PHA-767491

For research use only.

Catalog No.S2742 Synonyms: CAY10572, NMS 1116354

27 publications

PHA-767491 Chemical Structure

CAS No. 942425-68-5

PHA-767491 (CAY10572, NMS 1116354) is a potent ATP-competitive dual Cdc7/CDK9 inhibitor with IC50 of 10 nM and 34 nM in cell-free assays, respectively.It displays ~20-fold selectivity against CDK1/2 and GSK3-β, 50-fold selectivity against MK2 and CDK5, 100-fold selectivity against PLK1 and CHK2.

Selleck's PHA-767491 has been cited by 27 publications

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Biological Activity

Description PHA-767491 (CAY10572, NMS 1116354) is a potent ATP-competitive dual Cdc7/CDK9 inhibitor with IC50 of 10 nM and 34 nM in cell-free assays, respectively.It displays ~20-fold selectivity against CDK1/2 and GSK3-β, 50-fold selectivity against MK2 and CDK5, 100-fold selectivity against PLK1 and CHK2.
Features The first inhibitor that directly affects the mechanisms controlling initiation as opposed to elongation in DNA replication.
Targets
Cdc7 [1]
(Cell-free assay)
CDK9 [1]
(Cell-free assay)
GSK-3β [1]
(Cell-free assay)
CDK2 [1]
(Cell-free assay)
CDK1 [1]
(Cell-free assay)
10 nM 34 nM 220 nM 240 nM 250 nM
In vitro

PHA-767491 displays approximately 20-fold selectivity for Cdk1, Cdk2 and GSK3-β, 50-fold selectivity for MK2 and Cdk5 and 100-fold selectivity for PLK1 and CHK2. PHA-767491 inhibits cell proliferation in a variety of human cell lines with IC50 of 0.86 μM for SF-268 to 5.87 μM for K562, and significantly induces apoptosis in a p53-independent manner in almost all cell lines in contrast with 5-FU or gemcitabine which only works in a few of cell lines. Unlike current DNA synthesis inhibitors, PHA-767491 treatment at 5 μM blocks the initiation of DNA replication but not replication fork progression, due to specific inhibition of Cdc7 kinase and Mcm2 phosphorylation at the Cdc7-dependent Ser40 site. [1] The up-regulated Mcl-1 levels in ABT-737-resistant OCI-LY1 and SU-DHL-4 cells can be significantly decreased by PHA-767491 treatment at 3 μM possibly due to the inhibition of Cdk9, leading to the restoration of the sensitivity to ABT-737. [2] The direct mitochondrial dependent pro-apoptosis effect of PHA-767491 is also observed when applied at 1 μM in quiescent chronic lymphocytic leukemia (CLL) cells through the similar mechanism with EC50 of 0.34-0.97 μM. While in proliferating CLL cells stimulated by CD154 and interleukin-4, PHA-767491 treatment at 5 μM abolishes DNA synthesis by inhibiting Cdc7 rather than triggering cell death. [3]

