Radiation induced shedding of TGF alpha results in a prolonged period of refractoriness to further release of TGF alpha

MP Hagan, MD, PhD, and A Yacoub, PhD

Medical College of Virginia, VCU, Richmond, Virgina United States

Purpose: We have shown that through an ERK1/2 dependent mechanism TGF alpha is shed from DU145 prostate carcinoma cells in response to ionizing radiation. Once shed, soluble TGF alpha confers sustained ERK1/2 activation on unirradiated bystander cells, resulting in the up-regulation of several DNA repair proteins and enhanced tumor growth. Here, we characterize the radiation dose dependence of the TGF alpha release and its down-stream effects. Methods: Following 60Co gamma-irradiation of DU145 cells, TGF alpha release was measured using an ELISA assay. Expression of the DNA repair proteins XRCC1 and ERCC1 was determined by semi-quantitative RT-PCR and Western analyses. EGFR activation was inhibited by the tyrphostin AG1478. ERK1/2 activation was inhibited by the MEK1/2- specific inhibitor PD98059. Results: A single exposure to ionizing radiation results in a dose-dependent release of TGF alpha. TGF alpha stimulates EGFR-ERK1/2 signaling, resulting in the up-regulation of XRCC1 and ERCC1 in unirradiated cells and enhanced up-regulation in irradiated cells. Blockade of the EGFR by AG1478 eliminates the up-regulation of XRCC1 and ERCC1 by TGF alpha, and substantially reduces the up-regulation of both proteins by irradiation. Following irradiation cells are refractory to further stimulation by irradiation for 8-12 hr. Irradiation during this refractory period does not result in increased ERK1/2 activity or expression of XRCC1 and ERCC1. Conclusions: These results further characterize the role of TGF alpha as a mediator of radiation-induced EGFR activation. We provide here the first demonstration of a radio-refractory period for TGF alpha production. This period last for several hours during which additional irradiation fails to induce the expression of DNA repair proteins.

KEY WORDS: Transforming Growth Factor alpha (TGF-alpha), Epithelial Growth Factor Receptor(EGFR), DNA repair, X-ray cross complementing group enzyme 1 (XRCC1), Nucleotide excision repair cross-complementing group enzyme 1(ERCC1).

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Paper presented at the International Symposium on Predictive Oncology and Intervention Strategies; Paris, France; February 9 - 12, 2002; in the section on Novel Therapies, Part 1.