Rapamycin (AY-22989)

Licensed by Pfizer Catalog No.S1039 Synonyms: Rapamune, Sirolimus, NSC-2260804

For research use only.

Rapamycin (AY-22989, Rapamune, Sirolimus, NSC-2260804) is a specific mTOR inhibitor with IC50 of ~0.1 nM HEK293 cells.

Rapamycin (AY-22989) Chemical Structure

CAS No. 53123-88-9

Selleck's Rapamycin (AY-22989) has been cited by 1562 publications

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Biological Activity

Description Rapamycin (AY-22989, Rapamune, Sirolimus, NSC-2260804) is a specific mTOR inhibitor with IC50 of ~0.1 nM HEK293 cells.
Targets
mTOR [1]
(HEK293 cells)
~0.1 nM
In vitro

Rapamycin inhibits endogenous mTOR activity in HEK293 cells with IC50 of ~0.1 nM, more potently than iRap and AP21967 with IC50 of ~5 nM and ~10 nM, respectively. [1] In Saccharomyces cerevisiae, Rapamycin treatment induces a severe G1/S cell cycle arrest and inhibition of translation initiation to levels below 20% of control. [2] Rapamycin significantly inhibits the cell viability of T98G and U87-MG in a dose-dependent manner with IC50 of 2 nM and 1 μM, respectively, while displaying little activity against U373-MG cells with IC50 of >25 μM despite the similar extent of the inhibition of mTOR signaling. Rapamycin (100 nM) induces G1 arrest and autophagy but not apoptosis in Rapamycin-sensitive U87-MG and T98G cells by inhibiting the function of mTOR. [3]

