Abstract A146: Targeting AKT signaling overcomes acquired resistance to cetuximab in non-small lung cancer cells.
Authors: Iida Mari , Campbell David A. , Brand Toni M. , Li Chunrong , Wheeler Deric L. 
Published: June 2012

The epidermal growth factor receptor (EGFR) is a central regulator of tumor progression in human cancers. Cetuximab is an anti-EGFR antibody that has been approved by FDA for the treatment of patients with head and neck squamous cell carcinoma and metastatic colorectal cancer. However, increasing evidence suggests that the vast majority of patients do not respond to cetuximab and those who initially respond subsequently acquire resistance. To determine how tumor cells acquire resistance to cetuximab we developed models of acquired resistance using the non-small cell lung cancer line NCI-H226. During investigations into the molecular mechanisms of acquired resistance we found that cetuximab-resistant clones manifested strong activation of HER3 through transphosphorylation by the EGFR and constitutive PI(3)K/AKT activity and ultimate escape from cetuximab therapy. To determine the extent of AKT signaling in cetuximab-resistant clones, we analyzed the activation of the AKT pathway using a human AKT phospho-antibody array, which measured the activity of 137 proteins in the AKT pathway. Results showed strong activation of several key AKT substrates including GSK3β, eIF4E, S6, IKKα, IRS-1, Bad, β-catenin and Raf1 in cetuximab-resistant clones. To determine if cetuximab-resistant clones had dependence on AKT signaling we utilized RNAi technology to knockdown AKT expression. The results demonstrated that loss of AKT signaling in cetuximab-resistant clones resulted in decreased proliferative potential. To determine if AKT inhibition could be a therapeutic approach to overcome cetuximab resistance we investigated the AKT inhibitor MK-2206, a recently developed allosteric inhibitor, in this model of acquired resistance to cetuximab. We treated cetuximab-resistant clones with 2.5 μM of MK2206 and interrogated the AKT pathway using the human AKT phospho-antibody array. Results from this antibody array showed that MK-2206 inhibited multiple downstream AKT targets including GSK3β, eIF4E, S6, IKKα, IRS-1, β-catenin and Raf1. We next measured the effects of MK2206 on cellular proliferation and apoptosis. We found that cetuximab-resistant clones treated with MK-2206 showed robust inhibition of cell proliferation and increased apoptosis as compared to vehicle controls. Furthermore, the combination of cetuximab and MK-2206 resulted in further decreases in proliferation and increased apoptosis than either drug alone. Thus, MK-2206 treatment of cetuximab-resistant clones re-sensitized resistant clones to cetuximab. This combinatorial treatment resulted in decreased phospho-AKT and phospho-MAPK, highlighting the importance of these two pathways in cetuximab-resistance cells. These results indicate that dual targeting of the EGFR and AKT might produce greater impact than either drug alone. Collectively, our findings indicate that AKT activation is an important pathway in acquired resistance to cetuximab and suggest a rationale for clinical strategies that investigate combinatorial therapy directed at both the EGFR and AKT in patients with acquired resistance to cetuximab.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr A146.