Inhibition of HER2 in HER2-amplified breast cancer has been remarkably successful clinically, as demonstrated by the efficacy of HER-kinase inhibitors and HER2-antibody treatments. Whilst resistance to HER2 inhibition is common in the metastatic setting, the specific programs downstream of HER2 driving resistance are not established. Through genomic profiling of 733 HER2-amplified breast cancers, we identify enrichment of somatic alterations that promote MEK/ERK signaling in metastatic tumors with shortened progression-free survival on anti-HER2 therapy. These mutations, including NF1 loss and ERBB2 activating mutations, are sufficient to mediate resistance to FDA-approved HER2 kinase inhibitors including tucatinib and neratinib. Moreover, resistant tumors lose AKT dependence while undergoing a dramatic sensitization to MEK/ERK inhibition. Mechanistically, this driver pathway switch is a result of MEK-dependent activation of CDK2 kinase. These results establish genetic activation of MAPK as a recurrent mechanism of anti-HER2 therapy resistance that may be effectively combated with MEK/ERK inhibitors.

Anti-HER2 therapies have markedly improved prognosis of HER2-positive breast cancer. However, different mechanisms play a role in treatment resistance. Here, we identified AXL overexpression as an essential mechanism of trastuzumab resistance. AXL orchestrates epithelial-to-mesenchymal transition and heterodimerizes with HER2, leading to activation of PI3K/AKT and MAPK pathways in a ligand-independent manner. Genetic depletion and pharmacological inhibition of AXL restored trastuzumab response in vitro and in vivo. AXL inhibitor plus trastuzumab achieved complete regression in trastuzumab-resistant patient-derived xenograft models. Moreover, AXL expression in HER2-positive primary tumors was able to predict prognosis. Data from the PAMELA trial showed a change in AXL expression during neoadjuvant dual HER2 blockade, supporting its role in resistance. Therefore, our study highlights the importance of targeting AXL in combination with anti-HER2 drugs across HER2-amplified breast cancer patients with high AXL expression. Furthermore, it unveils the potential value of AXL as a druggable prognostic biomarker in HER2-positive breast cancer.

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