Structural insights into the potency and selectivity of covalent pan-FGFR inhibitors

Lingzhi Qu,Yongheng Chen,Xiaojuan Chen,Hudie Wei,Jun Li,Ming Guo,Zhuchu Chen,Longying Jiang,Sitong Yue,Shuyan Dai

doi:10.1038/s42004-021-00623-x

Lingzhi Qu, Yongheng Chen + Show 8 more

Open Access

https://doi.org/10.1038/s42004-021-00623-x

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Abstract

FIIN-2, TAS-120 (Futibatinib) and PRN1371 are highly potent pan-FGFR covalent inhibitors targeting the p-loop cysteine of FGFR proteins, of which TAS-120 and PRN1371 are currently in clinical trials. It is critical to analyze their target selectivity and their abilities to overcome gatekeeper mutations. In this study, we demonstrate that FIIN-2 and TAS-120 form covalent adducts with SRC, while PRN1371 does not. FIIN-2 and TAS-120 inhibit SRC and YES activities, while PRN1371 does not. Moreover, FIIN-2, TAS-120 and PRN1371 exhibit different potencies against different FGFR gatekeeper mutants. In addition, the co-crystal structures of SRC/FIIN-2, SRC/TAS-120 and FGFR4/PRN1371 complexes reveal structural basis for kinase targeting and gatekeeper mutations. Taken together, our study not only provides insight into the potency and selectivity of covalent pan-FGFR inhibitors, but also sheds light on the development of next-generation FGFR covalent inhibitors with high potency, high selectivity, and stronger ability to overcome gatekeeper mutations.

Highlights

FIIN-2, TAS-120 (Futibatinib) and PRN1371 are highly potent pan-fibroblast growth factor receptor (FGFR) covalent inhibitors targeting the p-loop cysteine of FGFR proteins, of which TAS-120 and PRN1371 are currently in clinical trials
To test whether FIIN-2, TAS-120 and PRN1371 form covalent protein-inhibitor adducts with SRC protein, we carried out mass spectrometry, using FGFR1 as a positive control
When the p-loop cysteine of SRC and FGFR1 proteins was mutated to alanine, the molecular weight of the SRC and FGFR1 proteins did not change after incubation with these inhibitors (Supplementary Fig. 1)

Summary

Introduction

FIIN-2, TAS-120 (Futibatinib) and PRN1371 are highly potent pan-FGFR covalent inhibitors targeting the p-loop cysteine of FGFR proteins, of which TAS-120 and PRN1371 are currently in clinical trials. The co-crystal structures of SRC/FIIN-2, SRC/TAS-120 and FGFR4/PRN1371 complexes reveal structural basis for kinase targeting and gatekeeper mutations. PRN1371 and TAS120 have entered into clinical investigation[20] These three inhibitors possess an electrophilic acrylamide group designed to undergo Michael addition reaction and achieve covalent binding with a p-loop cysteine of FGFRs (Fig. 1a, b). Out of 518 human kinases, only 9 kinases share this p-loop cysteine (Fig. 1a), suggesting that these inhibitors might be highly selective Among these 9 kinases, 4 of them (FGFR1-4) belong to FGFR family, and 3 of them (SRC, YES, FGR) belong to the SRC family kinases (SFKs). We determined crystal structures of SRC/FIIN-2, SRC/ TAS-120, and FGFR4/PRN1371 complexes with high resolution x-ray diffraction data These three crystal structures detail the binding modes of these inhibitors, and provide structural basis for kinase targeting and overcoming gatekeeper mutations

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Results

Conclusion