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Signaling and Regulation

Stimulated PI3K-AKT Signaling Mediated through Ligand or Radiation-Induced EGFR Depends Indirectly, but not Directly, on Constitutive K-Ras Activity

¹ Divison of Radiobiology and Molecular Environmental Research, Department of Radiation Oncology, University of Tübingen, Tübingen, Germany and ² Department of Radiation Oncology, University of Dresden, Dresden, Germany

Requests for reprints: H. Peter Rodemann, Division of Radiobiology and Molecular Environmental Research, Department of Radiation Oncology, Eberhard-Karls University Tuebingen, Roentgenweg 11, 72076 Tuebingen, Germany. Phone: 49-0-7071-29-85962; Fax: 49-0-7071-29-5900. E-mail: hans-peter.rodemann{at}uni-tuebingen.de

Previous results showed an inducible radiation sensitivity selectively observable for K-RAS–mutated cell lines as a function of epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor blockade of phosphatidylinositol 3-kinase (PI3K)-AKT signaling. Therefore, the role of K-Ras activity for a direct (i.e., through activation of PI3K by K-Ras) or an indirect stimulation of PI3K-AKT signaling (through K-Ras activity–dependent EGFR ligand production) was investigated by means of small interfering RNA and inhibitor approaches as well as ELISA measurements of EGFR ligand production. K-RAS_mt tumor cells presented a constitutively activated extracellular signal–regulated kinase-1/2 signaling, resulting in enhanced production and secretion of the EGFR ligand amphiregulin (AREG). Medium supernatants conditioned by K-RAS_mt tumor cells equally efficiently stimulated EGFR signaling into the PI3K-AKT and mitogen-activated protein kinase pathways. Knocking down K-Ras expression by specific small interfering RNA markedly affected autocrine production of AREG, but not PI3K-AKT signaling, after treatment of K-RAS–mutated or wild-type cells with EGFR ligands or exposure to ionizing radiation. These results indicate that PI3K-mediated activation of AKT in K-RAS_mt human tumor cells as a function of EGFR ligand or radiation stimulus is independent of a direct function of K-Ras enzyme activity but depends on a K-Ras–mediated enhanced production of EGFR ligands (i.e., most likely AREG) through up-regulated extracellular signal–regulated kinase-1/2 signaling. The data provide new differential insight into the importance of K-RAS mutation in the context of PI3K-AKT–mediated radioresistance of EGFR-overexpressing or EGFR-mutated tumors. (Mol Cancer Res 2007;5(8):863–72)