EGFR Activation Results in Enhanced Cyclooxygenase-2 Expression through p38 Mitogen-Activated Protein Kinase–Dependent Activation of the Sp1/Sp3 Transcription Factors in Human Gliomas

Abstract

Expression of cyclooxygenase-2 (COX-2) has been linked to many cancers and may contribute to malignant phenotypes, including enhanced proliferation, angiogenesis, and resistance to cytotoxic therapies. Malignant gliomas are highly aggressive brain tumors that display many of these characteristics. One prominent molecular abnormality discovered in these astrocytic brain tumors is alteration of epidermal growth factor (EGF) receptor (EGFR) through gene amplification and/or mutation resulting in excessive signaling from this receptor. We found that EGF-mediated stimulation of EGFR tyrosine kinase in human glioma cell lines induces expression of both COX-2 mRNA and protein. The p38 mitogen-activated protein kinase (p38-MAPK) pathway was a strong downstream factor in this activation with inhibition of this pathway leading to strong suppression of COX-2 induction. The p38-MAPK pathway can activate the Sp1/Sp3 transcription factors and this seems necessary for EGFRdependent transactivation of the COX-2 promoter. Analysis of COX-2 promoter/luciferase constructs revealed that transcriptional activation of the COX-2 promoter by EGFR requires the Sp1 binding site located at À245/À240. Furthermore, Sp1/Sp3 binding to this site in the promoter is enhanced by EGFR activation both in vitro and in vivo. Enhanced DNA binding by Sp1/Sp3 requires p38-MAPK activity and correlates with increased phosphorylation of the Sp1 transcription factor. Thus, EGFR activation in malignant gliomas can transcriptionally activate COX-2 expression in a process that requires p38-MAPK and Sp1/Sp3. Finally, treatment of glioma cell lines with prostaglandin E2, the predominant product of COX-2 activity, results in increased vascular endothelial growth factor expression, thus potentially linking elevations in COX-2 expression with tumor angiogenesis in malignant gliomas.

Prolidase Directly Binds and Activates Epidermal Growth Factor Receptor and Stimulates Downstream Signaling

“…It is also gratifying that we were able to show that not only could PEPD induce Cox-2 but this induction also depended on EGFR activation. Although the detailed mechanism by which PEPD induces Cox-2 remains to be elucidated, Xu and Shu (22) reported that EGFR activation in human glioma cells transcriptionally activated Cox-2 via p38 MAPK and Sp1/Sp3, and Lo et al (32) reported that nuclear EGFR bound together with STAT3 to the Cox-2 gene promoter and stimulated Cox-2 gene transcription in human glioma cells.…”

“…Interestingly, several previous studies have indicated that EGFR activation leads to transcriptional activation of Cox-2 (22)(23)(24). To find out if PEPD could bind to EGFR, cell lysates were prepared from Hepa1c1c7 and RT-4 cells, and each lysate (0.5 mg of protein) was incubated with the recombinant PEPD (20 g of protein) in a 0.5-ml volume and the mixture was then subjected to immunoprecipitation with anti-EGFR or an isotype-matched IgG, followed by Western blotting with anti-His (for detection of the recombinant PEPD).…”

Epidermal Growth Factor-activated Aryl Hydrocarbon Receptor Nuclear Translocator/HIF-1β Signal Pathway Up-regulates Cyclooxygenase-2 Gene Expression Associated with Squamous Cell Carcinoma

“…In fact, a previous study has reported that there is no correlation between EGFR and COX-2 in clinical lung cancer tumor samples or cell lines (46). The difference in results might lie in the alternation of EGFR-downstream signals such as p38 mitogenactivated protein kinase and Sp1/Sp3 to induce COX-2 expression (47). In clinical trials, dual blockage of EGFR and COX-2 in lung cancer leads to disappointing outcomes, since this is not able to overcome the innate resistance of EGFR tyrosine kinase inhibitor (48,49).…”

Cancer-related inflammation

“…These results show that an early genetic event that is necessary and sufficient for the development of a human tumour directly promotes the build-up of an inflammatory microenvironment 8 . Although these 8,9 and other results 10 connect the activation of protein-tyrosine-kinase-encoding oncogenes to inflammation, the precise roles of the various components of the inflammatory microenvironment in the progression of tumours remain to be defined.…”