Cell Data
Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
human SF268 cells MkfGVJJwdGmoZYLheIlwdiCjc4PhfS=> Ml3PO|IhcA>? MnPqRY51cXC{b3zp[oVz[XSrdnWgZYN1cX[rdImgZYdicW6|dDDwOVMh\GWoaXPp[Y51KGi3bXHuJHNHOjZ6IHPlcIx{KGGodHXyJFczKGi{czygTWM2OD1yLki2JO69VQ>? MV2xPFQ3QThyOR?=
human HCT116 cells MnPJVJJwdGmoZYLheIlwdiCjc4PhfS=> NIHQ[GE4OiCq MmXkRY51cXC{b3zp[oVz[XSrdnWgZYN1cX[rdImgZYdicW6|dDDoeY1idiCKQ2SxNVYh[2WubIOg[ZhxemW|c3nu[{BxPTNiZ3Xu[UBi\nSncjC3NkBpenNiYomgdJJwdGmoZYLheIl3\SCjc4PhfUwhUUN3ME2wMlk4KM7:TR?= MVixPFQ3QThyOR?=
human HCT16 cells M3;sSXBzd2yrZnXyZZRqd25iYYPzZZk> NYrIbWNwPzJiaB?= M2fYPGFvfGmycn;sbYZmemG2aY\lJIFkfGm4aYT5JIFo[Wmwc4SgbJVu[W5iSFPUNVYh[2WubIOgZYZ1\XJiN{KgbJJ{KGK7IHz1Z4ln\XKjc3WgZoF{\WRiYYPzZZktKEmFNUC9NUDPxE1? MmnNNVkyOTV6NEW=
human SW403 cells MVTQdo9tcW[ncnH0bY9vKGG|c3H5 M1fVfVczKGh? NYTiSJF{SW62aYDyc4xq\mW{YYTpeoUh[WO2aY\peJkh[WejaX7zeEBpfW2jbjDTW|QxOyClZXzsd{Bi\nSncjC3NkBpenNiYomgcJVkcW[ncnHz[UBj[XOnZDDhd5NigSxiSVO1NF0yKM7:TR?= MXqxPVEyPTh2NR?=
human A2780 cells NVvx[5kxWHKxbHnm[ZJifGmxbjDhd5NigQ>? NF;afpY4OiCq MkXSRY51cXC{b3zp[oVz[XSrdnWgZYN1cX[rdImgZYdicW6|dDDoeY1idiCDMke4NEBk\WyuczDlfJBz\XO|aX7nJJA2OyCpZX7lJIFnfGW{IEeyJIhzeyCkeTDwdo9tcW[ncnH0bZZmKGG|c3H5MEBKSzVyPUGuNFch|ryP Mlf3NVg1Pjl6MEm=
human SW48 cells M{LzXnBzd2yrZnXyZZRqd25iYYPzZZk> M2T4N|czKGh? MVjBcpRqeHKxbHnm[ZJifGm4ZTDhZ5Rqfmm2eTDh[4FqdnO2IHj1cYFvKFOZNEigZ4VtdHNiYX\0[ZIhPzJiaILzJIJ6KGy3Y3nm[ZJie2ViYnHz[YQh[XO|YYmsJGlEPTB;MT6yJO69VQ>? MXyxPVEyPTh2NR?=
human MCF7 cells M2ntcnBzd2yrZnXyZZRqd25iYYPzZZk> NF7XTIE4OiCq MnfvRY51cXC{b3zp[oVz[XSrdnWgZYN1cX[rdImgZYdicW6|dDDoeY1idiCPQ1[3JINmdGy|IHHmeIVzKDd{IHjyd{BjgSCudXPp[oVz[XOnIHLhd4VlKGG|c3H5MEBKSzVyPUGuN{DPxE1? M{DqNlE6OTF3OES1
human U2OS cells NWjMcWQzWHKxbHnm[ZJifGmxbjDhd5NigQ>? NYHD[25IPzJiaB?= NEPkfVJCdnSrcILvcIln\XKjdHn2[UBi[3Srdnn0fUBi\2GrboP0JIh2dWGwIGWyU3Mh[2WubIOg[ZhxemW|c3nu[{BxPTNiZ3Xu[UBi\nSncjC3NkBpenNiYomgdJJwdGmoZYLheIl3\SCjc4PhfUwhUUN3ME2xMlQ6KM7:TR?= MmDENVg1Pjl6MEm=
