Oncogenic c-ros oncogene1 (ROS1) fusion kinases have been identified in a 
variety of human cancers and are attractive targets for cancer therapy. The 
MET/ALK/ROS1 inhibitor crizotinib (Xalkori, PF-02341066) has demonstrated 
promising clinical activity in ROS1 fusion-positive non-small cell lung cancer. 
However, emerging clinical evidence has shown that patients can develop 
resistance by acquiring secondary point mutations in ROS1 kinase. In this study 
we characterized the ROS1 activity of PF-06463922, a novel, orally available, 
CNS-penetrant, ATP-competitive small-molecule inhibitor of ALK/ROS1. In vitro, 
PF-06463922 exhibited subnanomolar cellular potency against oncogenic ROS1 
fusions and inhibited the crizotinib-refractory ROS1(G2032R) mutation and the 
ROS1(G2026M) gatekeeper mutation. Compared with crizotinib and the 
second-generation ALK/ROS1 inhibitors ceritinib and alectinib, PF-06463922 
showed significantly improved inhibitory activity against ROS1 kinase. A crystal 
structure of the PF-06463922-ROS1 kinase complex revealed favorable interactions 
contributing to the high-affinity binding. In vivo, PF-06463922 showed marked 
antitumor activity in tumor models expressing FIG-ROS1, CD74-ROS1, and the 
CD74-ROS1(G2032R) mutation. Furthermore, PF-06463922 demonstrated antitumor 
activity in a genetically engineered mouse model of FIG-ROS1 glioblastoma. Taken 
together, our results indicate that PF-06463922 has potential for treating ROS1 
fusion-positive cancers, including those requiring agents with CNS-penetrating 
properties, as well as for overcoming crizotinib resistance driven by ROS1 
mutation.