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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Cited by 18 Publications

4 Customer Reviews

  • 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.

  • 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.

  • 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  MVnD[YxtKF[rYXLpcIl1gSCDc4PhfS=> NGD3PZQxNjIkgKO1NOKh|ryP MX63NkBp MnHB[IVkemWjc3XzJINmdGxidnnhZoltcXS7IHnuJIEh\G:|ZTDk[ZBmdmSnboSgcYFvdmW{ NX70fm9EOjZ{N{i4NVk>
HepG2  NWDJWWhkS2:ub375JGZwem2jdHnvckBCe3OjeR?= MXKx5qCUPTEEoN88US=> Mlu5NVAh\A>? NX\sUmRK\GWlcnXhd4V{KHSqZTDueY1j\XJib3[geoli[mynIFjldGczKGOxbH;ubYV{yqC|aXfubYZq[2GwdHz5 MUmyOlI4QDhzOR?=
HepG2  NXLtSW5PSXCxcITvd4l{KEG|c3H5 NUjuRpVtOC5z4pETOVDDqM7:TR?= NV;YNVVvPDhiaB?= NF\YU|NqdmS3Y3XzJIFxd3C2b4Ppd{BqdiCjIHTvd4Uh\GWyZX7k[Y51KG2jbn7ldi=> M4nzdlI3Ojd6OEG5
HepG2  NVrafHBvTnWwY4Tpc44hSXO|YYm= NYHndJdkPS9zMDFOwG0> M1\1bFI1KGh? NETUUZVlemGvYYTpZ4FtdHliaX7obYJqfHNicHjvd5Bpd3K7bHH0bY9vKG:oIFHLWEBifCCVZYKtOFc{ MUWyOlI4QDhzOR?=
HepG2  MnnBSpVv[3Srb36gRZN{[Xl? MXi1M|ExKM7:TR?= MorpNlQhcA>? M{jGbIRwf26{ZXf1cIF1\XNidHjlJIxmfmWuczDITWYy|rFiYX7kJIN6[2yrbjDENeKh MXKyOlI4QDhzOR?=
HepG2  M2rQc2Z2dmO2aX;uJGF{e2G7 NWXFb2pSOC5z4pETOVDDqM7:TR?= Mmn6NlQhcA>? NH20fXFqdmS3Y3XzJJA3OiCmb4fudoVofWyjdHnvckwhSmWlbHnuMVEh\XiycnXzd4lwdiCjbnSgUGM{Si2LIITvJGxEO0JvSVmgZ49vfmW{c3nvckBqdiCjIHTvd4Uh\GWyZX7k[Y51KG2jbn7ldi=> MnvyNlYzPzh6MUm=
HepG3  NY\v[IF1TnWwY4Tpc44hSXO|YYm= NH34VJIxNjIkgKO1NOKh|ryP MYK0PEBp NFTweZpqdmS3Y3XzJINmdGxiYYX0c5Bp[We7IHnuJIEh\G:|ZTDk[ZBmdmSnboSgcYFvdmW{ MlHGNlYzPzh6MUm=
AGS M4i4ZWdzd3e2aDDJcohq[mm2aX;uJGF{e2G7 NVrQPYx1UUN3ME2xOU4xKMLzIEKuPVEh|ryP MkDDNlQ2QTd2N{i=
HGC27 MVnHdo94fGhiSX7obYJqfGmxbjDBd5NigQ>? M3qxeWlEPTB;MUWuNEDDuSB2LkiyJO69VQ>? NEHvXmIzPDV7N{S3PC=>
MKN45 MlW2S5Jwf3SqIFnubIljcXSrb36gRZN{[Xl? NUjyZopuUUN3ME2wMlgzKMLzIECuNFEh|ryP MUeyOFU6PzR5OB?=
NUGC4 M{XuUGdzd3e2aDDJcohq[mm2aX;uJGF{e2G7 M4X2VmlEPTB;Mj65N{DDuSByLkOxJO69VQ>? NYPFSJZtOjR3OUe0O|g>
PC9 M1ewdGdzd3e2aDDJcohq[mm2aX;uJGF{e2G7 MX23NkBp M{jLUWlEPTB;MUCuNVXDuTBwNkKgcm0> NFrR[IczOzh5NEi4NC=>
