ZDHHC5-Mediated BRAF Palmitoylation Drives KRASG12C-Inhibitor Resistance and Sustains Oncogenic MAPK Signalling

Therapeutic resistance to single-agent targeted therapies (e.g., KRASG12C inhibitors) frequently arises in tumors through adaptive mechanisms such as cellular plasticity and genetic instability. Here, we demonstrated that KRASG12C inhibitors induce BRAF membrane localization, leading to MAPK pathway reactivation in KRASG12C-mutant cancers. The engineered tethering of BRAF to the membrane significantly diminished the antitumour efficacy of the KRASG12C inhibitors. Mechanistically, ZDHHC5-mediated palmitoylation at residues Cys194/195, located within the RAS-binding domain (RBD) of BRAF, governs BRAF membrane recruitment and protein stability. Notably, BRAF palmitoylation sustains oncogenic signalling across diverse cancer types, especially in cholangiocarcinoma, and high ZDHHC5 expression is correlated with aggressive clinical behavior in KRASG12C-mutant and other MAPK-driven malignancies. Depletion of ZDHHC5 or disruption of BRAF palmitoylation at these critical sites suppressed MAPK signalling, restored sensitivity to KRASG12C inhibition, and impaired tumor progression in vitro and in vivo. Our findings identify BRAF palmitoylation as a dual regulator of therapeutic resistance and tumor progression, highlighting ZDHHC5 as a therapeutic target to overcome KRASG12C inhibitor resistance and treat BRAF palmitoylation-dependent cancers.