Pharmacological inhibition of EGFR tyrosine kinase affects ILK-mediated cellular radiosensitization in vitro

Purpose: Integrin-linked kinase ( ILK) mediates signals from beta integrins and links integrins to epidermal growth factor receptor ( EGFR). Previous studies have identified an antisurvival effect of ILK in irradiated cells. The aim of this study was to evaluate the role of EGFR tyrosine kinase ( tk) activity for ILK-mediated radiosensitization. Materials and methods: Human FaDu squamous cell carcinoma ( SCC) cells stably transfected with hyperactive ILK ( ILK-hk) and ILK(fl/fl) and ILK(-/-) mouse fibroblasts were treated with the pharmacological EGFR-tk inhibitor BIBX1382BS without or in combination with single doses of X-rays. Clonogenic radiation survival, protein expression and phosphorylation ( EGFR, v-akt murine thymoma viral oncogene homolog 1 ( Akt), p42/44 mitogen-activated protein kinase), DNA-double strand break ( DSB) repair measured by gamma H2AX foci, cell morphology and cell cycle distribution were examined. Results: Expression of ILK-hk or ILK(fl/fl) status resulted in significant radiosensitization relative to vector controls or ILK(-/-). Following BIBX1382BS, clonogenic survival of normal fibroblasts and vector controls remained unaffected while ILK-hk-related radiosensitization was significantly diminished. In contrast to BIBX1382BS, which did not affect DNA-DSB repair, ILK-hk-mediated radiosensitization was associated with reduced DNA-DSB repair. At 10 days after BIBX1382BS treatment, FaDu transfectants, in contrast to fibroblasts, showed reduced cell size, accumulation of G1 phase cells and reduced Akt-serine( S) 473 phosphorylation. Conclusions: Our findings confirm ILK as a cell type-independent antisurvival factor in irradiated cells, which actions in terms of radiosensitization critically depend on proper EGFR-tk activity.