Pelitinib (EKB-569)

Catalog No.S1392

For research use only.

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.

Pelitinib (EKB-569) Chemical Structure

CAS No. 257933-82-7

Selleck's Pelitinib (EKB-569) has been cited by 8 Publications

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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.
Features An improved version of EKI-785.
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]

Cell Data
Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
A431 NY\EcGs1TnWwY4Tpc44h[XO|YYm= MWXJcohq[mm2aX;uJI9nKEWJRmKgZZV1d3Cqb4PwbI9zgWyjdHnvckBqdiCqdX3hckBCPDNzIHPlcIx{NCCLQ{WwQVAvODB6MENOwG0v NXrYT2FsRGFidHHy[4V1RSehYnzhcosoKGi{ZX[9K4h1fHC|Oj:vdJVjdWWmLn7jZokvdmyvLn7pbE5od3ZxMkC3PVc5PzFpPkKwO|k4QDdzPD;hQi=>
Sf9 MonISpVv[3Srb36gZZN{[Xl? M4DtZ|ExKG2rboO= MULJcohq[mm2aX;uJI9nKHKnY3;tZolv[W62IHj1cYFvKEirc{[teIFo\2WmIFXHSnIh[3m2b4DsZZNucWNiZH;tZYlvKCh4NEWgeI8hOTF6NjDy[ZNq\HWnczmg[ZhxemW|c3XkJIlvKGKjY4Xsc5ZqenW|IHnu[oVkfGWmIGPmPUBqdnOnY4SgZ4VtdHNiYYPz[ZN{\WRiYYOgdoVlfWO2aX;uJIlvKGG3dH;wbI9{eGixconsZZRqd25icILlbY5kfWKjdHXkJIZweiBzMDDtbY5{KG[xbHzve4VlKGK7IFHUVE1O\0OuMjDh[IRqfGmxbjDhcoQhdWWjc4Xy[YQtKEmFNUC9NE4xOzh3zszNMi=> NGHneGg9[SC2YYLn[ZQ:L1:kbHHub{chcHKnZk2nbJR1eHN8Lz;weYJu\WRwbnPibU5vdG1wbnnoModwfi9|ME[wNFE1QSd-M{C2NFAyPDl:L3G+
A431 MUnGeY5kfGmxbjDhd5NigQ>? NYXx[m45QTBibXnudy=> NYq4N5dKUW6qaXLpeIlwdiCxZjDFS2ZTKGmwIHj1cYFvKEF2M{GgZ4VtdHNiYYPz[ZN{\WRiYYOgdoVlfWO2aX;uJIlvKEWJRj3zeIlufWyjdHXkJGVITlJiYYX0c5Bpd3OyaH;yfYxifGmxbjDwdoVqdmO3YXL0[YQh\m:{IEmwJI1qdnNiZn;scI94\WRiYomgSWdHNXO2aX31cIF1cW:wIHL5JJNidmS5aXPoMWVNUVODLDDJR|UxRTBwMEO5{txONg>? NGqwOlI9[SC2YYLn[ZQ:L1:kbHHub{chcHKnZk2nbJR1eHN8Lz;weYJu\WRwbnPibU5vdG1wbnnoModwfi9|MEm3N|c{PSd-M{C5O|M4OzV:L3G+
DiFi MVvGeY5kfGmxbjDhd5NigQ>? MUCyJIhzew>? MVTJcohq[mm2aX;uJIF2fG:yaH;zdIhwenmuYYTpc44hd2ZiRVfGVkBqdiCqdX3hckBFcU[rIHPlcIx{KGGodHXyJFIhcHK|IHL5JGVNUVODLDDJR|UxRTBwMEe5OFPPxE1w NXfDSpJmRGFidHHy[4V1RSehYnzhcosoKGi{ZX[9K4h1fHC|Oj:vdJVjdWWmLn7jZokvdmyvLn7pbE5od3ZxMUe2PFk5OzZpPkG3Olg6QDN4PD;hQi=>
UCH1 M{CzUmFvfGmycn;sbYZmemG2aY\lJIF{e2G7 NH25U4o4OiCqcoO= NIrQd5VCdnSrcILvcIln\XKjdHn2[UBi[3Srdnn0fUBi\2GrboP0JIh2dWGwIGXDTFEh[2WubIOgcYVie3W{ZXSgZYZ1\XJiN{KgbJJ{KGK7IHHsZY1ieiCkbIXlJIF{e2G7LDDJR|UxRTBwMEpOwG0v M1[3W|xiKHSjcnfleF0oZ2KuYX7rK{BpemWoPTfoeJRxezpxL4D1Zo1m\C6wY3LpMo5tdS6waXiu[493NzNyOUezO|M2Lz5|MEm3N|c{PTxxYU6=
UCH2 Ml;2RY51cXC{b3zp[oVz[XSrdnWgZZN{[Xl? NWLNToxNPzJiaILz MlPVRY51cXC{b3zp[oVz[XSrdnWgZYN1cX[rdImgZYdicW6|dDDoeY1idiCXQ1iyJINmdGy|IH3lZZN2emWmIHHmeIVzKDd{IHjyd{BjgSCjbHHtZZIh[my3ZTDhd5NigSxiSVO1NF0yNjcQvF2u NV\1[5BuRGFidHHy[4V1RSehYnzhcosoKGi{ZX[9K4h1fHC|Oj:vdJVjdWWmLn7jZokvdmyvLn7pbE5od3ZxM{C5O|M4OzVpPkOwPVc{PzN3PD;hQi=>
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]

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 Research:

[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 Research:

[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

Solubility (25°C)

In vitro

Chemical Information

Molecular Weight 467.92
Formula

C24H23ClFN5O2

CAS No. 257933-82-7
Storage 3 years -20°C powder
2 years -80°C in solvent
Smiles CCOC1=C(C=C2C(=C1)N=CC(=C2NC3=CC(=C(C=C3)F)Cl)C#N)NC(=O)C=CCN(C)C

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Tech Support

Answers to questions you may have can be found in the inhibitor handling instructions. Topics include how to prepare stock solutions, how to store inhibitors, and issues that need special attention for cell-based assays and animal experiments.

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