Pelitinib (EKB-569)

Catalog No.S1392 Batch:S139202

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Technical Data

Formula

C24H23ClFN5O2
Molecular Weight 467.92 CAS No. 257933-82-7
Solubility (25°C)* In vitro DMSO 13 mg/mL (27.78 mM)
Water Insoluble
Ethanol Insoluble
* <1 mg/ml means slightly soluble or insoluble.
* Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations.
* Room temperature shipping (Stability testing shows this product can be shipped without any cooling measures.)

Preparing Stock Solutions

Biological Activity

Description Pelitinib (EKB-569) is a potent irreversible EGFR inhibitor with IC50 of 38.5 nM. Pelitinib (EKB-569) also slightly inhibits Src, MEK/ERK and ErbB2 with IC50s of 282 nM, 800 nM and 1255 nM, respectively. Phase2.
Targets
EGFR [1] Src [1] MEK/ERK [1] ErbB2 [1] Raf [1]
38.5 nM 282 nM 800 nM 1.255 μM 3.353 μM
In vitro Pelitinib displays much higher inhibitory activity against EGFR, compared with the closely related c-erbB-2, as well as other kinases such as Src, Cdk4, c-Met, Raf, and MEK/ERK, with IC50 ranging from 282 nM for Src to >20 μM for Cdk4. Consistently, Pelitinib treatment significantly inhibits the autophosphorylation of EGFR but not c-Met in A431 cells. [1] Pelitinib potently inhibits the proliferation of normal human keratinocytes (NHEK), as well as A431 and MDA-468 tumor cells with IC50 of 61 nM, 125 nM, and 260 nM, respectively, while displaying little activity against MCF-7 cells with IC50 of 3.6 μM. Pelitinib inhibits EGF-induced phosphorylation of EGFR in A431 and NHEK cells with IC50 of 20-80 nM, as well as the phosphorylation of STAT3 with IC50 of 30-70 nM. Pelitinib at 75-500 nM also specifically inhibits the activation of AKT and ERK1/2, without affecting NF-κB pathway. In NHEK cells, Pelitinib also potently inhibits TGF-α mediated EGFR activation with IC50 of 56 nM, as well as activation of STAT3 and ERK1/2 with IC50 of 60 nM and 62 nM, respectively. [2]
In vivo A single oral dose of 10 mg/kg Pelitinib potently inhibits the EGFR phosphorylation in A431 xenografts with over-expressed EGFR, by 90% within 1 hour, and by >50% after 24 hours. Administration of Pelitinib at 20 mg/kg/day inhibits tumorigenesis in APCMin/+ mice by 87%, equivalent to the effect of used with 2 times doses of EKI-785 (40 mg/kg/day), consistent with greater in vivo potency. [1] Pelitinib selectively inhibits EGFR signaling in airway epithelial cells in vivo. In the mouse model of airway epithelial remodeling that is inducible by viral infection and features a delayed but permanent switch to goblet cell metaplasia, Pelitinib treatment at 20 mg/kg/day corrects all 3 aspects of epithelial remodeling, by completely blocking the increase of ciliated cells and decrease of Clara cells, and significantly inhibiting the metaplasia of goblet cells. [3]
Features An improved version of EKI-785.

Protocol (from reference)

Kinase Assay:

[1]
  • Autophosphorylation of EGFR in cells

    For experiments using cells in culture, A431 cells are treated with various concentrations of Pelitinib for 2.75 hours before co-incubation with 100 ng/mL EGF for 0.25 hour. Cells are washed twice with cold phosphate-buffered saline (PBS) before adding to lysis buffer (10 mM Tris, pH 7.5, 5 mM ethylenediamine tetra-acetic acid (EDTA), 150 mM NaCl, 1% Triton X-100, 1% Sodium deoxycholate, 0.1 % SDS, 1 mM PMSF, 10 mg/mL pepstatin A, 10 mg/mL leupeptin, 20 KIU/mL aprotinin, 2 mM sodium orthovanadate, and 100 mM sodium fluoride) for 20 minutes on ice, before immunoprecipitation and SDS-PAGE-immunoblotting. For immunoprecipitation, cultured cells are placed in cold lysis buffer and immediately homogenized on ice with a polytron with several pulses. The homogenate is first centrifuged at 2500 rpm (20 minutes, 4 °C) and then again at 14,000 rpm in a microcentrifuge (10 minutes, 4 °C). Supernatants (1000 μg protein) are incubated for 2 hours at 4 °C with 15 mL of EGFR polyclonal antibody. After 2 hours, 50 μL of protein G plus/protein A agarose beads is added and incubated with constant rotation for 2 hours at 4 °C. After washing with lysis buffer, beads are boiled for 2 minutes in Laemmli sample buffer. Proteins are then resolved by SDS-PAGE, transferred to immobilon membrane and probed overnight with an anti-phosphotyrosine antibody conjugated with horseradish peroxidase (HRP). Membranes are developed using the ECL reagent. Total EGFR protein is determined by stripping membranes and re-probing with receptor-specific antibodies. Quantitation of bands is done by densitometry, using ImageQuant software with a Molecular Dynamics laser transmittance scanner.
Cell Assay:

