employ chemoproteomics or broad profiling panels if available.

Recommendation:

• For batch-specific affinity/IC50/Kd data and any off-target profiling for Keap1-Nrf2-IN-3, consult the CoA/Spec Sheet or contact Technical Support with the lot number.