Abstract 956: Receptor tyrosine kinases, not KRAS, activate PI3K in KRAS mutant colorectal cancers
Authors: Ebi Hiromichi , Corcoran Ryan B. , Singh Anurag , Chen Zhao , Song Youngchul , Lifshits Eugene , Ryan David P. , Meyerhardt Jeffrey A. , Benes Cyril , Settleman Jeffrey , Cantley Lewis C. , Wong Kwok-Kin , Engelman Jeffrey A. 
Published: June 2012

Introduction: Therapies inhibiting receptor tyrosine kinases (RTKs) are effective when they lead to simultaneous downregulation of phosphoinositide 3-kinase (PI3K)-AKT and mitogen-activated protein kinase (MEK)-ERK signaling. Although cetuximab has demonstrated activity in KRAS wildtype cancers, it remains unknown if RTK inhibition alone can suppress one or both of these pathways in colorectal cancers, especially in cancers with KRAS mutations.

Experimental Procedures and Results: Colorectal cancer cells were treated with gefitinib (1 μM) or Cetuximab (10 μg/mL) to compare the consequences of EGFR inhibition on downstream signaling events between KRAS wt (n=6) and mt (n=8) cell lines. In KRAS wt cancers, either cetuximab or gefitinib led to loss of ERK phosphorylation, but had only a modest effect on PI3K signaling. However, in most KRAS mutant cancers, EGFR inhibition minimally impacted ERK signaling and had neglible effects on PI3K signaling. To determine if KRAS constitutively activates ERK and PI3K signaling in KRAS mutant colorectal cancers, we utilized shRNA in eight KRAS mutant cancer cell lines and abruptly turned off KRAS expression in an vivo transgenic model. In all models, the phosphorylation of ERK was markedly decreased by KRAS knockdown, but this was not associated with downregulation of PI3K signaling. To determine how PI3K was activated in KRAS mutant cancers, we immunoprecipitated the p85 regulatory subunit and identified that IRS proteins co-precipitated with PI3K in KRAS mutant cancer. These complexes were disrupted by an IGF-IR inhibitor, NVP-AEW541, but not by gefitinib, and accordingly, NVP-AEW541 led to loss of AKT phosphorylation. Of the eight KRAS mutant cell lines, we found p85/IRS-1 or p85/IRS-2 complexes in 7 cell lines. We also observed that a monoclonal anti-IGF-IR antibody R1507 (18 mg/kg twice a week) downregulated PI3K signaling in SW837 xenografts in vivo. Biochemical assessment of patient specimens with KRAS mutations also suggested that PI3K was regulated primarily by IGF-IR. Importantly, by combining RTK inhibitors with MEK inhibitors in KRAS mutant cancers, both the PI3K and ERK pathways were concomitantly downregulated leading to marked growth suppression and robust apoptosis. Furthermore, combining R1507 (18 mg/kg twice a week) with a MEK inhibitor (AZD6244 25 mg/kg twice daily) induced marked tumor regressions of a KRAS mutant colorectal cancer in vivo.

Conclusions: In this study, we provide evidence that KRAS does not drive PI3K signaling in KRAS mutant cancers, and demonstrate that specific RTK inhibition potently suppresses PI3K signaling in vitro and in vivo. These findings provide a framework for utilizing RTK inhibitors in the treatment of KRAS mutant colorectal cancers.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 956. doi:10.1158/1538-7445.AM2011-956