Discovery of 3-pyrazolyl-substituted pyrazolo[1,5-a]pyrimidine derivatives as potent TRK inhibitors to overcome clinically acquired resistance

Abstract

Several secondary tropomyosin receptor kinase (TRK) mutations located in the solvent front, xDFG, and gatekeeper regions, are a common cause of clinical resistance. Mutations in the xDFG motif in particular limit sensitivity to second-generation TRK inhibitors, which represent an unmet clinical need. We designed a series of 3-pyrazolyl-substituted pyrazolo[1,5-a]pyrimidine derivatives toward these secondary mutations using ring-opening and scaffold-hopping strategies. Compound 5n was the most potent, with IC₅₀ values of 2.3 nM, 0.4 nM, and 0.5 nM against TRKA^G667C, TRKA^F589L, and TRKA^G595R, compared to selitrectinib with IC₅₀ values of 12.6 nM, 5.8 nM, and 7.6 nM, respectively (approximately 5.4, 14.5, and 15.2-fold increases). Furthermore, 5n displayed favorable pharmacokinetic properties and satisfactory antitumor efficacy (tumor growth inhibition of 97% at 30 mg/kg and 73% at 100 mg/kg) in TRKA^WT and TRKA^G667C xenograft mouse models. Collectively, 5n is a promising TRK inhibitor lead compound for overcoming clinically acquired resistance to second-generation inhibitors, particularly for resistant tumors harboring the TRKA^G667C mutation in the xDFG motif.

Keywords: Clinical resistance; Ring-opening and Scaffold-hopping; Secondary mutations; TRK inhibitors.