KU-0063794

Catalog No.S1226

KU-0063794 Chemical Structure

Molecular Weight(MW): 465.54

KU-0063794 is a potent and highly specific dual-mTOR inhibitor of mTORC1 and mTORC2 with IC50 of ~10 nM in cell-free assays; no effect on PI3Ks.

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In DMSO USD 180 In stock
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4 Customer Reviews

  • mTORC2-mediated stretch-induced SGK-1 phosphorylation. SMCs infected with Ad-SGK-1 were pretreated with PPP (10 μmol/L), rapamycin (100 nmol/L),or Ku-0063794 (10 μmol/L) for 30 minutes and then were subjected to stretch or IGF-1 (10 ng/mL) for 30 minutes.

     

     

    Circ Res 2010 107, 1265-1274. KU-0063794 purchased from Selleck.

    Effects of PI3K and mTOR inhibitors on IGF1R and AKT signaling in RMS cells. Rh41 cells were treated with 0.3 uM PI3K inhibitor BKM120, 0.3 uM mTOR inhibitor KU0063794 for the indicated time. Lysates were made and analyzed for p-AKT, S6RP and IGF1R.

    Oncogene 2013 10.1038/onc.2013.509. KU-0063794 purchased from Selleck.

  •  

    mTORC2 is required for mechanical activation of Akt. A, immunoblots of mdMSC subjected to strain for 45 min following treatment with the mTOR inhibitor KU0063794 (2uM). B, densitometric analysis of phosphorylated GSK3 (Ser-9) normalized to total GSK3. D, immunoblots of mdMSC treated with KU0063794 and then stimulated with insulin (50 nM) for 60 min.

    J Biol Chem 2011 286, 39450-6. KU-0063794 purchased from Selleck.

    For MTT assays, cells (2,000 ~ 5,000 cells/well) were subcultured into 96-well plates according to their growth properties. Cell proliferation was assayed at 72 hr after treatment of KU-0063794 by adding 20 μl of 5 mg/ml 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) solution per 100 μl of growth medium. After incubating for 3-4 h at 37°C, the media were removed and 150 µl/well of MTT solvent (either absolute DMSO or isopropanol containing 4 μM HCl and 0.1% Nonidet-40) was added to dissolve the formazan. The absorbance of each well was measured by ELx808 (BioTek, Winooski, VT) or Wallac Victor2 (Perkin-Elmer Life Sciences, Boston, MA) Microplate Reader. Viable cells are presented as percent of control, vehicle-treated cells.

     

     

    Dr. Yong-Weon Yi from Georgetown University Medical Center. KU-0063794 purchased from Selleck.

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Biological Activity

Description KU-0063794 is a potent and highly specific dual-mTOR inhibitor of mTORC1 and mTORC2 with IC50 of ~10 nM in cell-free assays; no effect on PI3Ks.
Targets
mTORC1 [1]
(Cell-free assay)
mTORC2 [1]
(Cell-free assay)
~10 nM ~10 nM
In vitro

Compared with the mTOR inhibitor PP242, KU-0063794 exhibits higher specificity for mTOR, as being inactive against PI3Ks or 76 other kinases. In HEK-293 cells, KU-0063794 at 30 nM is sufficient to rapidly ablate S6K1 activity by blocking the phosphorylation of the hydrophobic motif (Thr389) and subsequently the phosphorylation of the T-loop residue (Thr229). In case of IGF1 stimulation of serum-starved HEK-293 cells, 300 nM of KU-0063794 is needed to inhibit the S6K1 activity by ~90%. KU-0063794 at 100-300 nM also completely inhibits the amino-acid-induced phosphorylation of S6K1 and S6 protein. Similar to S6K1, KU-0063794 inhibits the phosphorylation of mTORC1 at Ser2448 and mTORC2 at Ser2481 in a dose-dependent and time-dependent manner. In the presence of serum or following IGF1 stimulation, KU-0063794 induces a dose-dependent inhibition of the activity and phosphorylation of Akt at Ser473 and unexpected Thr308 as well as the phosphorylation of the Akt substrates PRAS40 at Thr246, GSK3α/GSK3β at Ser21/Ser9 and Foxo-1/3a at Thr24/Thr32. KU-0063794 but not rapamycin inhibits SGK1 activity and Ser422 phosphorylation as well as its physiological substrate NDGR1 in a dose-dependent manner, to the same extent as S6K1 and Akt phosphorylation, whereas KU-0063794 dose not inhibit phorbol ester induced ERK or RSK phosphorylation and RSK activation. Compared with rapamycin, KU-0063794 exhibits more significant potency to induce the complete dephosphorylation of 4E-BP1 at Thr37, Thr46 and Ser65. KU-0063794 inhibits cell growth of both wild-type and mLST8-deficient MEFs and induces a G1 cell cycle arrest, more significantly than rapamycin. [1]