PC9GR MlrFS5Jwf3SqIFnubIljcXSrb36gRZN{[Xl? MlfYO|IhcA>? NVjC[49PUUN3ME22MlIyyrFzLkOwJI5O NHnNS3gzOzh5NEi4NC=>
H1650 M2XLO2dzd3e2aDDJcohq[mm2aX;uJGF{e2G7 MWi3NkBp MlPPTWM2OD15Lk[xxtExNjZ{IH7N NIDmRYkzOzh5NEi4NC=>
H1975 MnLsS5Jwf3SqIFnubIljcXSrb36gRZN{[Xl? MWi3NkBp MV3JR|UxRTFzLkG1xtExNjl|IH7N MWWyN|g4PDh6MB?=
PC9 MVPGeY5kfGmxbjDBd5NigQ>? MUixNE4yPSCwTR?= NVf1UpljPzJiaB?= MWrpcohq[mm2czDtWG9TKHCqb4PwbI9zgWyjdHnvckB{fGG2dYRCpC=> M2Tn[|I{QDd2OEiw
PC9GR MoXoSpVv[3Srb36gRZN{[Xl? NWjoW5hNPi5{MTDuUS=> MXO3NkBp MojNbY5pcWKrdIOgcXRQWiCyaH;zdIhwenmuYYTpc44he3SjdIXzxsA> MlzZNlM5PzR6OEC=
H1650 M1flTGZ2dmO2aX;uJGF{e2G7 NIXHUFc4NjZzIH7N MVS3NkBp MUfpcohq[mm2czDtWG9TKHCqb4PwbI9zgWyjdHnvckB{fGG2dYRCpC=> Mk\ZNlM5PzR6OEC=
H1975 MVzGeY5kfGmxbjDBd5NigQ>? NYDkNZZkOTFwMUWgcm0> MWC3NkBp NH\1V5lqdmirYnn0d{BuXE:UIIDoc5NxcG:{eXzheIlwdiC|dHH0eZPDqA>? MWOyN|g4PDh6MB?=
PC9 MYjGeY5kfGmxbjDBd5NigQ>? NULOboR5OTBwMUWgcm0> M1TPclczKGh? MY\pcohq[mm2czDwbI9{eGixconsZZRqd25ib3[gdFcxWz[N NVztdWJOOjN6N{S4PFA>
PC9GR MmXjSpVv[3Srb36gRZN{[Xl? NV22WGVqPi5{MTDuUS=> MkPqO|IhcA>? NVjONYt7cW6qaXLpeJMheGixc4Doc5J6dGG2aX;uJI9nKHB5MGO2Ty=> MmLtNlM5PzR6OEC=
H1650 MmX5SpVv[3Srb36gRZN{[Xl? M3jk[FcvPjFibl2= M2DYXFczKGh? NGfCbJdqdmirYnn0d{BxcG:|cHjvdplt[XSrb36gc4YheDdyU{\L NXHydY5QOjN6N{S4PFA>
H1975 NHu1U5pHfW6ldHnvckBCe3OjeR?= M17DflEyNjF3IH7N NFHvW|E4OiCq MWrpcohq[mm2czDwbI9{eGixconsZZRqd25ib3[gdFcxWz[N M4HWSlI{QDd2OEiw
LNCaP NVm1SYR[S2WubDDWbYFjcWyrdImgRZN{[Xl? MlvxNE0yOCEQvF2= NFTtbWgzPCCqwrC= MmHn[IVkemWjc3XzJINmdGxidnnhZoltcXS7IHnuJIEh\G:|ZTDk[ZBmdmSnboSgcYFvdmW{ MlTLNlM5PDB4MEW=
PC-3 M3G4eGNmdGxiVnnhZoltcXS7IFHzd4F6 M3z6dFAuOTBizszN NGHHU2kzPCCqwrC= MUnk[YNz\WG|ZYOgZ4VtdCC4aXHibYxqfHliaX6gZUBld3OnIHTldIVv\GWwdDDtZY5v\XJ? MmfxNlM5PDB4MEW=
MDA-MB-468  MVrD[YxtKF[rYXLpcIl1gSCDc4PhfS=> MUSwMVExKM7:TR?= MkfJNlQhcMLi NGHtPZZl\WO{ZXHz[ZMh[2WubDD2bYFjcWyrdImgbY4h[SCmb4PlJIRmeGWwZHXueEBu[W6wZYK= NVXqUlhLOjN6NEC2NFU>
