NTRK kinase domain mutations in cancer variably impact sensitivity to type I and type II inhibitors

Romel Somwar et al. find that cancer-causing NTRK gene fusions resistant to one form of inhibitor therapy can be resistant to other inhibitor types. Using molecular simulations, they show that some NTRK1 mutations resistant to the type I inhibitor larotrectinib are hypersensitive to the type II inhibitor altiratinib, potentially due to the introduction of a sulfur moiety in the kinase binding pocket. Tyrosine kinase domains dynamically fluctuate between two main structural forms that are referred to as type I (DFG-in) or type II (DFG-out) conformations. Comprehensive data comparing type I and type II inhibitors are currently lacking for NTRK fusion-driven cancers. Here we used a type II NTRK inhibitor, altiratinib, as a model compound to investigate its inhibitory potential for larotrectinib (type I inhibitor)-resistant mutations in NTRK. Our study shows that a subset of larotrectinib-resistant NTRK1 mutations (V573M, F589L and G667C) retains sensitivity to altiratinib, while the NTRK1(V573M) and xDFG motif NTRK1(G667C) mutations are highly sensitive to type II inhibitors, including altiratinib, cabozantinib and foretinib. Moreover, molecular modeling suggests that the introduction of a sulfur moiety in the binding pocket, via methionine or cysteine substitutions, specifically renders the mutant kinase hypersensitive to type II inhibitors. Future precision treatment strategies may benefit from selective targeting of these kinase mutants based on our findings.