An Extracellular Signal–Regulated Kinase 2 Survival Pathway Mediates Resistance of Human Mesothelioma Cells to Asbestos-Induced Injury

Abstract

We hypothesized that normal human mesothelial cells acquire resistance to asbestos-induced toxicity via induction of one or more epidermal growth factor receptor (EGFR)-linked survival pathways (phosphoinositol-3-kinase/AKT/mammalian target of rapamycin and extracellular signal-regulated kinase [ERK] 1/2) during simian virus 40 (SV40) transformation and carcinogenesis. Both isolated HKNM-2 mesothelial cells and a telomerase-immortalized mesothelial line (LP9/TERT-1) were more sensitive to crocidolite asbestos toxicity than an SV40 Tag-immortalized mesothelial line (MET5A) and malignant mesothelioma cell lines (HMESO and PPM Mill). Whereas increases in phosphorylation of AKT (pAKT) were observed in MET5A cells in response to asbestos, LP9/TERT-1 cells exhibited dose-related decreases in pAKT levels. Pretreatment with an EGFR phosphorylation or mitogenactivated protein kinase kinase 1/2 inhibitor abrogated asbestosinduced phosphorylated ERK (pERK) 1/2 levels in both LP9/TERT-1 and MET5A cells as well as increases in pAKT levels in MET5A cells. Transient transfection of small interfering RNAs targeting ERK1, ERK2, or AKT revealed that ERK1/2 pathways were involved in cell death by asbestos in both cell lines. Asbestos-resistant HMESO or PPM Mill cells with high endogenous levels of ERKs or AKT did not show doseresponsive increases in pERK1/ERK1, pERK2/ERK2, or pAKT/AKT levels by asbestos. However, small hairpin ERK2 stable cell lines created from both malignant mesothelioma lines were more sensitive to asbestos toxicity than shERK1 and shControl lines, and exhibited unique, tumorspecific changes in endogenous cell death-related gene expression. Our results suggest that EGFR phosphorylation is causally linked to pERK and pAKT activation by asbestos in normal and SV40 Tag-immortalized human mesothelial cells. They also indicate that ERK2 plays a role in modulating asbestos toxicity by regulating genes critical to cell injury and survival that are differentially expressed in human mesotheliomas.

Cyclooxygenase-2, Epidermal Growth Factor Receptor, and Aromatase Signaling in Inflammation and Mesothelioma

“…For example, in malignant mesothelioma, asbestos fibers activate the EGFR causing the activation of extracellular signal-regulated kinases (ERK) downstream (15). Similarly, COX-2/PGE2 and its downstream effectors can be regulated independently of EGFR signaling.…”

“…Various studies suggest that receptor tyrosine kinase activation participates in the oncogenic progression of nonneoplastic mesothelial progenitor cells to malignant mesothelioma. Asbestos fibers interact with the external domain of the EGFR to trigger dimerization, activation, and increased EGFR mRNA and protein levels in rat and human SV40 immortalized mesothelial cells (15). Upregulated EGFR and resulting tyrosine phosphorylation leads to the activation of downstream mitogen-activated protein kinase (MAPK) signaling pathway associated with proliferation (42) and to the involvement of EGFR activation in mitogenicity and carcinogenesis induced by asbestos (43).…”

The Role of Cyclooxygenase-2, Epidermal Growth Factor Receptor and Aromatase in Malignant Mesothelioma

“…Although there is a crosstalk between EGFR and COX-2 in carcinogenesis it is important to stress that EGFR and its downstream effectors can be activated independently of COX-2/PGE2. For example, in MM, asbestos fibers activate the EGFR resulting in activation of extracellular signal regulated kinase downstream (Shukla et al, 2011). Similarly, COX-2/PGE2 and its downstream effectors can be regulated independently of EGFR signaling.…”

“…Various studies suggest that receptor tyrosine kinase activation participates in the oncogenic progression of non neoplastic mesothelial progenitor cells to malignant mesothelioma. Asbestos fiber interact with the external domain of the EGFR to cause dimerization, activation and increased EGFR mRNA and protein levels in rat and human SV-40 immortalized mesothelial cells (Shukla et al, 2011). Up-regulated EGFR and resulting tyrosine phosphorylation leads to the Ras activation which phosphorylates directly and activates Raf (Rapidly Accelerated Fibrosarcoma).…”

The Role of Inflammation in Development and Therapy of Malignant Mesothelioma

“…Hill et al (2003) reported that amosite asbestos causes pleural inflammation by increased secretion of Intercellular Adhesion Molecule-1 (ICAM-1), Monocyte Chemoattractant Protein-1 (MCP-1) and Macrophage Inhibitory Protein-2 (MIP-2) in pleural lavage fluid as well as in vitro mesothelial cell culture (Hill et al, 2003). Recent studies from our group have also shown that human mesothelial cells acquire resistance to asbestos-induced toxicity via induction of one or more Epidermal Growth Factor Receptor (EGFR)-linked survival pathways (PI3K/AKT/ERK1/2) during Simian Virus 40 (SV40) transformation and carcinogenesis (Shukla et al, 2011). Many cytokines and growth factors are shown to be implicated in asbestos-induced MM pathogenesis including Tumor Necrosis Factor alpha (TNF-α), Transforming Growth Factor Beta (TGF-β), Platelet Derived Growth Factor (PDGF), Insulin like Growth Factor (IGF) (Liu and Klominek, 2004), interleukin-6 (IL-6), interleukin-8 (IL-8) (Galffy et al, 1999), Vascular Endothelial Growth Factor (VEGF) (Strizzi et al, 2001) and Hepatocyte Growth Factor (HGF) (Cacciotti et al, 2001).…”