Abstract A150: Molecular mechanisms mediating the pharmacodynamic interactions between oxaliplatin (Ox) and epidermal growth factor receptor (EGFR) inhibitors in KRAS mutant colorectal cancer (CRC) cells.
Authors: Ogbagabriel Selam , Xie Jiansong , Juan Gloria , Begley C. Glenn , Radinsky Robert , Freeman Daniel J. 
Published: June 2012

Background: Platinum agents are a standard of care for treatment many cancers. Clinical data has shown that patients (pts) with CRC with mutant (MT) KRAS do not respond to EGFR inhibitors (Amado et al 2009). Furthermore, results from phase 3 clinical trials with erlotinib, cetuximab, or panitumumab have shown that pts whose tumors express MT KRAS have shorter progression-free survival and overall survival when an EGFR inhibitor is added to a platinum-containing regimen vs chemotherapy alone (Eberhard et al 2005, Bokemeyer et al 2011, Douillard et al 2010). However, KRAS status is not predictive of outcomes in pts receiving Ox- or irinotecan-containing regimens without an EGFR inhibitor (Richman et al 2009). This negative interaction has not been observed in pts whose tumors express either wild type (WT) KRAS receiving an EGFR inhibitor and chemotherapy or whose tumors express MT KRAS receiving an EGFR inhibitor with irinotecan (Peeters et al 2010, Van Cutsem et al 2009). Our goal was to gain a molecular mechanistic understanding of the negative interaction between EGFR inhibitors and Ox in KRAS MT CRC cells.

Methods: To investigate the negative interaction between Ox and EGFR inhibitors, isogenic MT and WT KRAS-expressing HCT116 CRC cells were treated with Ox, SN-38 (the active metabolite of irinotecan), panitumumab, or gefitinib as single agents or in combination for 72 hrs. Viability was measured using an ATPlite assay. To determine the cellular distribution of EGFR, cells were serum-starved, labeled with a fluorescent anti-EGFR mAb, and visualized by a confocal microscopy. To analyze the effects of single agent or combination treatment on downstream phosphorylation of the PI3K or MAPK pathways, cells were treated for 24 hrs and phospho-proteins were detected by Western blotting. To investigate whether negative interaction between Ox and EGFR inhibitors could be reversed, cells were treated with Ox and gefitinib in combination with inhibitors to MEK, PI3K or Src for 72 hrs.

Results: Treatment with gefitinib, but not panitumumab, reversed the anti-proliferative effects of Ox in the MT KRAS-expressing CRC cells vs Ox alone (p&lt;0.0005). Neither treatment reversed the anti-proliferative effects of Ox observed in the WT KRAS-expressing cells or SN-38 in either the MT or WT KRAS-expressing cells. In the MT KRAS-expressing cells, EGFR was predominantly expressed intracellularly; in contrast, distinct cell surface staining was observed in the W T-KRAS expressing cells. This difference correlated with the inability of panitumumab to reverse the anti-proliferative effects of Ox in the CRC cells. Combination treatment of MT KRAS CRC cells with Ox and gefitinib increased pAKT which was not observed with Ox alone. Treatment of the mutant KRAS-expressing cells with a MEK, PI3K or Src inhibitor reversed the negative interaction between Ox and gefitinib (p&lt;0.0005).

Conclusion: We developed a preclinical model to further understand the negative interaction between MT KRAS-expressing cancer cells, platinum agents, and EGFR inhibitors. Feedback though AKT may contribute to the increased resistance to Ox in KRAS MT cells. Inhibitors of MEK, PI3K, or Src reversed the negative interaction between the EGFR inhibitors and Ox in a mutant-KRAS genetic background.

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 A150.