ARTICLE ABSTRACTPurpose: In colorectal cancer, the activation of the intracellular RAS–RAF and PIK3CA–AKT pathways has been implicated in the resistance to anti-EGFR mAbs. We have investigated the role of regorafenib, an oral multikinase inhibitor, in combination with cetuximab, an anti-EGFR mAb, to overcome anti-EGFR resistance.Experimental Design: We have tested, in vitro and in vivo, the effects of regorafenib in a panel of human colorectal cancer cell lines with a KRAS mutation (SW480, SW620, HCT116, LOVO, and HCT15) or with a BRAF mutation (HT29), as models of intrinsic resistance to cetuximab treatment, and in two human colorectal cancer cell lines (GEO and SW48) that are cetuximab-sensitive, as well as in their derived cells with acquired resistance to cetuximab (GEO-CR and SW48-CR).Results: Treatment with regorafenib determined a dose-dependent growth inhibition in all colorectal cancer cell lines. The combined treatment with cetuximab and regorafenib induced synergistic antiproliferative and apoptotic effects in cetuximab-resistant cell lines by blocking MAPK and AKT pathways. Nude mice were injected s.c. with HCT116, HCT15, GEO-CR, and SW48-CR cells. The combined treatment caused significant tumor growth inhibition. Synergistic antitumor activity of regorafenib plus cetuximab was also observed in an orthotopic colorectal cancer model of HCT116 cells. In particular, the combined treatment induced a significant tumor growth inhibition in the primary tumor site (cecum) and completely prevented metastasis formation.Conclusions: The combined treatment with cetuximab and regorafenib could be a strategy to overcome resistance to anti-EGFR therapies in metastatic colorectal cancer patients. Clin Cancer Res; 21(13); 2975–83. ©2015 AACR.