1-Methyl-3-((4-(quinolin-4-yloxy)phenyl)amino)-1H-pyrazole-4-carboxamide derivatives as new rearranged during Transfection (RET) kinase inhibitors capable of suppressing resistant mutants in solvent-front regions.

REarranged during Transfection (RET) is a validated target for anticancer drug discovery and two selective RET inhibitors were approved by US FDA in 2020. However, acquired resistance mediated by secondary mutations in the solvent-front region of the kinase (e.g. G810C/S/R) becomes a major challenge for selective RET inhibitor therapies. Herein, we report a structure-based design of 1-methyl-3-((4-(quinolin-4-yloxy)phenyl)amino)-1H-pyrazole-4-carboxamide derivatives as new RET kinase inhibitors which are capable of suppressing the RETG810 C/R resistant mutants. One of the representative compounds, 8q, potently suppressed wild-type RET kinase with an IC50 value of 13.7 nM. It also strongly inhibited the proliferation of BaF3 cells stably expressing various oncogenic fusions of RET kinase with solvent-front mutations, e.g. CCDC6-RETG810C , CCDC6-RETG810R , KIF5B-RETG810C and KIF5B-RETG810R , with IC50 values of 15.4, 53.2, 54.2 and 120.0 nM, respectively. Furthermore, 8q dose-dependently inhibited the activation of RET and downstream signals and obviously triggered apoptosis in Ba/F3-CCDC6-RETG810 C/R cells. The compound also exhibited significant anti-tumor efficacy with a tumor growth inhibition (TGI) value of 66.9% at 30 mg/kg/day via i. p. in a Ba/F3-CCDC6-RETG810C xenograft mouse model. Compound 8q may be utilized as a lead compound for drug discovery combating acquired resistance against selective RET inhibitor therapies.