Temsirolimus

Catalog No.S1044 Batch:S1044

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Technical Data

Formula

C56H87NO16
Molecular Weight 1030.29 CAS No. 162635-04-3
Solubility (25°C)* In vitro Ethanol 75 mg/mL (72.79 mM)
DMSO 67 mg/mL (65.03 mM)
Water Insoluble
* <1 mg/ml means slightly soluble or insoluble.
* 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.
* Room temperature shipping (Stability testing shows this product can be shipped without any cooling measures.)

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Biological Activity

Description Temsirolimus is a specific mTOR inhibitor with IC50 of 1.76 μM in a cell-free assay. This compound induces autophagy and apoptosis.
Targets
mTOR [1]
(Cell-free assay)
1.76 μM
In vitro In the absence of FKBP12, Temsirolimus potently inhibits mTOR kinase activity with IC50 of 1.76 μM, similar to that of rapamycin with IC50 of 1.74 μM. This compound treatment at nanomolar concentrations (10 nM to <5 μM) displays a modest and selective antiproliferative activity via FKBP12-dependent mechanism, but can completely inhibit the proliferation of a broad panel of tumor cells at low micromolar concentrations (5-15 μM), involving FKBP12-independent suppression of mTOR signaling. This chemical treatment at micromolar but not nanomolar concentrations (20 μM) causes a marked decline in global protein synthesis and disassembly of polyribosomes, accompanied by rapid increase in the phosphorylation of translation elongation factor eEF2 and the translation initiation factor eIF2A. [1] It inhibits the phosphorylation of ribosomal protein S6, more potently in PTEN-positive DU145 cells than in PTEN-negative PC-3 cells, and inhibits cell growth and clonogenic survival of both cells in a concentration-dependent manner. [2] This compound (100 ng/mL) potently inhibits proliferation and induces apoptosis in primary human lymphoblastic leukemia (ALL) cells. [3]
In vivo In the NOD/SCID xenograft models with human ALL, Temsirolimus treatment at 10 mg/kg/day produces a decrease in peripheral blood blasts and in splenomegaly [3] Administration of this compound (20 mg/kg i.p. 5 days/week) significantly delays the growth of DAOY xenografts by 160% after 1 week and 240% after 2 weeks, compared with controls. Single high-dose of this chemical (100 mg/kg i.p) treatment induces 37% regression of tumor volume within 1 week. This compound treatment for 2 weeks also delays the growth of rapamycin-resistant U251 xenografts by 148%. [4] Inhibition of mTOR by this agent improves performance on four different behavioral tasks and decreases aggregate formation in a mouse model of Huntington disease. [5] Administration of this compound induces significant dose-dependent, antitumor responses against subcutaneous growth of 8226, OPM-2, and U266 xenografts with ED50 of 20 mg/kg and 2 mg/kg for 8226 and OPM-2, respectively, which are associated with inhibited proliferation and angiogenesis, induction of apoptosis, and reduction in tumor cell size. [6]

Protocol (from reference)

Kinase Assay:

[1]
  • In vitro assay of mTOR catalytic activity

    The Flag-tagged wild-type human mTOR (Flag-mTOR) DNA constructs are transiently transfected into HEK293 cells. Protein extraction and purification of Flag-mTOR are carried out 48 hours later. In vitro kinase assays of purified Flag-mTOR in the presence of various concentrations of Temsirolimus without FKBP12 are performed in 96-well plate and detected by dissociation-enhanced lanthanide fluorescent immunoassay (DELFIA) using His6-S6K1 as the substrate. Enzymes is first diluted in kinase assay buffer (10 mM Hepes (pH 7.4), 50 mM NaCl, 50 mM β-glycerophosphate, 10 mM MnCl2, 0.5 mM DTT, 0.25 μM microcystin LR, and 100 μg/mL BSA). To each well, 12 μL of the diluted enzyme is mixed briefly with 0.5 μL of this compound. The kinase reaction is initiated by adding 12.5 μL kinase assay buffer containing ATP and His6-S6K to give a final reaction volume of 25 μL containing 800 ng/mL FLAG-mTOR, 100 μM ATP, and 1.25 μM His6-S6K. The reaction plate is incubated for 2 hours (linear at 1-6 hours) at room temperature with gentle shaking and then terminated by adding 25 μL Stop buffer (20 mM Hepes (pH 7.4), 20 mM EDTA, and 20 mM EGTA). The DELFIA detection of the phosphorylated (Thr-389) His6-S6K is performed at room temperature using a monoclonal anti-P(T389)-p70S6K antibody labeled with Europium-N1-ITC (Eu) (10.4 Eu per antibody). 45 μL of the terminated kinase reaction mixture is transferred to a MaxiSorp plate containing 55 μL PBS. The His6-S6K is allowed to attach for 2 hours after which the wells are aspirated and washed once with PBS. 100 μL of DELFIA buffer with 40 ng/mL Eu-P(T389)-S6K antibody is added. The antibody binding is continued for 1 hour with gentle agitation. The wells are then aspirated and washed four times with PBS containing 0.05% Tween 20 (PBST). 100 μL of DELFIA Enhancement solution is added to each well and the plates are read in a PerkinElmer Victor model plate reader.
Cell Assay:

