AREG mediates transient hepatocellular carcinoma resistance to receptor tyrosine kinase inhibitors.

Lenvatinib is a first-line therapy for hepatocellular carcinoma (HCC), yet resistance remains a major clinical challenge. Emerging studies suggest resistance evolves through dynamic multi-step processes. Here, using multi-timepoint and multi-omics sampling, we characterized the trajectory of lenvatinib resistance in HCC cells and identified three adaptive states marked by state transitions. Multi-omics analyses and functional validation identified amphiregulin (AREG) as a key transient mediator of resistance in our models. AREG is a sufficient and dominant ligand to promote cell survival through epidermal growth factor receptor (EGFR) activation. Pharmacological inhibition of the AREG-EGFR axis specifically targeted early-resistant cells. Furthermore, AREG upregulation was validated in additional replicate experiments and another HCC cell line. Mechanistically, a focal deletion on chromosome 3p may contribute to NF-κB activation and AREG expression. This study delineates a recurrent yet transient adaptive mechanism and suggests new therapeutic opportunities-such as combination therapies targeting transient resistance mechanisms-to prevent or overcome drug resistance.