Different sensitivities of four protein tyrosine kinase inhibitors towards drug-resistant RET mutations

The RET protein tyrosine kinase (PTK) is a clinically validated target of therapy in non-small cell lung cancer (NSCLC) and thyroid cancer. Mutations in the targeted PTKs is a mechanism of drug resistance in cancer therapy with tyrosine kinase inhibitors (TKIs). Mutation-sensitive secondary drugs have been used successfully to overcome acquired drug-resistance to the first line TKIs. Cabozantinib, lenvatinib, vandetanib, and nintedanib are FDA-approved multikinase TKIs with anti-RET activity. Using RET kinase-dependent BaF3/KIF5B-RET cells, we isolated thirteen mutations resistant to one of these TKIs. Cross-analysis of sensitivities of these four TKIs on these drug-resistant RET mutants and the RETM918T mutant, which is found in medullary thyroid carcinoma, revealed different TKI resistance-sensitivity profiles. In particular, the RETM918T mutant was resistant to cabozantinib, lenvatinib, and vandetanib but did not affect the potency of nintedanib. RETL881V was isolated as a vandetanib-resistant mutation. The RETL881V mutation also induced resistance to cabozantinib and lenvatinib but did not affect the nintedanib sensitivity. Examination of the RET-vandetanib co-crystal structure and the chemical structures of vandetanib and nintedanib suggested that a phenyl group in nintedanib, which corresponds to a methoxy group in vandetanib, may form hydrophobic interaction with the shorter side chain of Val881 and thus allows nintedanib to inhibit RETL881V. To test this possibility, we synthesized two nintedanib analogs (Compound 1 and Compound 2) without the phenyl group and tested their activities. The data showed that RETL881V was resistant to Compound 1 and Compound 2, supporting a role of the phenyl group of nintedanib in mediating inhibition of RETL881V. Taken together, we have identified 13 RET mutations that display different resistance-sensitivity profiles against four anti-RET TKIs. Moreover, nintedanib is effective in inhibiting RETM918T and RETL881V mutants that are resistant to the other three TKIs. Citation Format: Xuan Liu, Tao Shen, Qingling Huang, Teng Peng, Frank Hilberg, Jianfeng Cai, Blaine H. Mooers, Jie Wu. Different sensitivities of four protein tyrosine kinase inhibitors towards drug-resistant RET mutations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2117.