PQ 401 196868-63-0

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Biological Activity


PQ 401 is an IGF-1R Inhibitor and inhibits autophosphorylation of the IGF-IR kinase domain at concentrations


PQ 401 (50 mg/Kg, 100 mg/Kg) significantly inhibits MCNeuA tumor growth in a dose-dependent manner.


IGF-IR Peptide Autophosphorylation


One microgram of constitutively active IGF-IR kinase domain peptide
isincubated +/− varying concentrations of PQ 401 in 2% DMSO in 40 mM
Tris (pH 7.4), 80μMEGTA, 0.25% 2-mercaptoethanol, 80μM Na3VO4, 10 mM MgCl2, and 2 mM MnCl2 for 20 minutes. ATP isthen added at a final concentration of 20μM.
Autophosphorylation of the kinase domain peptide isallowed to occur for
20 minutes at 22℃.


The reaction isstopped by the addition of
SDS-reducing buffer and the samples are run on SDS-PAGE. Following
transfer to nitrocellulose membrane, peptide autophosphorylation
isdetermined by Western blotting employing an antibody against
phosphotyrosine (PY20).






Method


The inhibitory effects of diaryl urea on breast cancer cell growth are
determined using a CyQuant cell proliferation assay kit. MCF-7 or MCNeuA
cells are plated in 96-well plates (5 × 103 per well) in
phenol red-free DMEM supplemented with 10% FCS. One plate isprepared for
each harvest day. Cells are allowed to adhere overnight and are then
treated with various concentrations of diaryl urea or DMSO as a vehicle
control.


Microplate cultures are harvested on days 0, 1, 2, and 3 by
inverting the microplate onto paper towels with gentle blotting to
remove growth medium without disrupting adherent cells. Each plate
iskept at −80 ℃ until the end of the experiment (day 3) when all of the
plates are thawed and assayed together.


After thawing, 200 μL of CyQuant
GR solution are added to each well and the plates are incubated in the
dark for 2 to 5 minutes. Fluorescence ismeasured with a SpectraMax
Gemini XS fluorescence microplate reader with 480-nm excitation and
520-nm emission. Proliferation index iscalculated as the percent of
nucleotide content versus control cells at day 0.

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Product Categories : Protein Tyrosine Kinase > IGF-1R Inhibitor