Cell Data
Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
HepG2  NGLiUnVE\WyuIG\pZYJqdGm2eTDBd5NigQ>? M3jse|AvOeLCk{WwxsDPxE1? NVSzfmlVPzJiaB?= NUG3SHlj\GWlcnXhd4V{KGOnbHygeoli[mmuaYT5JIlvKGFiZH;z[UBl\XCnbnTlcpQhdWGwbnXy MV6yOlI4QDhzOR?=
HepG2  MYjDc4xwdnliRn;ycYF1cW:wIFHzd4F6 M1KxWlHjiJN3MNMg{txO NXf6Z|NpOTBiZB?= NHXkUWxl\WO{ZXHz[ZMhfGinIH71cYJmeiCxZjD2bYFjdGViSHXwS|Ih[2:ub37p[ZPDqHOrZ37p[olk[W62bIm= M2DUclI3Ojd6OEG5
HepG2  NHywWlZCeG:ydH;zbZMhSXO|YYm= NXXuZ4d6OC5z4pETOVDDqM7:TR?= NGPTNo41QCCq M1Wxdolv\HWlZYOgZZBweHSxc3nzJIlvKGFiZH;z[UBl\XCnbnTlcpQhdWGwbnXy MljRNlYzPzh6MUm=
HepG2  NGi2ZYdHfW6ldHnvckBCe3OjeR?= NWHxOYF7PS9zMDFOwG0> M{jI[lI1KGh? MlvH[JJidWG2aXPhcIx6KGmwaHnibZR{KHCqb4PwbI9zgWyjdHnvckBw\iCDS2SgZZQhW2W{LUS3Ny=> NXXJfmJWOjZ{N{i4NVk>
HepG2  Mm\vSpVv[3Srb36gRZN{[Xl? NHnm[4w2NzFyIN88US=> NHzibnEzPCCq Ml\5[I94dnKnZ4XsZZRmeyC2aHWgcIV3\Wy|IFjJSlHPuSCjbnSgZ5lkdGmwIFSxxsA> M2fXc|I3Ojd6OEG5
HepG2  MXPGeY5kfGmxbjDBd5NigQ>? MkmyNE4y6oDVNUFCpO69VQ>? NXj2eWR2OjRiaB?= NHm0bYtqdmS3Y3XzJJA3OiCmb4fudoVofWyjdHnvckwhSmWlbHnuMVEh\XiycnXzd4lwdiCjbnSgUGM{Si2LIITvJGxEO0JvSVmgZ49vfmW{c3nvckBqdiCjIHTvd4Uh\GWyZX7k[Y51KG2jbn7ldi=> NIT4RogzPjJ5OEixPS=>
HepG3  NHf6UIlHfW6ldHnvckBCe3OjeR?= MnjqNE4y6oDVNUFCpO69VQ>? NInQTHM1QCCq Mof6bY5lfWOnczDj[YxtKGG3dH;wbIFogSCrbjDhJIRwe2ViZHXw[Y5l\W62IH3hco5meg>? NYi1e3M6OjZ{N{i4NVk>
AGS NH\3NZhIem:5dHigTY5pcWKrdHnvckBCe3OjeR?= Mle4TWM2OD1zNT6wJOKyKDJwOUGg{txO NE[xfWIzPDV7N{S3PC=>
HGC27 M4L1eGdzd3e2aDDJcohq[mm2aX;uJGF{e2G7 MWjJR|UxRTF3LkCgxtEhPC56MjFOwG0> MmG2NlQ2QTd2N{i=
MKN45 NVPhbWlmT3Kxd4ToJGlvcGmkaYTpc44hSXO|YYm= M4e1NGlEPTB;MD64NkDDuSByLkCxJO69VQ>? M3n1W|I1PTl5NEe4
NUGC4 Mn;tS5Jwf3SqIFnubIljcXSrb36gRZN{[Xl? MU\JR|UxRTJwOUOgxtEhOC5|MTFOwG0> M{DEWVI1PTl5NEe4
PC9 MYfHdo94fGhiSX7obYJqfGmxbjDBd5NigQ>? NVvEeW0yPzJiaB?= M3PqcGlEPTB;MUCuNVXDuTBwNkKgcm0> M1KxXFI{QDd2OEiw