[1]
  • Cell lines

    A549, H157, H460, H446, HCT116, HT29, SW480, DLD1, Caco2, LNCap, DU145, MDA468, MDA231, HEK293, and PC3-MM2

  • Concentrations

    Dissolved in DMSO, final concentrations ~20 μM

  • Incubation Time

    72 hours

  • Method

    Cells are exposed to various concentrations of Temsirolimus for 72 hours. After treatment, viable cell densities are determined by MTS dye conversion using CellTiter AQ assay kit.
Animal Study:

[4]
  • Animal Models

    Female athymic nude mice injected s.c. with DAOY, or U251 cells

  • Dosages

    20 mg/kg

  • Administration

    Injection daily 5 times per week

References

  • https://pubmed.ncbi.nlm.nih.gov/18413763/
  • https://pubmed.ncbi.nlm.nih.gov/15805283/
  • https://pubmed.ncbi.nlm.nih.gov/16195324/
  • https://pubmed.ncbi.nlm.nih.gov/11245461/
  • https://pubmed.ncbi.nlm.nih.gov/15146184/
  • https://pubmed.ncbi.nlm.nih.gov/15304393/

Customer Product Validation

mTOR inhibitors attenuate ganetespib-driven elevation of HSPs in multiple tumor cell types. A375 melanoma cells were treated with vehicle, ganetespib (25 nmol/L), BEZ235 (500 nmol/L), or temsirolimus (500 nmol/L), either alone or in combination, for 24 hours. The levels of HSP90α, HSP70, HSP27, and GAPDH were determined by immunoblotting.

Data from [ Mol Cancer Res , 2014 , 12, 703-13 ]

SCID mice with PC-9/Vec or PC-9/HGF tumors were administered 25 mg/kg erlotinib once daily for 4 days or 50 mg/kg temsirolimus once from day 8. Four hours after final erlotinib administration, tumors were harvested, and the relative levels of proteins in the tumor lysates were determined by western blotting.

Data from [ PLoS One , 2013 , 8, e62104 ]

SCID mice with PC-9/Vec or PC-9/HGF tumors were administered as described in Figure. Four hours after final administration of erlotinib, tumors were harvested, and tumor cell angiogenesis (CD31) were determined by immunohistochemistry.

Data from [ PLoS One , 2013 , 8, e62104 ]

<p>Breast cancer cells were pretreated with 100ng/ml EGF for 15 min and then treated with the indicated concentrations of Temsirolimus for 24 hours.</p><div><div> </div></div><p> </p>

, , Dr. Zhang of Tianjin Medical University

Selleck's Temsirolimus Has Been Cited by 110 Publications

Haploinsufficiency of miR-143 and miR-145 reveal targetable dependencies in resistant del(5q) myelodysplastic neoplasm [ Leukemia, 2025, 39(4):917-928] PubMed: 40000845
Multi-layer stratified oncology platform utilizing transcriptomics, prostate cancer organoids, and modeling of drug response [ J Exp Clin Cancer Res, 2025, 44(1):290] PubMed: 41094672
PI3K/AKT signaling mediates stress-inducible amyloid formation through c-Myc [ Cell Rep, 2025, 44(5):115617] PubMed: 40272983
Lactate shuttle between cytotrophoblast and syncytiotrophoblast in the placenta enhances ferroptosis resistance and maintains placental homeostasis: implications for early pregnancy loss [ Cell Commun Signal, 2025, 23(1):438] PubMed: 41088442
Molecular control of PDPNhi macrophage subset induction by ADAP as a host defense in sepsis [ JCI Insight, 2025, e186456] PubMed: 39903516
A high-throughput screening platform to identify MYCN expression inhibitors for liver cancer therapy [ Front Oncol, 2025, 15:1486671] PubMed: 40027135
Single-Molecule-Based, Label-Free Monitoring of Molecular Glue Efficacies for Promoting Protein-Protein Interactions Using YaxAB Nanopores [ ACS Nano, 2024, 18(45):31451-31465] PubMed: 39482865
The mTOR pathway controls phosphorylation of BRAF at T401 [ Cell Commun Signal, 2024, 22(1):428] PubMed: 39223665
Patient-derived rhabdomyosarcoma cells recapitulate the genetic and transcriptomic landscapes of primary tumors [ iScience, 2024, 27(10):110862] PubMed: 39319271
Enzymatic depletion of circulating glutamine is immunosuppressive in cancers [ iScience, 2024, 27(6):109817] PubMed: 38770139

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