Cell Data
Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
KMS-12-PE NWS3TYZbT3Kxd4ToJGlvcGmkaYTpc44hSXO|YYm= M3ftdGlEPTB;NEmuOVMxOiEQvF2= M{fhUFxiKHSjcnfleF0oZ2KuYX7rK{BpemWoPTfoeJRxezpxL4f3e{5m[mlwYXOueYsw[2inbXLsM4NwdXCxdX7kY5JmeG:{dG;jZZJlN0OKRV3CUFQyOy9pPmPBUmdGWjxxYU6=
P31-FUJ M3XOVWdzd3e2aDDJcohq[mm2aX;uJGF{e2G7 Mlf6TWM2OD1|MT61N|c1KM7:TR?= NEnsTWY9[SC2YYLn[ZQ:L1:kbHHub{chcHKnZk2nbJR1eHN8Lz;3e5cv\WKrLnHjMpVsN2OqZX3icE9kd22yb4Xu[H9z\XCxcoTfZ4Fz\C:FSFXNRmw1OTNxJ{7TRW5ITVJ:L3G+
LOXIMVI MmL1S5Jwf3SqIFnubIljcXSrb36gRZN{[Xl? NYDDUGd6UUN3ME2yO{45ODVzIN88US=> NI\3flg9[SC2YYLn[ZQ:L1:kbHHub{chcHKnZk2nbJR1eHN8Lz;3e5cv\WKrLnHjMpVsN2OqZX3icE9kd22yb4Xu[H9z\XCxcoTfZ4Fz\C:FSFXNRmw1OTNxJ{7TRW5ITVJ:L3G+
SK-N-FI M3nsO2dzd3e2aDDJcohq[mm2aX;uJGF{e2G7 NXTtcHZVUUN3ME2yO{4{QDFzIN88US=> NXfpVFRxRGFidHHy[4V1RSehYnzhcosoKGi{ZX[9K4h1fHC|Oj:ve5d4NmWkaT7hZ{52cy:laHXtZoww[2:vcH;1coRgemWyb4L0Y4NiemRxQ1jFUWJNPDF|Lze+V2FPT0WUPD;hQi=>
RPMI-8866 MYTHdo94fGhiSX7obYJqfGmxbjDBd5NigQ>? MX7JR|UxRTJ4Lke0NkDPxE1? MlnXQIEhfGG{Z3X0QUdg[myjbnunJIhz\WZ;J3j0eJB{Qi9xd4f3MoVjcS6jYz71b{9kcGWvYnyvZ49ueG:3bnTfdoVxd3K2X3PhdoQwS0iHTVLMOFE{Nyd-U1HOS2VTRC:jPh?=
NCI-H2171 M{jxNWdzd3e2aDDJcohq[mm2aX;uJGF{e2G7 MXTJR|UxRTJ2LkK0PFUh|ryP NXe0VolsRGFidHHy[4V1RSehYnzhcosoKGi{ZX[9K4h1fHC|Oj:ve5d4NmWkaT7hZ{52cy:laHXtZoww[2:vcH;1coRgemWyb4L0Y4NiemRxQ1jFUWJNPDF|Lze+V2FPT0WUPD;hQi=>
RH-1 M37NdGdzd3e2aDDJcohq[mm2aX;uJGF{e2G7 NUjHW|VtUUN3ME2yN{44QDZ4IN88US=> NV\5[IZ1RGFidHHy[4V1RSehYnzhcosoKGi{ZX[9K4h1fHC|Oj:ve5d4NmWkaT7hZ{52cy:laHXtZoww[2:vcH;1coRgemWyb4L0Y4NiemRxQ1jFUWJNPDF|Lze+V2FPT0WUPD;hQi=>
NCI-H128 Ml7DS5Jwf3SqIFnubIljcXSrb36gRZN{[Xl? Mn\xTWM2OD1{Mz61PFU{KM7:TR?= MYq8ZUB1[XKpZYS9K39jdGGwazegbJJm\j1paIT0dJM7Ny:5d4eu[YJqNmGlLoXrM4Np\W2kbD;jc41xd3WwZG;y[ZBwenShY3Hy[E9EUEWPQly0NVMwLz6VQV7HSXI9N2F-
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HT-29 M{PKWmN6fG:2b4jpZ{BCe3OjeR?= NE\4d2wyOCCwTR?= NW\pbpJNPzJiaB?= MY\EUXNQ MkjWVI91\W62aXH0[ZMhPS2obIXvdo92emGlaXytbY5lfWOnZDDjfZRwfG:6aXPpeJk> M3PhUFxiKHSjcnfleF0oZ2KuYX7rK{BpemWoPTfoeJRxezpxL4D1Zo1m\C6wY3LpMo5tdS6waXiu[493NzJ2OUCwPFc{Lz5{NEmwNFg4OzxxYU6=
HT-29 NVzhe4Q1S3m2b4TvfIlkKEG|c3H5 MXSxNEBvVQ>? MkjZO|IhcA>? Mlj5SG1UVw>? NVvxO5hoWG:2ZX70bYF1\XNiZHnnbZRwgGmwLXnu[JVk\WRiY4n0c5RwgGmlaYT5 NH\nNZY9[SC2YYLn[ZQ:L1:kbHHub{chcHKnZk2nbJR1eHN8Lz;weYJu\WRwbnPibU5vdG1wbnnoModwfi9{NEmwNFg4Oyd-MkS5NFA5PzN:L3G+
HT-29 MVnDfZRwfG:6aXOgRZN{[Xl? MXexNEBvVQ>? NH7OcVI4OiCq Mn3nSG1UVw>? NV3hPJNFWG:2ZX70bYF1\XNiY3HtdJRwfGinY3nuMYlv\HWlZXSgZ5l1d3SxeHnjbZR6 M33VZVxiKHSjcnfleF0oZ2KuYX7rK{BpemWoPTfoeJRxezpxL4D1Zo1m\C6wY3LpMo5tdS6waXiu[493NzJ2OUCwPFc{Lz5{NEmwNFg4OzxxYU6=
HEK293 MXPGeY5kfGmxbjDBd5NigQ>? NUjVdJJqUUN3MDC9JFAvODByMTFOwG0> MUG8ZUB1[XKpZYS9K39jdGGwazegbJJm\j1paIT0dJM7Ny:ydXLt[YQvdmOkaT7ucI0vdmmqLnfvek8yPzN3MEm1N{c,OTd|NUC5OVM9N2F-
HEK293T, PBMCs MXrBcpRqfmm{YXygZYN1cX[rdImgZZN{[Xl? NEjVWWc3KGSjeYO= M2riXGVEPTBiPTCwMlAxODFizszN M2fCVlxiKHSjcnfleF0oZ2KuYX7rK{BpemWoPTfoeJRxezpxL4D1Zo1m\C6wY3LpMo5tdS6waXiu[493NzF5NEi1OVAyLz5zN{S4OVUxOTxxYU6=
HEK293T, PBMCs NHTtRplCdnSrdnnyZYwh[WO2aY\peJkh[XO|YYm= M1XLfFYh\GG7cx?= NWrUTJJkTUN3MDC9JFAvODBzMzFOwG0> NEHse3E9[SC2YYLn[ZQ:L1:kbHHub{chcHKnZk2nbJR1eHN8Lz;weYJu\WRwbnPibU5vdG1wbnnoModwfi9zN{S4OVUxOSd-MUe0PFU2ODF:L3G+