PC9GR MU\Hdo94fGhiSX7obYJqfGmxbjDBd5NigQ>? MV[3NkBp MY\JR|UxRTZwMkJCtVEvOzBibl2= MkjTNlM5PzR6OEC=
H1650 MmnOS5Jwf3SqIFnubIljcXSrb36gRZN{[Xl? M1PwUlczKGh? MV3JR|UxRTdwNkJCtVAvPjJibl2= M4PLXVI{QDd2OEiw
H1975 NGjMb4dIem:5dHigTY5pcWKrdHnvckBCe3OjeR?= NF3aZVM4OiCq MVXJR|UxRTFzLkG1xtExNjl|IH7N NFLTOWozOzh5NEi4NC=>
PC9 M3v3emZ2dmO2aX;uJGF{e2G7 M2nWdVExNjF3IH7N NHXa[lg4OiCq NULPR3FscW6qaXLpeJMhdVSRUjDwbI9{eGixconsZZRqd25ic4TheJV{yqB? NWjKeI9ROjN6N{S4PFA>
PC9GR MnTqSpVv[3Srb36gRZN{[Xl? MWO2MlIyKG6P NXvobWJFPzJiaB?= M3fGbolvcGmkaYTzJI1VV1JicHjvd5Bpd3K7bHH0bY9vKHO2YYT1d:Kh MmX0NlM5PzR6OEC=
H1650 M{XtRmZ2dmO2aX;uJGF{e2G7 NIH6XlM4NjZzIH7N Mm\iO|IhcA>? MYPpcohq[mm2czDtWG9TKHCqb4PwbI9zgWyjdHnvckB{fGG2dYRCpC=> NUXaV5RCOjN6N{S4PFA>
H1975 MXjGeY5kfGmxbjDBd5NigQ>? NIHYd|UyOS5zNTDuUS=> NVfleY9XPzJiaB?= MkHQbY5pcWKrdIOgcXRQWiCyaH;zdIhwenmuYYTpc44he3SjdIXzxsA> MnnmNlM5PzR6OEC=
PC9 MYXGeY5kfGmxbjDBd5NigQ>? NH3LN2EyOC5zNTDuUS=> NGrQ[|E4OiCq NX6yVmhHcW6qaXLpeJMheGixc4Doc5J6dGG2aX;uJI9nKHB5MGO2Ty=> Mkj4NlM5PzR6OEC=
PC9GR MWjGeY5kfGmxbjDBd5NigQ>? M33hRlYvOjFibl2= MkT3O|IhcA>? MX;pcohq[mm2czDwbI9{eGixconsZZRqd25ib3[gdFcxWz[N NVzrN|RKOjN6N{S4PFA>
H1650 NX6wSVNMTnWwY4Tpc44hSXO|YYm= Mlq4O{43OSCwTR?= MWi3NkBp MXHpcohq[mm2czDwbI9{eGixconsZZRqd25ib3[gdFcxWz[N MoL0NlM5PzR6OEC=
H1975 MUfGeY5kfGmxbjDBd5NigQ>? MkX0NVEvOTVibl2= MY[3NkBp NHrWOZBqdmirYnn0d{BxcG:|cHjvdplt[XSrb36gc4YheDdyU{\L M371RlI{QDd2OEiw
LNCaP M4XZfGNmdGxiVnnhZoltcXS7IFHzd4F6 M1S5[|AuOTBizszN MoDkNlQhcMLi MUfk[YNz\WG|ZYOgZ4VtdCC4aXHibYxqfHliaX6gZUBld3OnIHTldIVv\GWwdDDtZY5v\XJ? MXqyN|g1ODZyNR?=
PC-3 NFXIVYJE\WyuIG\pZYJqdGm2eTDBd5NigQ>? MWWwMVExKM7:TR?= NYXK[YQ{OjRiaNMg Mn\G[IVkemWjc3XzJINmdGxidnnhZoltcXS7IHnuJIEh\G:|ZTDk[ZBmdmSnboSgcYFvdmW{ NWjqWXBJOjN6NEC2NFU>
MDA-MB-468  MUTD[YxtKF[rYXLpcIl1gSCDc4PhfS=> NUXaWW8xOC1zMDFOwG0> MWiyOEBpyqB? NEL3WYtl\WO{ZXHz[ZMh[2WubDD2bYFjcWyrdImgbY4h[SCmb4PlJIRmeGWwZHXueEBu[W6wZYK= NF[xdZgzOzh2ME[wOS=>
