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Signal-transduction genes such as EGFR and RAS, which encode components of the MAPK pathway, are frequently over-expressed or mutated in a variety of cancers. A majority of colorectal cancers carry either a RAS or a BRAF mutation. The BRAF gene codes for a serine/threonine kinase just downstream from RAS that activates p42/44 ERK via phosphorylation. In some instances, ionizing radiation activates the EGFR via its phosphorylation, which is a major effector of the p42/44 mitogen-activated protein kinase (MAPK/ERK) pathway. This pathway has been proposed to be involved in cell proliferation or repopulation after radiation. We wished to understand the significance of the EGFR and MAPK pathways in radiation-induced cell death and/or survival. For this purpose we chose a panel of cell lines including SW620 colorectal cell line, MDA-MB-468 breast carcinoma, A431 epidermoid carcinoma, normal skin fibroblasts and ATM cells. SW620 cells do not express EGFR (absent by immunoblotting and RT-PCR), while both MDA-MB-468 and A-431 have EGFR amplification and overexpression. The surviving fractions of SW620 and MDA-MB-468 cells irradiated with 2 Gy were 0.37 ± 0.05 and 0.32 ± 0.06, respectively. Cells were irradiated with 2-8 Gy. Phospho p42/44-ERK levels were assessed by immunoblotting at various time interval post irradiation. We found that p42/44 ERK levels were decreased in a dose dependent manner relative to control within 5 min of irradiation in SW620, MDA-MB-468 and fibroblasts. In these cells the p42/44 ERK levels recovered by 2h at 2 or 4 Gy dose levels. However we found as reported before, that in confluent A-431 cells the p42/44 ERK levels were increased within 2 min post irradiation, the levels returned to baseline by 20 min. In contrast the pERK activity was not affected in ATM cells. These results demonstrate that p42/44 ERK can be modulated by ionizing radiation, but that this effect is independent of EGFR. These findings suggest that it will be critical to measure downstream effects, rather than simply EGFR presence or activation, in determining the effect of radiation on signal transduction.

[Proc Amer Assoc Cancer Res, Volume 45, 2004]