cells from thymus of normal BALB/c mice NH\RdVhHfW6ldHnvckBCe3OjeR?= M1n3OGlEPTBiPTCwMlAxOyEQvF2= MX:8ZUB1[XKpZYS9K39jdGGwazegbJJm\j1paIT0dJM7Ny:ydXLt[YQvdmOkaT7ucI0vdmmqLnfvek8yODB{MUm0PEc,OTByMkG5OFg9N2F-
PC3 M4TrWI1VV1JiaX7obYJqfGmxbjDhd5NigQ>? Mon3TWM2OCB;IECuNFEh|ryP M3PNW|xiKHSjcnfleF0oZ2KuYX7rK{BpemWoPTfoeJRxezpxL4D1Zo1m\C6wY3LpMo5tdS6waXiu[493NzJzOUe4Olg{Lz5{MUm3PFY5OzxxYU6=
PC3 Ml\LRY51cXC{b3zp[oVz[XSrdnWgZYN1cX[rdImgZZN{[Xl? MoPaTWM2OCB;IECuNFEh|ryP NHjGVIE9[SC2YYLn[ZQ:L1:kbHHub{chcHKnZk2nbJR1eHN8Lz;weYJu\WRwbnPibU5vdG1wbnnoModwfi9{MUm3PFY5Oyd-MkG5O|g3QDN:L3G+
HEK293 NGfWUmNHfW6ldHnvckBCe3OjeR?= NGDt[|A5KGh? NUnyW|BlTUN3MDC9JFAvODVizszN NVP1cIl3RGFidHHy[4V1RSehYnzhcosoKGi{ZX[9K4h1fHC|Oj:vdJVjdWWmLn7jZokvdmyvLn7pbE5od3ZxMkG1N|k{ODFpPkKxOVM6OzBzPD;hQi=>
HeLa NVjXfmhPS3m2b4TvfIlkcXS7IHHzd4F6 MVnJR|UxKD1iMD63O{DPxE1? MoTyQIEhfGG{Z3X0QUdg[myjbnunJIhz\WZ;J3j0eJB{Qi9xcIXicYVlNm6lYnmucoxuNm6raD7nc5YwOzByMUWwO|AoRjNyMEG1NFcxRC:jPh?=
HEK293 M2nBb2Z2dmO2aX;uJGF{e2G7 M3rab2lEPTBiPTCxMlEh|ryP MnzUQIEhfGG{Z3X0QUdg[myjbnunJIhz\WZ;J3j0eJB{Qi9xcIXicYVlNm6lYnmucoxuNm6raD7nc5YwOTR3M{C5NFcoRjF2NUOwPVA4RC:jPh?=
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T47D M1jwdWdzd3e2aDDpcohq[mm2aX;uJIF{e2G7 NHXxe5A4OiCq MnvKTWM2OCB;IEGuOVc3KM7:TR?= MkPkQIEhfGG{Z3X0QUdg[myjbnunJIhz\WZ;J3j0eJB{Qi9xcIXicYVlNm6lYnmucoxuNm6raD7nc5YwOjl|MEi4PVUoRjJ7M{C4PFk2RC:jPh?=
SiHa MkjnS5Jwf3SqIHnubIljcXSrb36gZZN{[Xl? Mn3HO|IhcA>? Mn7MTWM2OCB;IEGuO|U3KM7:TR?= MWO8ZUB1[XKpZYS9K39jdGGwazegbJJm\j1paIT0dJM7Ny:ydXLt[YQvdmOkaT7ucI0vdmmqLnfvek8zQTNyOEi5OUc,Ojl|MEi4PVU9N2F-
Lewis rat lymph node M3LIVWFvfGmycn;sbYZmemG2aY\lJIFkfGm4aYT5JIF{e2G7 MWnJR|UxKD1iMj62JO69VQ>? MUm8ZUB1[XKpZYS9K39jdGGwazegbJJm\j1paIT0dJM7Ny:ydXLt[YQvdmOkaT7ucI0vdmmqLnfvek8yPjF6NUi2OUc,OTZzOEW4OlU9N2F-
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PC3 MWHDfZRwfG:6aXPpeJkh[XO|YYm= NELmS2hGSzVyIE2gOFMvOSEQvF2= MlPDQIEhfGG{Z3X0QUdg[myjbnunJIhz\WZ;J3j0eJB{Qi9xcIXicYVlNm6lYnmucoxuNm6raD7nc5YwOjh5N{S0NlYoRjJ6N{e0OFI3RC:jPh?=
Assay
Methods Test Index PMID
Western blot p-mTOR(S2448)/mTOR 23991038
Growth inhibition assay Cell proliferation 30393233
Histomorphology Haematoxylin & Eosin 28418837
Immunofluorescence NeuN ; p62/Beclin 28418837 28819214
ELISA Type III collagen/Fibronectin 23364979
In vivo Treatment with Rapamycin in vivo specifically blocks targets known to be downstream of mTOR such as the phosphorylation and activation of p70S6K and the release of inhibition of eIF4E by PHAS-1/4E-BP1, leading to complete blockage of the hypertrophic increases in plantaris muscle weight and fibre size. [4] Short-term Rapamycin treatment, even at the lowest dose of 0.16 mg/kg, produces profound inhibition of p70S6K activity, which correlates with increased tumor cell death and necrosis of the Eker renal tumors. [5] Rapamycin inhibits metastatic tumor growth and angiogenesis in CT-26 xenograft models by reducing the production of VEGF and blockage of VEGF-induced endothelial cell signaling. [6] Rapamycin treatment at 4 mg/kg/day significantly reduces tumor growth of C6 xenografts, and tumor vascular permeability. [7]

