Purpose: Identify and characterize novel combinations of sorafenib with anti-inflammatory painkillers to target difficult-to-treat RAS-mutant cancer.

Experimental Design: The cytotoxicity of acetylsalicylic acid (aspirin) in combination with the multikinase inhibitor sorafenib (Nexavar) was assessed in RAS-mutant cell lines in vitro. The underlying mechanism for the increased cytotoxicity was investigated using selective inhibitors and shRNA-mediated gene knockdown. In vitro results were confirmed in RAS-mutant xenograft mouse models in vivo.

Results: The addition of aspirin but not isobutylphenylpropanoic acid (ibruprofen) or celecoxib (Celebrex) significantly increased the in vitro cytotoxicity of sorafenib. Mechanistically, combined exposure resulted in increased BRAF/CRAF dimerization and the simultaneous hyperactivation of the AMPK and ERK pathways. Combining sorafenib with other AMPK activators, such as metformin or A769662, was not sufficient to decrease cell viability due to sole activation of the AMPK pathway. The cytotoxicity of sorafenib and aspirin was blocked by inhibition of the AMPK or ERK pathways through shRNA or via pharmacologic inhibitors of RAF (LY3009120), MEK (trametinib), or AMPK (compound C). The combination was found to be specific for RAS/RAF–mutant cells and had no significant effect in RAS/RAF–wild-type keratinocytes or melanoma cells. In vivo treatment of human xenografts in NSG mice with sorafenib and aspirin significantly reduced tumor volume compared with each single-agent treatment.

Conclusions: Combination sorafenib and aspirin exerts cytotoxicity against RAS/RAF–mutant cells by simultaneously affecting two independent pathways and represents a promising novel strategy for the treatment of RAS-mutant cancers.