Relationship of EGFR-independent activation of PI3K in KRAS wild-type colorectal cancers and cetuximab resistance.

Abstract

385 Background: Therapies targeting receptor tyrosine kinases (RTKs) are typically effective only when they lead to simultaneous inhibition of PI3K-AKT and MEK-ERK signaling. Cetuximab is a monoclonal antibody against EGFR that has been approved for treatment of metastatic colorectal cancer (CRC). Studies have suggested that its benefit is restricted to patients with KRAS wildtype (WT) CRC. However, even in KRAS WT CRC, the response rate to single-agent cetuximab is low, and the mechanistic basis for this is not well-understood. Methods: The ability of cetuximab to inhibit PI3K-AKT and MEK-ERK signaling in a panel of KRAS WT CRC cell lines was assessed by immunoblotting. Paired biopsies from CRC patients obtained pre-treatment and after 3 weeks of cetuximab were evaluated by immunohistochemistry and proteomic techniques to assess inhibition of PI3K activity. We preformed immunoprecipitations (IPs) of the regulatory subunit of PI3K from cell lines and primary CRCs to identify RTKs involved in PI3K activation. Cetuximab-based therapeutic combinations were evaluated in KRAS WT CRC cell lines and xenograft models. Results: Cetuximab led to effective inhibition of MEK-ERK signaling (>75% reduction in phospho-ERK), but led to incomplete inhibition of PI3K-AKT signaling (0-50% reduction in phospho-AKT) in all KRAS WT CRC cell lines tested. Analysis of paired biopsies from patients with KRAS WT CRC obtained pre-treatment and after 3 weeks of cetuximab demonstrated that the PI3K pathway remained active in many patients, despite cetuximab therapy. PI3K regulatory subunit IPs from cell lines and primary CRCs revealed multiple RTK inputs to PI3K, most notably from IGFIR. Combined treatment with cetuximab and an IGFIR inhibitor or a PI3K inhibitor led to improved efficacy in vitro and to tumor regressions in KRAS WT CRC xenograft models. Conclusions: Cetuximab fails to inhibit PI3K signaling in some KRAS WT CRCs. Additional RTKs other than EGFR play a role in PI3K activation in these cancers, which may contribute to cetuximab resistance. Combining cetuximab with targeted therapies directed against the PI3K pathway may lead to improved efficacy in KRAS WT CRC, and should be evaluated in future clinical trials.