human COLO205 cells NYLPNpNTWHKxbHnm[ZJifGmxbjDhd5NigQ>? M2r4SFczKGh? MnrVRY51cXC{b3zp[oVz[XSrdnWgZYN1cX[rdImgZYdicW6|dDDoeY1idiCFT1zPNlA2KGOnbHzzJIFnfGW{IEeyJIhzeyCkeTDseYNq\mW{YYPlJIJie2WmIHHzd4F6NCCLQ{WwQVEvPSEQvF2= NGi3[YsyQTFzNUi0OS=>
human OVCAR8 cells M2XQ[XBzd2yrZnXyZZRqd25iYYPzZZk> M1XUU|czKGh? MYnBcpRqeHKxbHnm[ZJifGm4ZTDhZ5Rqfmm2eTDh[4FqdnO2IIC1N{Bl\W[rY3nlcpQhcHWvYX6gU3ZESVJ6IHPlcIx{KGGodHXyJFczKGi{czDifUBxem:uaX\ldoF1cX[nIHHzd4F6NCCLQ{WwQVEvPTZizszN NHvFU2EyQDR4OUiwPS=>
human L363 cells M2HT[nBzd2yrZnXyZZRqd25iYYPzZZk> NHv6eG04OiCq NHXhUFJCdnSrcILvcIln\XKjdHn2[UBi[3Srdnn0fUBi\2GrboP0JIh2dWGwIFyzOlMh[2WubIOgZYZ1\XJiN{KgbJJ{KGK7IHz1Z4ln\XKjc3WgZoF{\WRiYYPzZZktKEmFNUC9NU43KM7:TR?= MUixPVEyPTh2NR?=
human NHDF cells NYiySIt2WHKxbHnm[ZJifGmxbjDhd5NigQ>? MV63NkBp M1XhWWFvfGmycn;sbYZmemG2aY\lJIFkfGm4aYT5JIFo[Wmwc4SgbJVu[W5iTljESkBk\WyuczDh[pRmeiB5MjDodpMh[nlibIXjbYZmemG|ZTDiZZNm\CCjc4PhfUwhUUN3ME2xMlYh|ryP MlnnNVkyOTV6NEW=
human NCI-H929 cells MX;Qdo9tcW[ncnH0bY9vKGG|c3H5 MkS1O|IhcA>? Mor6RY51cXC{b3zp[oVz[XSrdnWgZYN1cX[rdImgZYdicW6|dDDoeY1idiCQQ1mtTFkzQSClZXzsd{Bi\nSncjC3NkBpenNiYomgcJVkcW[ncnHz[UBj[XOnZDDhd5NigSxiSVO1NF0yNjhizszN M2XmNFE6OTF3OES1
human SF539 cells Mn60VJJwdGmoZYLheIlwdiCjc4PhfS=> NHewNVc4OiCq MmLURY51cXC{b3zp[oVz[XSrdnWgZYN1cX[rdImgZYdicW6|dDDoeY1idiCVRkWzPUBk\WyuczDlfJBz\XO|aX7nJJA2OyCpZX7lJIFnfGW{IEeyJIhzeyCkeTDwdo9tcW[ncnH0bZZmKGG|c3H5MEBKSzVyPUKuN|Qh|ryP MkjoNVg1Pjl6MEm=
human SW480 cells NWr2VJhKWHKxbHnm[ZJifGmxbjDhd5NigQ>? NGP0XFA4OiCq M1HZeWFvfGmycn;sbYZmemG2aY\lJIFkfGm4aYT5JIFo[Wmwc4SgdFU{KGSnZnnjbYVvfCCqdX3hckBUXzR6MDDj[YxteyCjZoTldkA4OiCqcoOgZpkheHKxbHnm[ZJifGm4ZTDhd5NigSxiSVO1NF0zNjZ5IN88US=> NFX0enkyQDR4OUiwPS=>
human NCI60 cells MYPQdo9tcW[ncnH0bY9vKGG|c3H5 Mke2RY51cXC{b3zp[oVz[XSrdnWgZYN1cX[rdImgZYdicW6|dDDoeY1idiCQQ1m2NEBk\WyuczygTWM2OD1|LkGg{txO MnjPNVg1Pjl6MEm=
