Capmatinib

Capmatinib (INCB28060) is an orally active, highly selective, ATP-competitive c-Met tyrosine kinase inhibitor with an IC50 value of 0.13 nM. Capmatinib effectively inhibits the proliferation and migration of c-Met-dependent tumor cells, induces apoptosis, and exhibits antitumor activity.

c-Met:0.13 nM

Methods: HUVEC and THP-1 cells were treated with Capmatinib at a concentration gradient (0–10 nM) for 24 hours, followed by CCK-8 assay to assess cell viability.
Results: Capmatinib at 0–5 nM showed no effect on cell viability in either cell line, while 10 nM Capmatinib significantly reduced cell viability. [1]
Methods: HUVECs were pretreated with Capmatinib (1, 2.5, 5 nM) and LPS (200 ng/mL) for 24 hours, then co-incubated with THP-1 cells for 30 minutes. THP-1 cells were labeled with a green fluorescent dye and co-incubated with CAP-pretreated HUVECs. Adherent THP-1 cells were counted under a microscope.
Results: LPS significantly enhanced THP-1 adhesion to HUVECs, and Capmatinib dose-dependently reversed this effect. [1]

Methods: Nude mice were used to establish a brain metastasis model via left ventricular injection of PC9-BrM cells (stably expressing GFP-luciferase). Following successful model establishment, mice received intraperitoneal injections of Capmatinib (6 mg/kg, five times weekly for 5 weeks), tail vein injections of anetumab (0.2 mg/kg, twice weekly for 5 weeks), or combined therapy.
Results: Capmatinib monotherapy significantly inhibited brain metastasis and prolonged mouse survival. Capmatinib + anetumab combination therapy demonstrated optimal efficacy, significantly extending mouse survival compared to the control group while exhibiting the least BBB leakage. [2]

c-Met Kinase Assay: The assay buffer contains 50 mM Tris-HCl, 10 mM MgCl2, 100 mM NaCl, 0.1 mg/ml BSA, 5 mM DTT, pH 7.8. For HTS 0.8 μL of 5 mM of INCB28060 dissolved in DMSO are dotted on 384-well plates. DMSO titration suggests that the maximum tolerated concentration of the solvent is 4%. To measure IC50s the INCB28060 plate is prepared by 3-fold and 11-point serial dilutions. 0.8 μL of INCB28060 in DMSO is transferred from INCB28060 plate to the assay plate. The final concentration of DMSO is 2%. Solutions of 8 nM unphosphorylated c-Met or 0.5 nM phosphorylated c-Met are prepared in assay buffer. A 1 mM stock solution of peptide substrate Biotin-EQEDEPEGDYFEWLE-amide dissolved in DMSO is diluted to 1 μM in assay buffer containing 400 μM ATP (unphosphorylated c-Met) or 160 uM ATP (phosphorylated c-Met). A 20 μL volume of enzyme solution (or assay buffer for the enzyme blank) is added to the appropriate wells in each plate and then 20 μL/well of substrate solution to initiate the reaction. The plate is protected from light and incubated at 25 °C for 90 minutes. The reaction is stopped by adding 20 μL of a solution containing 45 mM EDTA, 50 mM Tris-HCl, 50 mM NaCl, 0.4 mg/ml BSA, 200 nM SA-APC and 3 nM EUPy20. The plate is incubated for 15-30 minutes at room temperature and HTRF (homogenous time resolved fluorescence) is measured on a Perkin Elmer Fusion α-FP instrument. The HTRF program settings used are as follows: Primary excitation filter 330/30, Primary window: 200 uSec, Primary delay: 50 uSec, Number of flashes: 15, Well read time: 2000

H441 cells are seeded in RPMI-1640 medium containing 10% FBS and grown to complete confluence. Gaps are introduced by scraping cells with a P200 pipette tip. Cells are then stimulated with 50 ng/mL recombinant human HGF to induce migration across the gap in the presence of various concentrations of INCB28060. After an overnight incubation, representative photographs are taken and a semiqualitative assessment of inhibition of cell migration is conducted.(Only for Reference)

DMSO: 4.13 mg/mL (10.01 mM), Sonication is recommended.