Discovery of novel 2,4-diarylaminopyrimidine analogues as ALK and ROS1 dual inhibitors to overcome crizotinib-resistant mutants including G1202R.

Aiming to explore novel anaplastic lymphoma kinase (ALK) and proto-oncogene tyrosine-protein kinase ROS (ROS1) dual inhibitors to overcome crizotinib-resistant mutants, two series of 2,4-diarylaminopyrimidine (DAAP) analogues bearing thiazole or 1,2,3-triazole moieties were designed and synthesized based upon the cocrystal structure of ceritinib with ALK (PDB 4MKC) as well as the binding model of ceritinib with ALK. The cellular and enzymatic assays validated 34c (WY-135) as a promising ALK (IC = 1.4 nM) and ROS1 (IC = 1.1 nM) dual inhibitor superior to crizotinib and ceritinib. 34c showed significantly inhibitory activities on ALK-dependent cell lines KARPAS299 (IC = 21 nM) and H2228 (IC = 95 nM) as well as ROS1-positive cell line HCC78 (IC = 40 nM). In particular, 34c was potent against a variety of frequently observed crizotinib-resistant mutants, particularly the L1196M mutant (IC = 3.1 nM) identified as the "gatekeeper" mutation and the G1202R mutant (IC = 8.7 nM) which conferred resistance to all clinical stage ALK inhibitors. Furthermore, 34c was capable of inducing cell apoptosis and strongly inhibiting cellular ALK and ROS1 activity. In addition, the binding models of 34c with ALK, ALK and ALK provided structural bases for SARs observations.