human Jurkat cells MVPQdo9tcW[ncnH0bY9vKGG|c3H5 NHTCemQ4OiCq M2jmfGFvfGmycn;sbYZmemG2aY\lJIFkfGm4aYT5JIFo[Wmwc4SgdFU{KGSnZnnjbYVvfCCqdX3hckBLfXKtYYSgZ4VtdHNiYX\0[ZIhPzJiaILzJIJ6KHC{b3zp[oVz[XSrdnWgZZN{[XluIFnDOVA:Oy5{IN88US=> NWnYUodVOTh2Nkm4NFk>
human HCT15 cells NFXudoxRem:uaX\ldoF1cW:wIHHzd4F6 NE\tRXY4OiCq MWTBcpRqeHKxbHnm[ZJifGm4ZTDhZ5Rqfmm2eTDh[4FqdnO2IHj1cYFvKEiFVEG1JINmdGy|IHX4dJJme3OrbnegdFU{KGenbnWgZYZ1\XJiN{KgbJJ{KGK7IIDyc4xq\mW{YYTpeoUh[XO|YYmsJGlEPTB;Mz64NUDPxE1? NEjGV3EyQDR4OUiwPS=>
human OPM2 cells NXHPdIZnWHKxbHnm[ZJifGmxbjDhd5NigQ>? M4ftelczKGh? M3;IeWFvfGmycn;sbYZmemG2aY\lJIFkfGm4aYT5JIFo[Wmwc4SgbJVu[W5iT2DNNkBk\WyuczDh[pRmeiB5MjDodpMh[nlibIXjbYZmemG|ZTDiZZNm\CCjc4PhfUwhUUN3ME20MlUh|ryP NFLHc4oyQTFzNUi0OS=>
human HT-29 cells Ml23VJJwdGmoZYLheIlwdiCjc4PhfS=> NYDTbGFRPzJiaB?= NG\He3dCdnSrcILvcIln\XKjdHn2[UBi[3Srdnn0fUBi\2GrboP0JIh2dWGwIFjUMVI6KGOnbHzzJIFnfGW{IEeyJIhzeyCkeTDseYNq\mW{YYPlJIJie2WmIHHzd4F6NCCLQ{WwQVUh|ryP NES1[WsyQTFzNUi0OS=>
human K562 cells NEHqZYZRem:uaX\ldoF1cW:wIHHzd4F6 Mn23O|IhcA>? NILEdnlCdnSrcILvcIln\XKjdHn2[UBi[3Srdnn0fUBi\2GrboP0JJA2OyCmZX\pZ4lmdnRiaIXtZY4hUzV4MjDj[YxteyCjZoTldkA4OiCqcoOsJGlEPTB;NT64O{DPxE1? MlHrNVg1Pjl6MEm=
U937 cells MmPJSpVv[3Srb36gZZN{[Xl? MX3Jcohq[mm2aX;uJI9nKFSQRnHsdIhiKHC{b3T1Z5Rqd25iaX6gWVk{PyClZXzsd{whUUN3ME2xPUDPxE1? NEL3dogyPzR6MEC2OC=>
human HeLa cells NHe1OJFHfW6ldHnvckBie3OjeR?= M3u5elUh|ryP M{HKSlI1KGh? NUPLfYRVUW6mdXP0bY9vKG:oIHHwc5B1d3OrczDpckBpfW2jbjDI[WxiKGOnbHzzJIF{e2W|c3XkJIF{KGGycHXhdoFv[2Vib3[gVGFTWCCjdDC1JJVOKGGodHXyJFI1KGi{cx?= MXyxPFQ3QThyOR?=
NHDF MX\GeY5kfGmxbjDhd5NigQ>? NX7TfJB[PSEQvF2= NH;BOooyPiCq NX7uXHEzUW6mdXP0bY9vKG:oIHPlcIwh[3mlbHWgZZJz\XO2IHnuJJRpgW2rZHnu[UBl\W[rY3nlcpQhVkiGRjDhd5Nme3OnZDDhd{BFVkFic4nueIhme2m|IHnuJHMueGijc3WgZZQhPSC3TTDh[pRmeiBzNnjyd{BHSUOVIHHuZYx6e2m|IHnuJJBz\XOnbnPlJI9nKHOncoXt Mn\6NVg1Pjl6MEm=