LNCaP MVHGeY5kfGmxbjDBd5NigQ>? NFr0PJEzODEkgKO4NFAhdk1? NH7aPXQzPCCqwrC= NFHUO2Rl\WO{ZXHz[ZMhfGinIIDoc5NxcG:{eXzheIlwdiCuZY\lcEBw\iCyN{DTOmshcW5iYTDkc5NmKGSncHXu[IVvfCCvYX7u[ZI> MofwNlM5PDB4MEW=
PC-3 M3XJWGZ2dmO2aX;uJGF{e2G7 MoWyNlAx6oDVOECwJI5O NVq2[YlnOjRiaNMg NHXLb|ll\WO{ZXHz[ZMhfGinIIDoc5NxcG:{eXzheIlwdiCuZY\lcEBw\iCyN{DTOmshcW5iYTDkc5NmKGSncHXu[IVvfCCvYX7u[ZI> MkToNlM5PDB4MEW=
MDA-MB-468  M2nyZ2Z2dmO2aX;uJGF{e2G7 NEDkOWczODEkgKO4NFAhdk1? M2XvelI1KGkEoB?= Mljk[IVkemWjc3XzJJRp\SCyaH;zdIhwenmuYYTpc44hdGW4ZXygc4YheDdyU{\LJIlvKGFiZH;z[UBl\XCnbnTlcpQhdWGwbnXy M1u2b|I{QDRyNkC1
Caki-1  NX71[|JCTnWwY4Tpc44hSXO|YYm= MWmxNFAuOjByMDDuUS=> NIDVVYcyOC1zOECgcYlv M163OGROW09? NXrZcXljcW6qaXLpeJMh[m:2aDDtWG9TSzFiYX7kJI1VV1KFMjDhd{BqdmSrY3H0[YQh[nlidHjlJIRm[3KnYYPlJIlvKHCqb4PwbI9zgWyjdHnvckBw\iCmb4fud5Rz\WGvIHXm[oVkfG:{cx?= NWq2ZohLOjN|NEm5PFk>
786-O NULVbWxiTnWwY4Tpc44hSXO|YYm= NFrN[5oyODBvMkCwNEBvVQ>? NHz1TnkyOC1zOECgcYlv Ml33SG1UVw>? NGPHb|ZqdmirYnn0d{Bjd3SqIH3UU3JEOSCjbnSgcXRQWkN{IHHzJIlv\GmlYYTl[EBjgSC2aHWg[IVkemWjc3WgbY4heGixc4Doc5J6dGG2aX;uJI9nKGSxd37zeJJm[W1iZX\m[YN1d3K| Mo\oNlM{PDl7OEm=
Caki-1  NITSS3VE\WyuIG\pZYJqdGm2eTDBd5NigQ>? NWPsd5dIOzByLUSwNFAhdk1? Ml31NlQuQTZiaB?= NHj0T4JFVVOR M4fIcZN2eHC{ZYPz[ZMhfGinIHPlcIwhfmmjYnnsbZR6KGmwIHLveIghfGmvZTDhcoQh\G:|ZTDk[ZBmdmSnboSgcYFvdmW{ MW[yN|M1QTl6OR?=
786-O MYfD[YxtKF[rYXLpcIl1gSCDc4PhfS=> MUSzNFAuPDByMDDuUS=> NWHyXpBHOjRvOU[gbC=> NWPC[5liTE2VTx?= Mlfid5VxeHKnc4Pld{B1cGViY3XscEB3cWGkaXzpeJkhcW5iYn;0bEB1cW2nIHHu[EBld3OnIHTldIVv\GWwdDDtZY5v\XJ? MXeyN|M1QTl6OR?=
Caki-1  MVzGeY5kfGmxbjDBd5NigQ>? MmHSNkDDvU1? NWfOcpFGPzJiaB?= NF3tT3RFVVOR Mn;jbY5lfWOnczDHNUBk\WyuIHP5Z4xmKGG{cnXzeEBidmRiYYX0c5Bp[We7 MkizNlM{PDl7OEm=
786-O Mn3ZSpVv[3Srb36gRZN{[Xl? NVP1epNsOiEEtV2= MXG3NkBp NH;zVVdFVVOR NUPMTG9KcW6mdXPld{BIOSClZXzsJIN6[2ynIHHydoV{fCCjbnSgZZV1d3CqYXf5 NHLIc|kzOzN2OUm4PS=>

