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PF-477736 Chk inhibitor

Name: PF-477736
Brand: Selleck Chemicals
SKU: S2904
Availability: InStock
Rating: 5 (36 reviews)

Cat.No.S2904

PF-477736 (PF-736, PF-00477736) is a selective, potent and ATP-competitive Chk1 inhibitor with K_i of 0.49 nM in a cell-free assay and also inhibits VEGFR2, Aurora-A, FGFR3, Flt3, Fms (CSF1R), Ret and Yes. It shows ~100-fold selectivity for Chk1 than Chk2.

Chemical Structure

Molecular Weight: 419.48

Jump to Mechanism of Action Cell Culture & Working Concentration Solubility Chemical Information, Storage & Stability Specificity

Quality Control

Batch: Purity: 99.80%

99.80

Related Targets	DNA/RNA Synthesis CDK Microtubule Associated Ras HSP (HSP90) PD-1/PD-L1 Aurora Kinase Casein Kinase Rho Serine/threonin kinase eIF ROCK Myc Kinesin PLK DHFR PAK PERK DYRK RUNX PP2A PFKFB AP-1 MPS1 Wee1 p21 BMI-1 LIM kinase Nur77 APC
Related Products	AZD7762 Rabusertib (LY2603618) MK-8776 (SCH 900776) CHIR-124 Prexasertib (LY2606368) Dihydrochloride BML-277 (Chk2 Inhibitor II) Prexasertib (LY2606368) CCT245737 (SRA737) GDC-0575 BBI-355 Mad1 Antibody [M18F4] Phospho-Chk1 (Ser345) Antibody [P19M4] CCT241533 hydrochloride SAR-020106 Chk2 Antibody [K6G1] Phospho-Chk1 (Ser317) Antibody [N9E3] Phospho-Chk2 (Thr68) Antibody [L12K12] Chk2 Antibody [C4M24] BBI-2779 14-3-3 γ Antibody [J16F21] LY2880070 CDK5RAP3 Antibody [K12N22]

Cell Culture, Treatment & Working Concentration

Cell Lines	Assay Type	Activity Description	PMID
TC32	qHTS assay	qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for TC32 cells	29435139
U-2 OS	qHTS assay	qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for U-2 OS cells	29435139
A673	qHTS assay	qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for A673 cells	29435139
Saos-2	qHTS assay	qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for Saos-2 cells	29435139
RD	qHTS assay	qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for RD cells	29435139
SK-N-SH	qHTS assay	qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for SK-N-SH cells	29435139
OHS-50	qHTS assay	qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for OHS-50 cells	29435139
SJ-GBM2	qHTS assay	qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for SJ-GBM2 cells	29435139
SK-N-MC	qHTS assay	qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for SK-N-MC cells	29435139
NB-EBc1	qHTS assay	qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for NB-EBc1 cells	29435139
LAN-5	qHTS assay	qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for LAN-5 cells	29435139
Click to View More Cell Line Experimental Data

Chemical Information, Storage & Stability

Molecular Weight	419.48	Formula	C₂₂H₂₅N₇O₂	Storage (From the date of receipt)	3 years-20°Cpowder
CAS No.	952021-60-2	Download SDF		Storage of Stock Solutions	1 year-80°Cin solvent 1 month-20°Cin solvent
Synonyms	PF-736, PF-00477736			Smiles	CN1C=C(C=N1)C2=C3C=NNC(=O)C4=C3C(=CC(=C4)NC(=O)C(C5CCCCC5)N)N2

Solubility

In vitro
Batch:

DMSO : 84 mg/mL (200.24 mM)
(Moisture-contaminated DMSO may reduce solubility. Use fresh, anhydrous DMSO.)

Water : Insoluble

Ethanol : Insoluble

Molarity Calculator

Mass	Concentration	Volume	Molecular Weight
=	×	×

Dilution Calculator Molecular Weight Calculator

In vivo Batch:
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
Clear solution	5%DMSO 1 40%PEG300 2 5%Tween80 3 50%ddH2O Validated by Selleck labs. Should you need adjustments to this formulation, contact our sales team for custom testing.	7.500mg/ml (17.88mM)	Taking the 1 mL working solution as an example, add 50 μL of 150 mg/ml clarified DMSO stock solution to 400 μL of PEG300, mix evenly to clarify it; add 50 μL of Tween80 to the above system, mix evenly to clarify; then continue to add 500 μL of ddH2O to adjust the volume to 1 mL. The mixed solution should be used immediately for optimal results.
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.

In vivo Formulation Calculator (Clear solution)

Step 1: Enter information below (Recommended: An additional animal making an allowance for loss during the experiment)

Dosage: mg/kg Average weight of animals: g Dosing volume per animal:μL Number of animals:

Step 2: Enter the in vivo formulation (This is only the calculator, not formulation. Please contact us first if there is no in vivo formulation at the solubility Section.)

% DMSO + % + % Tween 80 + % ddH₂O

%DMSO+ %

Calculation results:

Working concentration: mg/ml;

Method for preparing DMSO master liquid: mg drug pre-dissolved in μL DMSO ( Master liquid concentration mg/mL, Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug. )

Method for preparing in vivo formulation: Take μL DMSO master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH₂O, mix and clarify.

Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.