[2]
  • Cell lines

    NHEK, A431, MCF-7, and MDA-468

  • Concentrations

    Dissolved in DMSO, final concentrations ~10 μM

  • Incubation Time

    5 days

  • Method

    Cells are seeded in 96-well dishes, and after 2 hours, Pelitinib is added and incubated for 5 days. After incubation, the medium is removed from each well and fresh medium (150 μL) + 1 mg/mL MTT solution (50 μL) is added. After incubation for 2 hours at 37 °C, the medium is replaced with 150 μL DMSO, and absorbance at 540 nm in each well is determined. The IC50 is calculated by linear regression of the data.
Animal Study:

[1]
  • Animal Models

    Athymic nu/nu female mice bearing subcutaneous A431 tumors, or APCMin/+ male mice, a murine model of human familial adenomatous polyposis (FAP)

  • Dosages

    10, or 20 mg/kg/day

  • Administration

    Oral gavage

Customer Product Validation

Data from [Data independently produced by Infect Immun, 2014, 82(3), 1243-55]

Data from [J Immunol, 2012, 188, 4581-4589]

Data from [Data independently produced by , , Korean J Parasitol, 2017, 55(5):491-503]

Selleck's Pelitinib (EKB-569) has been cited by 11 publications

Integrative analysis of drug response and clinical outcome in acute myeloid leukemia [ Cancer Cell, 2022, S1535-6108(22)00312-9] PubMed: 35868306
EGFR Inhibition Potentiates FGFR Inhibitor Therapy and Overcomes Resistance in FGFR2 Fusion-Positive Cholangiocarcinoma [ Cancer Discov, 2022, 12(5):1378-1395] PubMed: 35420673
The multi-kinase inhibitor afatinib serves as a novel candidate for the treatment of human uveal melanoma [ Cell Oncol (Dordr), 2022, 45(4):601-619] PubMed: 35781872
Biomarker LEPRE1 induces pelitinib-specific drug responsiveness by regulating ABCG2 expression and tumor transition states in human leukemia and lung cancer [ Sci Rep, 2022, 12(1):2928] PubMed: 35190588
A Genome-Scale CRISPR Screen Identifies the ERBB and mTOR Signalling Networks as Key Determinants of Response to PI3K Inhibition in Pancreatic Cancer [ Mol Cancer Ther, 2020, 5;molcanther.1131.2019] PubMed: 32371585
Mapping phospho-catalytic dependencies of therapy-resistant tumours reveals actionable vulnerabilities. [ Nat Cell Biol, 2019, 21(6):778-790] PubMed: 31160710
Suppressors for Human Epidermal Growth Factor Receptor 2/4 (HER2/4): A New Family of Anti-Toxoplasmic Agents in ARPE-19 Cells [Kim YH Korean J Parasitol, 2017, 55(5):491-503] PubMed: 29103264
Sensitivity of Melanoma Cells to EGFR and FGFR Activation but Not Inhibition is Influenced by Oncogenic BRAF and NRAS Mutations. [Garay T, et al. Pathol Oncol Res, 2015, 10.1007/s12253-015-9916-9] PubMed: 25749811
Selective inhibition of human solute carrier transporters by multikinase inhibitors [Johnston RA, et al. Drug Metab Dispos, 2014, 42(11):1851-7] PubMed: 25165131
Role of epidermal growth factor receptor signaling in the interaction of Neisseria meningitidis with endothelial cells [Slanina H et al. Infect Immun, 2014, 82(3):1243-55] PubMed: 24379285

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