The Q61H mutation decouples KRAS from upstream regulation and renders cancer cells resistant to SHP2 inhibitors

Jeffrey E Lee,Mitsuhiko Ikura,Yoshihito Kano,Michael Ohh,Ahmet Mentes,Zhong-Yin Zhang,Richard Huang,Christopher B Marshall,Jonathan St-Germain,Molly L Udaskin,Claire M Robinson,Brian Raught,Teklab Gebregiworgis,Wenguang He,Jen Jen Yeh,Betty Poon ,Ivette Valencia-Sama,Meredith S Irwin,Nikolina Radulovich,Jiangyong Miao ,Melissa Huestis,Ming‐Sound Tsao

doi:10.1038/s41467-021-26526-y

Abstract

Cancer cells bearing distinct KRAS mutations exhibit variable sensitivity to SHP2 inhibitors (SHP2i). Here we show that cells harboring KRAS Q61H are uniquely resistant to SHP2i, and investigate the underlying mechanisms using biophysics, molecular dynamics, and cell-based approaches. Q61H mutation impairs intrinsic and GAP-mediated GTP hydrolysis, and impedes activation by SOS1, but does not alter tyrosyl phosphorylation. Wild-type and Q61H-mutant KRAS are both phosphorylated by Src on Tyr32 and Tyr64 and dephosphorylated by SHP2, however, SHP2i does not reduce ERK phosphorylation in KRAS Q61H cells. Phosphorylation of wild-type and Gly12-mutant KRAS, which are associated with sensitivity to SHP2i, confers resistance to regulation by GAP and GEF activities and impairs binding to RAF, whereas the near-complete GAP/GEF-resistance of KRAS Q61H remains unaltered, and high-affinity RAF interaction is retained. SHP2 can stimulate KRAS signaling by modulating GEF/GAP activities and dephosphorylating KRAS, processes that fail to regulate signaling of the Q61H mutant.

Highlights

Cancer cells bearing distinct KRAS mutations exhibit variable sensitivity to SHP2 inhibitors (SHP2i)
We and others have shown that dampening KRAS signaling by targeting SHP2 may provide a tractable strategy for the treatment of cancers driven by the major oncogenic KRAS mutants, including non-small-cell lung cancer (NSCLC), gastroesophageal cancer, and PDAC17–22
We observed that pancreatic ductal adenocarcinoma (PDAC) and PDAC patient-derived xenograft (PDX) cell lines harboring the KRAS Q61H mutation are less sensitive to the SHP2 catalytic inhibitor 11a-137, as well as the allosteric inhibitor SHP09938, in comparison to cells harboring mutations at the hotspot codon 12 (G12D/ V/C/R) (Fig. 1b and Supplementary Fig. 1)

Summary

Introduction

Cancer cells bearing distinct KRAS mutations exhibit variable sensitivity to SHP2 inhibitors (SHP2i). Phosphorylation of wild-type and Gly12-mutant KRAS, which are associated with sensitivity to SHP2i, confers resistance to regulation by GAP and GEF activities and impairs binding to RAF, whereas the near-complete GAP/GEF-resistance of KRAS Q61H remains unaltered, and highaffinity RAF interaction is retained. In-depth mutationspecific biochemical characterization of mutant RAS proteins is of paramount importance to understand the underlying mechanisms of pathogenesis, identify specific mutation-dependent therapeutic approaches, and, importantly, to identify patients who are likely to benefit from personalized or precision medicines[14,15]. We recently presented a model that demonstrates a direct catalytic role of SHP2 in reversing Src phosphorylation of KRAS This model adds an additional layer to previously proposed roles of SHP2 in RAS-MAPK signaling and provides a molecular mechanism by which SHP2 inhibition prevents cell growth

Methods

Results

Conclusion