... Click to View More Cell Line Experimental Data

Assay
Methods Test Index PMID
Western blot
p-MCM2 / CDC7 ; 

PubMed: 24902048     


HeLa cells were treated with PHA-767491 for the indicated time. Protein extracts were prepared and analysed by western blot with the indicated antibodies.

RNA Pol II / p-RNA Pol II / Caspase-3 / PARP / Mcl-1 / XIAP / Bcl-xL / Bcl-2 / NOXA; 

PubMed: 24202326     


KMS-18 and MM1S myeloma cells were incubated with 5 μM PHA-767491 for the indicated time. Protein extracts were prepared and analyzed by immunoblotting using the indicated antibodies

24902048 24202326
In vivo Administration of PHA-767491 twice a day for 5 days significantly inhibits the growth of HL60 xenograft in a dose-dependent manner with TGI of 50% and 92% at dose of 20 mg/kg and 30 mg/kg, respectively, the effect of which is also marked in A2780, Mx-1, and HCT-116 xenograft models as well as the DMBA-induced mammary carcinomas, and correlates with Cdc7 inhibition and subsequently decreased phosphorylation of Mcm2 at the Cdc7-dependent site Ser40 [1]

Protocol

Kinase Assay:[1]
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In vitro kinase assays:

The inhibition of Cdc7 and Cdk9 by PHA-767491 (IC50) is determined using the strong anion exchanger (Dowex 1-X8 resin, formate form)-based assay. For each enzyme, the absolute Km values for ATP and the specific substrate are initially determined, and each assay is then run at optimized ATP/33P-γ-ATP mix (2Km) and substrate (5Km) concentrations. Cdc7 kinase assay is performed in a buffer containing 50 mM Hepes pH 7.9, 15 mM MgCl2, 2 mM β- glycerylphosphate, 0.2 mg/mL BSA, 1 mM DTT, 3 μM Na3VO4, 2Km ATP/33P-γ-ATP mix, 5Km Mcm2 (aa 10-294), 37 nM of recombinant Cdc7/Dbf4 and increasing concentration of PHA-767491 in a final volume of 30 μL, and incubated for 1 hour at 25 °C. Cdk9 kinase assay is performed using 50 nM of recombinant Cdk9/cyclin T in 50 mM HEPES pH 7.5, 10 mM MgCl2, 1 mM DTT, 3 μM Na3VO4, 2Km ATP/33P-γ-ATP mix, 5Km RNA polymerase CDT peptide and increasing concentration of PHA-767491 in a final volume of 30 μL, and incubated for 1 hour at 25 °C. After incubation, an amount of 150 μL of resin/formate (pH 3.0) is added to stop the reaction and capture unreacted 33P-γ-ATP, separating it from the phosphorylated substrate in solution. After 1 hour of rest, a volume of 50 μL supernatant is transferred to Optiplate 96-well plates. After the additon of 150 μL of Microscint 40, the radioactivity is counted in the TopCount.
Cell Research:[1]
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  • Cell lines: HeLa, MCF7, HCT-116, U2OS, A2780, K562, SF-539, SF-268, Ovcar8, SW480, COLO205, HCT-15, Jurkat, PC3, and NHDF
  • Concentrations: Dissolved in DMSO, final concentrations ~ 20 μM
  • Incubation Time: 24 or 72 hours
  • Method: Cells are exposed to PHA-767491 for 24 or 72 hours. Cells are lysed and the ATP content in the well, used as a measure of viable cells, is determined using a thermostable firefly luciferase–based assay. Activation of caspase-3 and caspase-7 is measured as a ratio between treated sample and untreated control with a luciferase-based assay, containing a specific proluminescent substrate. DNA replication is measured as incorporation of nucleotide analog BrdU into DNA by flow cytometry.
    (Only for Reference)
Animal Research:[1]
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  • Animal Models: Female SCID mice subcutaneously implanted with HL60 cells, male Hsd, athymic nu-nu mice subcutaneously implanted with HCT116 cells, A2780 or Mx-1 cells, and female Sprague-Dawley rats with DMBA-induced mammary carcinomas
  • Dosages: ~50 mg/kg
  • Administration: Intravenous or oral administration twice a day
    (Only for Reference)

Solubility (25°C)

In vitro DMSO 24 mg/mL (96.11 mM)
Water Insoluble
Ethanol Insoluble
In vivo Add solvents to the product individually and in order(Data is from Selleck tests instead of citations):
5% DMSO+30% PEG 300+2% Tween 80+ddH2O
For best results, use promptly after mixing.
1mg/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 249.7
Formula

C12H11N3O.HCl

CAS No. 942425-68-5
Storage powder
in solvent
Synonyms CAY10572, NMS 1116354
Smiles C1CNC(=O)C2=C1NC(=C2)C3=CC=NC=C3.Cl

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Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID