PD0166285

For research use only.

Catalog No.S8148

1 publication

PD0166285 Chemical Structure

CAS No. 185039-89-8

PD0166285 is a potent Wee1 and Chk1 inhibitor with activity at nanomolar concentrations (IC50=24 nM for Wee1 and 72 nM for Myt1). PD0166285 is also a novel G2 checkpoint abrogator. PD0166285 induces apoptosis.

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Selleck's PD0166285 has been cited by 1 publication

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Biological Activity

Description PD0166285 is a potent Wee1 and Chk1 inhibitor with activity at nanomolar concentrations (IC50=24 nM for Wee1 and 72 nM for Myt1). PD0166285 is also a novel G2 checkpoint abrogator. PD0166285 induces apoptosis.
Targets
Wee1 [1]
(Cell-free assay)
Myt1 [1]
(Cell-free assay)
Chk1 [1]
(Cell-free assay)
24 nM 72 nM 3.4 μM
In vitro

PD0166285 is identified to inhibit Wee1 activity at nanomolar concentrations. The inhibitor abrogates G2/M checkpoint inducing early cell division. At the cellular level, 0.5 μM PD0166285 dramatically inhibits irradiation-induced Cdc2 phosphorylation at the Tyr-15 and Thr-14 in seven of seven cancer cell lines tested. This G2 checkpoint abrogation by PD0166285 is demonstrated to kill cancer cells. PD0166285 does not inhibit Cdc2/cyclin B but inhibits Chk1 kinase at a much higher concentration (3433 nM). the treatment of cells with the inhibitor is related to microtubule stabilization and decrease in cyclin D transcription. Thus, PD0166285 may be a potentially useful anti-cancer therapy[1][2].

Cell Data
Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
NIH3T3 cells Mny4SpVv[3Srb36gZZN{[Xl? M1;j[GlvcGmkaYTpc44hd2ZiUFTHSk1qdmS3Y3XkJIF2fG:yaH;zdIhwenmuYYTpc44hd2ZiUFTHSk1TKGmwIF7JTFNVOyClZXzsd{whUUN3ME2xMlghdk1? M1fjZlk4ODN2N{O=
C6 cells M17QUmZ2dmO2aX;uJIF{e2G7 MlXPTY5pcWKrdHnvckBw\iCSRFfGMYlv\HWlZXSgZZV1d3Cqb4PwbI9zgWyjdHnvckBw\iCSRFfGMXIhcW5iQ{[gZ4VtdHNuIFnDOVA:PSCwTR?= MlOzPVcxOzR5Mx?=
HEK293 cells MmnmSpVv[3Srb36gZZN{[Xl? M1HvbmJqdmSrbnegZYZncW6rdImgeI8hcHWvYX6g[pVtdC2uZX7neIghUGm|LYTh[4dm\CCPeYSxJItqdmG|ZTDlfJBz\XO|ZXSgbY4hUEWNMkmzJINmdGy|IHL5JHRTNU[URWSgZoF{\WRiYnnu[Ilv\yCjc4PhfUwhUUN3ME23MlIhdk1? MYqyNlc4ODZzMB?=
SW-620 MWLGeY5kfGmxbjDhd5NigQ>? MlzTTY5pcWKrdH;yfUBi[3Srdnn0fUBi\2GrboP0JHNYNTZ{MDDoeY1idiClb3zvckBi\GWwb3PhdoNqdm:vYTDj[YxteyC5YYOg[IV1\XKvaX7l[EwhUUN3ME2wMlI4KM7:TR?= M2n0XVExQTV4MkKy
HT-29 MnLkSpVv[3Srb36gZZN{[Xl? NHHBdFhKdmirYnn0c5J6KGGldHn2bZR6KGGpYXnud5QhUFRvMkmgbJVu[W5iY3;sc44h[WSnbn;jZZJkcW6xbXGgZ4VtdHNid3HzJIRmfGW{bXnu[YQtKEmFNUC9NE4zQCEQvF2= MWKxNFk2PjJ{Mh?=
C6 cell NYfGXodFTnWwY4Tpc44h[XO|YYm= NVuySYtMUW6qaXLpeIlwdiCxZjDDOkBk\WyuIIDyc4xq\mW{YYTpc44hcW5ic4Xs[o9zcG:mYX3pcoUhSiCjc4PhfUwhUUN3ME2wMlM1KM7:TR?= NV;SU291QTdyM{S3Ny=>
HCT-8 MkPTSpVv[3Srb36gZZN{[Xl? M4rxTGlvcGmkaYTvdpkh[WO2aY\peJkh[WejaX7zeEBJS1RvODDoeY1idiClb3zvckBi\GWwb3PhdoNqdm:vYTDj[YxteyC5YYOg[IV1\XKvaX7l[EwhUUN3ME2wMlQzKM7:TR?= NXy4dFIxOTB7NU[yNlI>

... Click to View More Cell Line Experimental Data

Assay
Methods Test Index PMID
Western blot
p-Cdc2 / Cdc2 / Wee1 / Cyclin B ; 

PubMed: 17177986     


Immunoblot analysis of anti-cdc2, anti-phosphorylated cdc2-tyr15, and Wee1 was performed in lysates of with 0.5 μM PD0166285 treated for 0 h, 2 h, or 4 h.

17177986
Immunofluorescence
Wee1 / α-tubulin ; 

PubMed: 17177986     


Effects of PD0166285 on the cellular location of Wee1 and adhesion. A: After 4 h of PD0166285 treatment, cells were fixed and doubly stained for immunofluorescence with PI (red) and anti-Wee1 (green). B: After 4 h of 0.5 μM PD0166285 treatment, morphologic changes and double stained with anti-α-tubulin (green) and anti-Wee1 (red) were observed. Shown are typical images obtained by laser confocal microscopy. Bar: 10 μm.

17177986
In vivo PD0166285 at 0.5 μM concentration can inhibit Cdc2Y15 /T14 phosphorylation in all cell lines tested, regardless of their p53 status[1] and pharmacological targeting of WEE1 by PD0166285 sensitizes U251-FM GBM tumors to IR in vivo[3].

Protocol

Kinase Assay:

[1]

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Wee1 Mass Screening:

Wee1 mass screening is performed using Amersham’s p34cdc2 kinase SPA kit with some modifications. Briefly, 45–60 nM full-length Wee1 kinase is incubated with 25 μM compounds, 20 μM ATP, and 122–441 nM Cdc2/cyclin B in a final volume of 50 μl of enzyme dilution buffer [50 mM Tris (pH 8.0), 10 mM NaCl, 10 mM MgCl2, 1 mM DTT, and 0.1 mM Na3VO4]. After 30 min incubation at 30°C, 30 μl of [33P]ATP containing kinase buffer [67 mM Tris (pH 8.0), 40 mM NaCl, 13 mM MgCl2, 1 mM DTT, and 0.13 mM Na3VO4] containing 1 μM biotinylated peptide, and 0.25 μCi of [γ-33P]ATP is added to the reaction and incubated for another 30 min at 30°C. The reaction is stopped by adding 200 μl of stop buffer [50 μM ATP, 5 mM EDTA, 0.1% Triton X-100, and 1.25 mg/ml SPA beads in PBS]. After centrifugation at 2400 rpm for 15 min, the plate is counted with Wallac’s Microbeta counter.
Cell Research:

[2]

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  • Cell lines: B16 mouse melanoma cells
  • Concentrations: 0.5, 1, 2 μM
  • Incubation Time: 4 h, 24 h
  • Method:

    B16 cells (1 × 105 cells in 100 mm-dishes) are maintained in medium overnight. In addition, the cells are treated with (0, 0.5, 1.0, or 2.0 μM) PD0166285 (including DMSO vehicle) for indicated times. The cells are washed twice with phosphate-buffered saline (PBS). Next, the cells are trypsinized, so the cell numbers in each dish are determined by using a computed cell-counter (Sysmex CDA-500) according to manufacturer's recommendation.


    (Only for Reference)
Animal Research:

[3]

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  • Animal Models: Nude mice with implanted GBM cells
  • Dosages: 0, 20, 100, 200, or 400 µM in 100 µl
  • Administration: s.c.
    (Only for Reference)

Solubility (25°C)

In vitro DMSO 100 mg/mL (195.14 mM)
Water Insoluble
Ethanol '100 mg/mL

* 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.

Chemical Information

Molecular Weight 512.43
Formula

C26H27Cl2N5O2

CAS No. 185039-89-8
Storage powder
in solvent
Synonyms N/A
Smiles CCN(CC)CCOC1=CC=C(C=C1)NC2=NC=C3C=C(C(=O)N(C3=N2)C)C4=C(C=CC=C4Cl)Cl

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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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Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID