In Vivo (Add solvents to the product individually and in order.)
Homogeneous suspension
CMC-NA
≥5mg/ml
Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
* <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.)
Preparing Stock Solutions
Biological Activity
Description
Zorifertinib (AZD3759) is a potent, orally active, CNS-penetrant EGFR inhibitor with IC50 values of 0.3 nM, 0.2 nM, and 0.2 nM for EGFR (WT), EGFR (L858R), and EGFR (exon 19Del), respectively. This compound is in Phase 1.
Targets
EGFR (L858R) (Cell-free assay)
EGFR (exon 19Del) (Cell-free assay)
EGFR (WT) (Cell-free assay)
0.2 nM
0.2 nM
0.3 nM
In vitro
In H3255 (L858R) cells, Zorifertinib (AZD3759) inhibits EGFR phosphorylation with an IC50 of 7.2 nM. This compound exhibited inhibitory effect on pEGFR pathway and proliferation of EGFR mutation derived cells PC-9 and H3255 with IC50s of 7.7 nM and 7 nM respectively, but no significant activity against proliferation of H838 cells.
In Vivo
Zorifertinib (AZD3759) shows good oral bioavailability in dogs and penetrates extensively into monkey brain. In a brain metastasis PC-9 (Exon19Del) model, this compound (15 mg/kg) causes significant dose-dependent antitumor efficacy.
IC50 determination of compounds against EGFR enzymes
The inhibition potency of Zorifertinib (AZD3759) against EGFR WT and mutant enzymes is assessed using CisBio homogenous time resolved fluorescence approach (HTRF, Cat No. 62TK0PEJ) according to manufacturer’s instruction. The final enzyme concentrations used in this assay are 0.1 nM, 0.03 nM, and 0.026 nM for EGFR wild type, L858R and Exon19Del, respectively, and 0.8 μM, 4 μM and 25 μM ATP, corresponding to the Km values of EGFR enzymes, are applied accordingly. In brief, 3 μL of ATP and 2 μM TK biotin-peptide substrate are incubated in the presence or absence of serially diluted compound at room temperature in 384-well Greiner white polystyrene assay plates. The reaction is initiated by addition of 3 μL kinase which could phosphorylate the substrate peptide, and the assay buffer contains 1 mM DTT, 5 mM MgCl2, 1 mM MnCl2, and 0.01% CHAPS. After 30 minutes incubation, the reaction is stopped by the addition of 6 μl detection reagent mix containing 250 nM Strep-XL665 and TK Ab Europium Cryptate diluted in detection buffer. The plates are incubated for 1 h before the fluorescence is then measured at 615 nm and 665 nm, respectively with excitation wavelength at 320 nm by EnVision Multilabel Reader from Perkin Elmer using standard HTRF settings. The calculated signal ratio of 665 nm/615 nm is proportional to the kinase activity. The concentration of this compound producing 50% inhibition of the respective kinase (IC50) is calculated using four-parameter logistic fit.
PC-9 (exon 19Del), H3255 (L858R) and H838 (wt) cells
Concentrations
~30 mM
Incubation Time
72 h
Method
Zorifertinib (AZD3759) cell proliferation is determined by MTS methods. Briefly, cells are seeded in 96-well plates (at a density to allow for logarithmic growth during the 72-hour assay) and incubated overnight at 37 °C and 5% CO2. Cells are then exposed to concentrations of this compound ranging from 30 to 0.0003 mM for 72 hours. For the MTS endpoint, cell proliferation is measured by the CellTiter AQueous Non-Radioactive Cell Proliferation Assay reagent in accordance with the manufacturer’s protocol. Absorbance is measured with a Tecan Ultra instrument. Predose measurements are made, and concentration needed to reduce the growth of treated cells to half that of untreated cells (GI50) values are determined using absorbance readings.
Sellecks Zorifertinib (AZD3759) Has Been Cited by 6 Publications
A Novel Network Pharmacology Strategy Based on the Universal Effectiveness-Common Mechanism of Medical Herbs Uncovers Therapeutic Targets in Traumatic Brain Injury
[ Drug Des Devel Ther, 2024, 18:1175-1188]
Aurora kinase blockade drives de novo addiction of cervical squamous cell carcinoma to druggable EGFR signalling
[ Oncogene, 2022, 10.1038/s41388-022-02256-3]
YH25448, an Irreversible EGFR-TKI with Potent Intracranial Activity in EGFR Mutant Non-Small Cell Lung Cancer
[ Clin Cancer Res, 2019, 25(8):2575-2587]
Brain distribution of a panel of EGFR inhibitors using cassette-dosing in wild-type and Abcb1/Abcg2 deficient mice
[Kim M Drug Metab Dispos, 2019, 10.1124/dmd.118.084210]
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SHIPPING AND STORAGE
Selleck products are transported at room temperature. If you receive the product at room temperature, please rest assured, the Selleck Quality Inspection Department has conducted experiments to verify that the normal temperature placement of one month will not affect the biological activity of powder products. After collecting, please store the product according to the requirements described in the datasheet. Most Selleck products are stable under the recommended conditions.
NOT FOR HUMAN, VETERINARY DIAGNOSTIC OR THERAPEUTIC USE.