Protocol (from reference)

Kinase Assay:

[1]

  • Immunoblotting for the mTOR kinase assay:

    HEK293 cells are plated at 2-2.5×105 cells/well of a 12-well plate and serum-starved for 24 hours in DMEM. Cells are treated with increasing concentrations of Rapamycin (0.05-50 nM) for 15 minutes at 37 °C. Serum is added to a final concentration of 20% for 30 minutes at 37 °C. Cells are lysed, and cell lysates are separated by SDS-PAGE. Resolved proteins are transferred to a polyvinylidene difluoride membrane and immunoblotted with a phosphospecific primary antibody against Thr-389 of p70 S6 kinase. Data are analyzed using ImageQuant and KaleidaGr

Cell Research:

[3]

  • Cell lines: U87-MG, T98G, and U373-MG
  • Concentrations: Dissolved in DMSO, final concentrations ~25 μM
  • Incubation Time: 72 hours
  • Method:

    Cells are exposed to various concentrations of Rapamycin for 72 hours. For the assessment of cell viability, cells are collected by trypsinization, stained with trypan blue, and the viable cells in each well are counted. For the determination of cell cycle, cells are trypsinized, fixed with 70% ethanol, and stained with propidium iodide using a flow cytometry reagent set. Samples are analyzed for DNA content using a FACScan flow cytometer and CellQuest software. For apoptosis detection, cells are stained with the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) technique using an ApopTag apoptosis detection kit. To detect the development of acidic vesicular organelles (AVO), cells are stained with acridine orange (1 μg/mL) for 15 minutes, and examined under a fluorescence microscope. To quantify the development of AVOs, cells are stained with acridine orange (1 μg/mL) for 15 minutes, removed from the plate with trypsin-EDTA, and analyzed using the FACScan flow cytometer and CellQuest software. To analyze the autophagic process, cells are incubated for 10 minutes with 0.05 mM monodansylcadaverine at 37 °C and are then observed under a fluorescence microscope.

Animal Research:

[7]

  • Animal Models: Athymic Nu/Nu mice inoculated subcutaneously with VEGF-A-expressing C6 rat glioma cells
  • Dosages: ~4 mg/kg/day
  • Administration: Injection i.p.

Solubility (25°C)

In vitro

In vivo

Add solvents to the product individually and in order
(Data is from Selleck tests instead of citations):
2% DMSO+30% PEG 300+5% Tween 80+ddH2O
For best results, use promptly after mixing.

5mg/mL

Chemical Information

Molecular Weight 914.18
Formula

C51H79NO13

CAS No. 53123-88-9
Storage 3 years -20°C powder
2 years -80°C in solvent
Smiles CC1CCC2CC(C(=CC=CC=CC(CC(C(=O)C(C(C(=CC(C(=O)CC(OC(=O)C3CCCCN3C(=O)C(=O)C1(O2)O)C(C)CC4CCC(C(C4)OC)O)C)C)O)OC)C)C)C)OC

In vivo Formulation Calculator (Clear solution)

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

mg/kg g μL

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% 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.

Molarity Calculator

Mass Concentration Volume Molecular Weight

Clinical Trial Information

NCT Number Recruitment Interventions Conditions Sponsor/Collaborators Start Date Phases
NCT05386914 Not yet recruiting Drug: Sirolimus Genetic Predisposition to Disease|Healthy Volunteers University of Missouri-Columbia July 2022 Phase 1
NCT04996719 Recruiting Drug: Rapamune Heart Failure Mayo Clinic May 24 2022 Phase 1
NCT05302791 Not yet recruiting Device: Muscle Cooling|Other: Resistance Exercise Exercise Training University of Nebraska May 2022 Not Applicable
NCT04989686 Withdrawn Other: Microsampling Immunosuppression Children''s Hospital of Philadelphia March 2022 --

(data from https://clinicaltrials.gov, updated on 2022-08-01)

Tech Support

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.

Handling Instructions

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