LNCaP NVf5d5BDTnWwY4Tpc44hSXO|YYm= MYeyNFDjiJN6MECgcm0> NIHQe2szPCCqwrC= MYjk[YNz\WG|ZYOgeIhmKHCqb4PwbI9zgWyjdHnvckBt\X[nbDDv[kBxPzCVNlugbY4h[SCmb4PlJIRmeGWwZHXueEBu[W6wZYK= NHe4VmkzOzh2ME[wOS=>
PC-3 MXXGeY5kfGmxbjDBd5NigQ>? MkTBNlAx6oDVOECwJI5O NW\VS3N{OjRiaNMg NE\HcZVl\WO{ZXHz[ZMhfGinIIDoc5NxcG:{eXzheIlwdiCuZY\lcEBw\iCyN{DTOmshcW5iYTDkc5NmKGSncHXu[IVvfCCvYX7u[ZI> M{\5fFI{QDRyNkC1
MDA-MB-468  MlzuSpVv[3Srb36gRZN{[Xl? Mni3NlAx6oDVOECwJI5O M4rDZVI1KGkEoB?= MoPS[IVkemWjc3XzJJRp\SCyaH;zdIhwenmuYYTpc44hdGW4ZXygc4YheDdyU{\LJIlvKGFiZH;z[UBl\XCnbnTlcpQhdWGwbnXy NVfjd285OjN6NEC2NFU>
Caki-1  M2HGUWZ2dmO2aX;uJGF{e2G7 NIjQUJMyODBvMkCwNEBvVQ>? NILiVnIyOC1zOECgcYlv MkTXSG1UVw>? M33HbIlvcGmkaYTzJIJwfGhibWTPVmMyKGGwZDDtWG9TSzJiYYOgbY5lcWOjdHXkJIJ6KHSqZTDk[YNz\WG|ZTDpckBxcG:|cHjvdplt[XSrb36gc4Yh\G:5boP0doVidSCnZn\lZ5RwenN? MVSyN|M1QTl6OR?=
786-O Mk\PSpVv[3Srb36gRZN{[Xl? NFPnR4gyODBvMkCwNEBvVQ>? MlfINVAuOThyIH3pci=> NX;xcHl7TE2VTx?= NUTVcpVMcW6qaXLpeJMh[m:2aDDtWG9TSzFiYX7kJI1VV1KFMjDhd{BqdmSrY3H0[YQh[nlidHjlJIRm[3KnYYPlJIlvKHCqb4PwbI9zgWyjdHnvckBw\iCmb4fud5Rz\WGvIHXm[oVkfG:{cx?= NILDRm0zOzN2OUm4PS=>
Caki-1  MVzD[YxtKF[rYXLpcIl1gSCDc4PhfS=> MU[zNFAuPDByMDDuUS=> MVKyOE06PiCq MXTEUXNQ M2TXd5N2eHC{ZYPz[ZMhfGinIHPlcIwhfmmjYnnsbZR6KGmwIHLveIghfGmvZTDhcoQh\G:|ZTDk[ZBmdmSnboSgcYFvdmW{ MXeyN|M1QTl6OR?=
786-O MmrUR4VtdCCYaXHibYxqfHliQYPzZZk> Mo\vN|AxNTRyMECgcm0> NU\2W|J{OjRvOU[gbC=> MXTEUXNQ M2HUZZN2eHC{ZYPz[ZMhfGinIHPlcIwhfmmjYnnsbZR6KGmwIHLveIghfGmvZTDhcoQh\G:|ZTDk[ZBmdmSnboSgcYFvdmW{ MV:yN|M1QTl6OR?=
Caki-1  M{O3cmZ2dmO2aX;uJGF{e2G7 MXyyJOK2VQ>? MU[3NkBp MV3EUXNQ MWTpcoR2[2W|IFexJINmdGxiY4njcIUh[XK{ZYP0JIFv\CCjdYTvdIhi\3l? NYDyeHNROjN|NEm5PFk>
786-O M1TzNGZ2dmO2aX;uJGF{e2G7 MoDtNkDDvU1? MVK3NkBp Ml7pSG1UVw>? MWrpcoR2[2W|IFexJINmdGxiY4njcIUh[XK{ZYP0JIFv\CCjdYTvdIhi\3l? NIO1dnkzOzN2OUm4PS=>

... 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(Data is from Selleck tests instead of citations):
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
in solvent
Synonyms N/A

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Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID