Ensartinib (X-396) is a potent and orally active dual ALK/MET inhibitor for the treatment of ALK-positive non-small cell lung cancer (NSCLC).

ALK:13.4 nM

Ensartinib is a potent ALK inhibitor that exhibits strong antiproliferative activity in various EML4-ALK–positive NSCLC cell lines. In H3122 cells, Ensartinib (IC₅₀ = 13.4 nM) significantly suppressed proliferation and induced PARP cleavage and apoptosis. Ensartinib also blocked ALK autophosphorylation and downstream signaling via AKT and ERK, indicating robust ALK pathway inhibition[1].

In H3122 xenograft mouse models, oral administration of Ensartinib (50 or 100 mg/kg, once daily for 14 days) significantly inhibited tumor growth (p < 0.001), with complete tumor regression observed in some mice at the 100 mg/kg dose. Ensartinib demonstrated good oral bioavailability, potent ALK targeting in vivo, and low toxicity.[1].

For viability experiments, cells are seeded in 96-well plates at 25%-33% confluency and exposed to drugs.The human lung adenocarcinoma cell lines H3122 and H2228 are treated with Ensartinib (10, 30, 100, 300 and 1000 nM). SUDHL-1 lymphoma cells are treated with Ensartinib (5, 10, 30, 100 and 300 nM). SY5Y neuroblastoma cells are treated with Ensartinib (30, 100, 300 and 1000 nM). At 72 hours post Ensartinib addition, Cell Titer Blue Reagent is added and fluorescence is measured on a Spectramax spectrophotometer. All experimental points are set up in hextuplicate replicates and are performed at least two independent times. IC50s are calculated using GraphPad Prism version 5 for Windows. The curves are fit using a nonlinear regression model with a log (inhibitor) vs. response formula[1].