Note: 1. Please make sure the liquid is clear before adding the next solvent.
2. Be sure to add the solvent(s) in order. You must ensure that the solution obtained, in the previous addition, is a clear solution before proceeding to add the next solvent. Physical methods such
as vortex, ultrasound or hot water bath can be used to aid dissolving.

Mechanism of Action

Targets/IC₅₀/Ki	Chk1 ^[1] (Cell-free assay) 0.49 nM(Ki) VEGFR2 ^[1] (Cell-free assay) 8 nM(Ki) Fms ^[1] (Cell-free assay) 10 nM(Ki) YES ^[1] (Cell-free assay) 14 nM(Ki) Chk2 ^[1] (Cell-free assay) 47 nM(Ki)
In vitro	PF-477736 (128 nM) abrogates the camptothecin-induced DNA damage checkpoint in a dose-dependent manner in CA46 and HeLa cells. This compound effectively abrogates the -induced S-phase arrest with a corresponding increase in apoptotic cell populations in HT29 cells. This chemical (540 nM) enhances -induced cytotoxicity in a time- and dose-dependent manner in HT29 cells. It potentiates the growth-inhibitory activity of a panel of chemotherapeutic agents across a broad spectrum of p53-deficient human cancer cell lines in the MTT assay. Addition of this compound (360 nM) to -arrested cells induces a dramatic increase in the intensity of H2AX phosphorylation, reflecting a greater number of γ-H2AX molecules near sites of DNA damage. ^[1] This chemical (0.5 nM) selectively blocks p73 and P53 phosphorylation in presence of curcumin in HL-60 cells. ^[2] It (360 nM) suppresses -induced phosphorylation of histone H3 (Ser10) and Cdc25C (Ser216) and potentiates apoptosis in COLO205 cells. ^[3] This compound (250 nM) combined with MK-1775 has marked synergistic cytotoxic activity in OVCAR-5 cells. It (250 nM) combined with MK-1775 causes accumulation of cells with a DNA content between 2N and 4N in OVCAR-5 cells. This chemical (250 nM) combined with MK-1775 causes premature mitosis before the end of DNA replication, with damaged DNA leading to apoptotic cell death in OVCAR-5 cells. ^[4]
Kinase Assay	Binding assay
Kinase Assay	The assay is performed in a 96-well plate for 20 minutes at 30℃ in 0.1 mL of assay buffer containing 50 mM TRIS pH 7.5, 0.4 M NaCl, 4 mM PEP, 0.15 mM NADH, 28 units of lactate dehydrogenase/mL, 16 units of pyruvate kinase/mL, 3 mM DTT, 0.125 mM Syntide-2, 0.15 mM ATP and 25 mM magnesium chloride. Assays are initiated with 1 nM of CHK1 kinase domain. The inhibition of CHK1 activity is determined by measuring initial velocities in the presence of varying concentrations of this compound. The data is analyzed using Enzyme Kinetic and Excel software and fit to a kinetic model for competitive inhibition to obtain a Ki value. The kinase selectivity of this chemical is evaluated by screening it at 1 μM or 10 μM against a panel 2 of about 100 protein kinases.
In vivo	PF-477736 (4 mg/kg i.v.) results in terminal half-life (T1/2) of 2.9 hours, AUC of 5.72 μg×hr/mL and CLp of 11.8 mL/min/kg in rats. This compound dose-dependently enhances the antitumor activity of a maximum tolerated dose in the Colo205 xenograft mouse model. This chemical (12 mg/kg) induces an increase in the phosphorylation of histone H3 (Ser10) and of phospho-histone H2AX in the Colo205 xenograft mouse model. ^[1] This compound (15 mg/kg i.p.) enhances induced tumor growth inhibition and tumor growth delay in COLO205 and MDA-MB-231 xenograft models. ^[3] This chemical (10 mg/kg once daily i.p.) combined with MK-1775 (30 mg/kg twice a day oral) leads to greater tumor growth inhibition in mice bearing OVCAR-5 xenografts. ^[4]
References	[1] https://pubmed.ncbi.nlm.nih.gov/18723486/ [2] https://pubmed.ncbi.nlm.nih.gov/20675383/ [3] https://pubmed.ncbi.nlm.nih.gov/19584159/ [4] https://pubmed.ncbi.nlm.nih.gov/22713237/

Applications

Methods	Biomarkers	PMID
Western blot	p-53 / p-p53 / p21 / MDM2 p-ATM / γ-H2AX	31362335
Growth inhibition assay	Cell viability	29167438
Immunofluorescence	p-p53	31362335

Clinical Trial Information

(data from https://clinicaltrials.gov, updated on 2024-05-22)

NCT Number	Recruitment	Conditions	Sponsor/Collaborators	Start Date	Phases
NCT00437203	Terminated	Neoplasms	Pfizer	December 2006	Phase 1

Tech Support

Handling Instructions

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Frequently Asked Questions

Question 1:
Can you advise the vehicle suggested for S2904: 2% DMSO/40% PEG 300 at 5mg/ml, is it a suspension or clear solution? I wanted to know how to dissolve it for an i.p. injection in mice.

Answer:
It can dissolve in 2% DMSO/40% PEG 300 at 5mg/ml as a clear solution. If you are going to use this kind of vehicle, please dissolve this compound in DMSO clearly first, then add PEG300, then water.

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