Radiation-induced Release of Transforming Growth Factor α Activates the Epidermal Growth Factor Receptor and Mitogen-activated Protein Kinase Pathway in Carcinoma Cells, Leading to Increased Proliferation and Protection from Radiation-induced Cell Death

Abstract

Exposure of A431 squamous and MDA-MB-231 mammary carcinoma cells to ionizing radiation has been associated with short transient increases in epidermal growth factor receptor (EGFR) tyrosine phosphorylation and activation of the mitogen-activated protein kinase (MAPK) and c-Jun NH 2 -terminal kinase (JNK) pathways. Irradiation (2 Gy) of A431 and MDA-MB-231 cells caused immediate primary activations (0 -10 min) of the EGFR and the MAPK and JNK pathways, which were surprisingly followed by later prolonged secondary activations (90 -240 min). Primary and secondary activation of the EGFR was abolished by molecular inhibition of EGFR function. The primary and secondary activation of the MAPK pathway was abolished by molecular inhibition of either EGFR or Ras function. In contrast, molecular inhibition of EGFR function abolished the secondary but not the primary activation of the JNK pathway. Inhibition of tumor necrosis factor ␣ receptor function by use of neutralizing monoclonal antibodies blunted primary activation of the JNK pathway. Addition of a neutralizing monoclonal antibody versus transforming growth factor ␣ (TGF␣) had no effect on the primary activation of either the EGFR or the MAPK and JNK pathways after irradiation but abolished the secondary activation of EGFR, MAPK, and JNK. Irradiation of cells increased pro-TGF␣ cleavage 120 -180 min after exposure. In agreement with radiation-induced release of a soluble factor, activation of the EGFR and the MAPK and JNK pathways could be induced in nonirradiated cells by the transfer of media from irradiated cells 120 min after irradiation. The ability of the transferred media to cause MAPK and JNK activation was blocked when media were incubated with a neutralizing antibody to TGF␣. Thus radiation causes primary and secondary activation of the EGFR and the MAPK and JNK pathways in autocrine-regulated carcinoma cells. Secondary activation of the EGFR and the MAPK and JNK pathways is dependent on radiation-induced cleavage and autocrine action of TGF␣. Neutralization of TGF␣ function by an anti-TGF␣ antibody or inhibition of MAPK function by MEK1/2 inhibitors (PD98059 and U0126) radiosensitized A431 and MDA-MB-231 cells after irradiation in apoptosis, 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT), and clonogenic assays. These data demonstrate that disruption of the TGF␣-EGFR-MAPK signaling module represents a strategy to decrease carcinoma cell growth and survival after irradiation.© 1999 by The American Society for Cell Biology 2493 INTRODUCTIONIonizing radiation has been shown to activate multiple signaling pathways within cells in vitro, which can lead to either increased cell death or increased proliferation depending on the cell type, the radiation dose, and the culture conditions (Xia et al., 1995;Rosette and Karin, 1996;Santana et al., 1996;Chmura et al., 1997;Schmidt-Ullrich et al., 1997;Carter et al., 1998;Haimovitz-Friedman, 1998;Kavanagh et al., 1998). Recently, a novel cellular target for ionizing radiation has been shown to be the ...

Sequence-dependent effects of ZD1839 (‘Iressa’) in combination with cytotoxic treatment in human head and neck cancer

“…Previous studies have shown that stimulation of the EGFR signalling pathway leads to radioresistance both in vitro and in vivo (Wollman et al, 1994;Miyaguchi et al, 1998;Akimoto et al, 1999;Dent et al, 1999). Interestingly, in the present study, we observed a strong positive association between EGFR level and radiosensitivity, which is in agreement with previous results of Bonner et al (1994Bonner et al ( , 1998Bonner et al ( , 2000.…”

The influence of fractionated radiation therapy on plasma vascular endothelial growth factor (VEGF) concentration in dogs with spontaneous tumors and its impact on outcome

“…In vitro, a 2.5-fold increased VEGF concentration has been measured in lung squamous carcinoma cells exposed to a single fraction of 15 Gy [2]. This increase was measurable up to 24 h and it was dependent on the activation of the ERK cascade through activation of the EGFR [2,9]. The activation of the ERK pathway was dose dependent with increased activity after exposure to a low radiation dose (1 Gy) [4].…”

“…VEGF is involved in development and growth of a wide variety of different tumors [15,17]. VEGF is actively secreted from tumor cells and its soluble form (VEGF 165 ) is detectable in the blood compartment [9].…”

“…This is the first step in activating the ERK pathway, a mitogen activated protein kinase (MAPK) pathway. This pathway is mediating cell survival and upregulation of VEGF expression in tumor cells [4,9]. In human squamous carcinoma cells the activation was dependent on the radiation dose, with low doses (1 Gy) causing prolonged activation of the ERK pathway and higher doses (6 Gy) having a weaker activation [4].…”

Chenodeoxycholic acid stimulates the progression of human esophageal cancer cells: A possible mechanism of angiogenesis in patients with esophageal cancer

“…5 After phosphorylation by EGFR, cascades may transduce signals to the nucleus via downstream pathways, including the mitogen-activated protein kinase (MAPK) pathway. 6 Activation of receptor tyrosine kinases such as EGFR is recognized to have important roles in the development and progression of cancer. 7 MAPK may regulate cyclooxygenase (COX)-2 expression via both transcriptional and posttranscriptional processes.…”