Home Cell Cycle Chk inhibitor CHIR-124 For research use only.

CHIR-124

CHIR-124 is a novel and potent Chk1 inhibitor with IC50 of 0.3 nM in a cell-free assay. It shows 2,000-fold selectivity against Chk2, 500- to 5,000-fold less activity against CDK2/4 and Cdc2.

CHIR-124 Chemical Structure

CHIR-124 Chemical Structure

CAS No. 405168-58-3

Purity & Quality Control

CHIR-124 Related Products

Signaling Pathway

Chk Signaling Pathways

Chk Signaling Pathway

Cell Data

Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
human MDA-MB-435 cells Cytotoxic assay Cytotoxicity against human MDA-MB-435 cells, EC50=0.08 μM 16603354
human MDA-MB-435 cells Cytotoxic assay Cytotoxicity against human MDA-MB-435 cells in presence of camptothecin 16603354
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Biological Activity

Description CHIR-124 is a novel and potent Chk1 inhibitor with IC50 of 0.3 nM in a cell-free assay. It shows 2,000-fold selectivity against Chk2, 500- to 5,000-fold less activity against CDK2/4 and Cdc2.
Targets
Chk1 [1]
(Cell-free assay)		 FLT3 [1]
(Cell-free assay)		 PDGFR [1]
(Cell-free assay)		 GSK-3 [1]
(Cell-free assay)
0.3 nM 5.8 nM 6.6 nM 23.3 nM
In vitro
In vitro

CHIR-124 is a quinolone-based small molecule that is structurally unrelated to other known inhibitors of Chk1. CHIR-124 interacts synergistically with topoisomerase poisons (e.g., Camptothecin or SN-38) in causing growth inhibition in a variety of cancer cell lines, including breast carcinoma (MDA-MB-231 and MDA-MB-435) and colon carcinoma (SW-620 and Colo205), all of which contains the mutant p53 gene. CHIR-124 abrogates the SN-38-induced S and G2-M checkpoints and potentiates apoptosis in MDA-MD-435 breast cancer cells. The abrogation of the G2-Mcheckpoint and induction of apoptosis by CHIR-124 are enhanced by the loss of p53. [1] CHIR-124 also potently targets other kinases such as PDGFR and Flt3 with IC50 of 6.6 nM and 5.8 nM, respectively. [2]
Kinase Assay Chk1 Assay
For the Chk1 assay, the kinase domain is expressed in Sf9 insect cells, and a biotinylated cdc25c peptide containing the consensus Chk1/Chk2 phosphorylation site (*)(biotin-[AHX]SGSGS*GLYRSPSMP-ENLNRPR[CONH2]) is used as the substrate. A dilution series of CHIR-124 is mixed with a kinasereaction buffer containing a final concentration of 30 mM Tris-HCl(pH 7.5), 10 mM MgCl2, 2 mM DTT, 4 mM EDTA, 25 mMβ-glycerophosphate, 5 mM MnCl2, 0.01% bovine serum albumin, 1.35 nM CHK1 kinase domain, 0.5 μM peptide substrate, and 1 AM unlabeled ATP, plus 5 nM 33Pγ-labeled ATP (specific activity = 2,000 Ci/mmol). Reactions and detection of the phosphate transfer are carried out by a radioactive method. Reactions are incubated at room temperature for 1 to 4 hours and the phosphorylated peptide captured on streptavidin-coated microtiter plates containing stop reaction buffer (25 mM EDTA [ethylenediaminetetraacetic acid], 50 mMHEPES, pH 7.5). Phosphorylated peptide is measured with the DELFIA TRF system using a Europium-labeled anti-phosphotyrosine antibody PT66. The concentration of CHIR-124 for IC50 is calculated using nonlinear regression with XL-Fit data analysis software.
Cell Research Cell lines MDA-MB-231, MDA-MB-435, SW-620, and COLO 205 cells
Concentrations 0-2350 nM, dependent on cell types
Incubation Time 48 hours
Method

MDA-MB-231, MDA-MB-435, SW-620, and COLO 205 cells in log-phase are plated into 96-well microplates. CHIR-124 is serially diluted in the presence of six different concentrations of Camptothecin or 0 nM camptothecin. Camptothecin is also serially diluted in the absence of CHIR-124. CHIR-124 is added to cells in 96-well dishes and incubated at 37 °C for 48 hours. Each treatment condition is done in triplicate. Cell proliferation is monitored by the 3-(4,5-dimethylthiazol-2-yl)-5- (3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS), inner salt assay. MTS inner salt is added to the microplates, which are incubated for another 3 hours, and absorbance at 490 nm is read on a plate reader. The concentrations of each drug in the combinations required to produce 50% inhibition are plotted to generate the isoboles. Isobologram analysis of drug interaction is based the equation of Loewe additivity (1= D A /IC50, A + DB/IC50, B), where IC50, A and IC50, B are the concentrations of drugs to result in 50% inhibition for each drug alone, and DA and DB are concentrations of each drug in the combination that yield 50% overall inhibition. A diagonal line indicating Loewe additivity is included in each graph. Data points that fall below the line indicate synergy, whereas those that fall above the line will indicate antagonism
In Vivo
In vivo

CHIR-124 potentiates the growth inhibitory effects by abrogating the G2-M checkpoint and increasing tumor apoptosis in an orthotopic breast cancer xenograft model.
Animal Research Animal Models MDA-MB-435 cells are implanted in the mammary fat pad of 8- to 10-week-old female immunodeficient mice.
Dosages 10 mg/kg or 20 mg/kg
Administration CHIR-124 is given orally four times daily × 6 on days 2 to 7 in captisol.

Chemical Information & Solubility

Molecular Weight 419.91 Formula

C23H22ClN5O
CAS No. 405168-58-3 SDF Download CHIR-124 SDF
Smiles C1CN2CCC1C(C2)NC3=C(C(=O)NC4=C3C=C(C=C4)Cl)C5=NC6=CC=CC=C6N5
Storage (From the date of receipt)

In vitro
Batch:

DMSO : Insoluble ( Moisture-absorbing DMSO reduces solubility. Please use fresh DMSO.)

Water : Insoluble

Ethanol : Insoluble


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In vivo
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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.

Handling Instructions

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