Molecular Weight (MW)

469.55

Formula

C30H23N5O

CAS No.

915019-65-7

Storage

-20℃ for 3 years in powder form

-80℃ for 2 years in solvent

Solubility (In vitro)

DMSO: 0.01 mg/mL (0.02 mM)

Water:<1 mg/mL (slightly soluble or insoluble)

DMF: 18 mg/mL warming (38.33 mM)

Solubility (In vivo)

NMP+polyethylene glycol 300 (10/90, v/v): 30 mg/mL

Synonym/Chemical Name

BEZ235; NVP BEZ235; BEZ 235; NVP-BEZ235; BEZ-235; NVP-BEZ-235; NVP-BEZ 235; 2-methyl-2-[4-(3-methyl-2-oxo-8-quinolin-3-ylimidazo[4,5-c]quinolin-1-yl)phenyl]propanenitrile

Kinase Assay: PI3Kα, β, and δ proteins are composed of the iSH2 domain of p85 NH2-terminally fused to the full-length protein p110 protein, with the exception of α that also does not contain the last 20 amino acids. PI3Kγ is produced as full-length protein deleted for its first 144 amino acids. All constructs are fused to a COOH-terminal His tag for convenient purification and then cloned into the pBlue-Bac4.5 (for α, β, and δ isoforms) or pVL1393 (for γ isoform) plasmids. The different vectors are then cotransfected with BaculoGold WT genomic DNA using methods recommended by the vendor for production of the respective recombinant baculoviruses and proteins. BEZ235 are tested for their activity against PI3K using a Kinase-Glo assay. The kinase reaction is done in 384-well black plate. Each well is loaded with 50 μL of test items (in 90% DMSO) and 5 μL reaction buffer containing 10 μg/mL PI substrate (l-α-phosphatidylinositol; Avanti Polar Lipids; prepared in 3% octyl-glucoside) and the PI3K proteins (10, 25, 10, and 150 nM of p110α, p110β, p110δ, and p110γ, respectively) are then added to it. The reaction is started by the addition of 5 μL of 1 μM ATP prepared in the reaction buffer and is incubated for either 60 (for p110α, p110β, and p110δ) or 120 min (for p110γ). It is terminated by the addition of 10 μL Kinase-Glo buffer. The plates are then read in a Synergy 2 reader for luminescence detection.

Cell Assay: MOLT-4 and CEM-R cells; The solubility of this compound in DMSO is<10 mM. General tips for obtaining a higher concentration: Please warm the tube at 37 °C for 10 minutes and/or shake it in the ultrasonic bath for a while.Stock solution can be stored below -20°C for several months; 500 nM, for cell cycle inhibition 200 nM, 16 hours for pRb decrease.

BEZ235 significantly reduces the phosphorylation levels of the mTOR activated kinase p70S6K. BEZ235 results in a reduction of S235/S236P-RPS6 levels with IC50 of 6.5 nM. The activity of BEZ235 against mTOR is determined using a biochemical mTOR K-LISA assay with IC50 of 20.7 nM. BEZ235 shows slightly lower activity against its β paralogue with IC50 of 75 nM. The PI3K/Akt/mTOR pathway is often constitutively activated in human tumor cells. BEZ235 blocks PI3K and mTOR kinase activity by binding to the ATP-binding cleft of these enzymes. Both PTEN-null cell lines PC3M and U87MG show a dose-dependent reduction in cell proliferation when treated with increasing concentrations of BEZ235 with an average GI50 of 10-12 nM. BEZ235 is an mTORC1/2 catalytic inhibitor.

In Vivo

BEZ235 induces regression of the tumors (69%) without statistically significant effect on body weight gain. Altogether, these preliminary in vivo efficacy results show that BEZ235 causes disease stasis when administered orally as a single agent and can enhance the efficacy of other anticancer agents when used in combination studies.

Animal model

Female Harlan athymic nude mice

Formulation & Dosage

NMP/polyethylene glycol 300 (10/90, v/v); 45 mg/kg; oral gavage

References

Mol Cancer Ther, 2008, 7(7), 1851-1863.; PLoS One, 2011, 6(9), e25132.