... Click to View More Cell Line Experimental Data

In vivo Ku0063794 inhibits tumor growth and mTOR signaling in a preclinical renal cell carcinoma model. However, Ku0063794 was not more effective than temsirolimus in the animal study. A possible explanation for lack of greater activity in vivo for Ku0063794 is that temsirolimus has important effects on the tumor microenvironment. Temsirolimus decreased angiogenesis in the xenograft tumors while Ku0063794 did not. Temsirolimus treated tumors expressed less VEGF and PDGF than Ku0063794 treated tumors, thus stimulating less angiogenesis[2].

Protocol

Kinase Assay:

[1]

+ Expand

mTOR complexes kinase assays:

HEK-293 cells are freshly lysed in Hepes lysis buffer. Lysate (1-4 mg) is pre-cleared by incubating with 5-20 μL of Protein G-Sepharose conjugated to pre-immune IgG. The lysate extracts are then incubated with 5-20 μL of Protein G-Sepharose conjugated to 5-20 μg of either anti-Rictor or anti-Raptor antibody, or pre-immune IgG. All antibodies are covalently conjugated to Protein G-Sepharose. Immunoprecipitations are carried out for 1 hour at 4 °C on a vibrating platform. The immunoprecipitates are washed four times with Hepes lysis buffer, followed by two washes with Hepes kinase buffer. For Raptor immunoprecipitates used for phosphorylating S6K1, for the initial two wash steps the buffer includes 0.5 M NaCl to ensure optimal kinase activity. GST-Akt1 is isolated from serum-deprived HEK-293 cells incubated with PI-103 (1 μM for 1 hour). GST-S6K1 is purified from serum-deprived HEK-293 cells incubated with rapamycin (0.1 μM for 1 hour). mTOR reactions are initiated by adding 0.1 mM ATP and 10 mM MgCl2 in the presence of various concentrations of KU-0063794 and GST-Akt1 (0.5 μg) or GST-S6K1 (0.5 μg). Reaction are carried out for 30 minutes at 30 °C on a vibrating platform and stopped by addition of SDS sample buffer. Reaction mixtures are then filtered through a 0.22-μm-poresize Spin-X filter and samples are subjected to electrophoresis and immunoblot analysis with the indicated antibodies.
Cell Research:

[1]

+ Expand
  • Cell lines: Wild-type and mLST8 deficient MEFs
  • Concentrations: Dissolved in DMSO, final concentration ~3 μM
  • Incubation Time: 24, 48, and 72 hours
  • Method:

    Cells are treated with KU-0063794 for 24, 48, and 72 hours, and the medium is changed every 24 hours with freshly dissolved KU-0063794. For the measurement of cell growth, cells are washed once with PBS, and fixed in 4% (v/v) paraformaldehyde in PBS for 15 minutes. After washing once with water, the cells are stained with 0.1% Crystal Violet in 10% ethanol for 20 minutes and washed three times with water. Crystal Violet is extracted from cells with 0.5 mL of 10% (v/v) ethanoic (acetic) acid for 20 minutes. The eluate is then diluted 1:10 in water and absorbance at 590 nm is quantified. For the assessment of cell cycle distribution, cells are harvested by trypsinization, washed once in PBS, and re-suspended in ice-cold aq. 70% (v/v) ethanol. Cells are washed twice in PBS plus 1% (w/v) BSA and stained for 20 minutes in PBS plus 0.1% (v/v) Triton X-100 containing 50 g/mL propidium iodide and 50 g/mL RNase A. The DNA content of cells is determined using a FACSCalibur flow cytometer and CellQuest software. Red fluorescence (585 nm) is acquired on a linear scale, and pulse width analysis is used to exclude doublets. Cell-cycle distribution is determined using FlowJo software.


    (Only for Reference)
Animal Research:

[2]

+ Expand
  • Animal Models: Nu/Nu nude mice
  • Formulation: one part DMSO and then diluted (200 µg/100 µl) with 4 parts PEG1500 (50% (w/v) in 75 mM Hepes, pH 8.0
  • Dosages: 8 mg/kg
  • Administration: i.p.
    (Only for Reference)

Solubility (25°C)

In vitro DMSO 16 mg/mL (34.36 mM)
Water Insoluble
Ethanol Insoluble
In vivo Add solvents to the product individually and in order:
30% PEG400+0.5% Tween80+5% propylene glycol
For best results, use promptly after mixing.
30 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 465.54
Formula

C25H31N5O4

CAS No. 938440-64-3
Storage powder
Synonyms N/A

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Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID