Inverse correlation ofepidermal growth factor receptormessenger RNA induction and suppression of anchorage-independent growth by OSI-774, an epidermal growth factor receptor tyrosine kinase inhibitor, in glioblastoma multiforme cell lines
Marc-Eric Halatsch, Esther E. Gehrke, Vassilios I. Vougioukas, Ingolf C. Bötefür, Farhad A.- Borhani, Thomas Efferth, Erich Gebhart, Sebastian Domhof, Ursula Schmidt, Michael Buchfelder
2004
Neurosurgical Focus
In cancers in humans, the expression and/or function of the EGFR is frequently altered due to gene amplification, overexpression, autocrine or paracrine ligand stimulation, or constitutive activation. 23, 46, 51, 57, 61, 62 Because these functional and structural alterations have been strongly implicated in increased cellular proliferation and tumorigenesis, decreased apoptosis, and poor prognosis, 34,46 several strategies involving mAbs, 32 immunotoxin conjugates, 1 antisense oligonucleotides,
more »
... 52 and ribozymes 11, 14 have been attempted ultimately to intercept cellular effects mediated by quantitatively and qualitatively aberrant EGFR. The EGFR is a transmembranous glycoprotein composed of an external domain, which binds activating ligands such as EGF and TGF␣, and an intracellular TyK domain, which on activation phosphorylates both the receptor itself and a variety of downstream effector molecules such as phospholipase C␥, phosphatidylinositol-3-kinase, and mitogen-activated protein kinase. The ultimate cellular response to the activation of the EGFR signaling cascade is DNA synthesis and cell division. 56 Neurosurg Focus 16 (2):Preview of J Neurosurg 100:523-533, 2004, Click here to return to Table of Contents Inverse correlation of epidermal growth factor receptor messenger RNA induction and suppression of anchorageindependent growth by OSI-774, an epidermal growth factor receptor tyrosine kinase inhibitor, in glioblastoma multiforme cell lines Object. Quantitative and qualitative alterations in the epidermal growth factor receptor (EGFR) commonly occur in many cancers in humans, including malignant gliomas. The aim of the current study was to evaluate molecular and cellular effects of OSI-774, a novel EGFR tyrosine kinase inhibitor, on nine glioblastoma multiforme (GBM) cell lines. Methods. The effects of OSI-774 on expression of EGFR messenger (m)RNA and protein, proliferation, anchorageindependent growth, and apoptosis were examined using semiquantitative reverse transcription-polymerase chain reaction, immunocytochemical analysis, Coulter counting, soft agar cloning, and terminal deoxynucleotidyl transferasemediated deoxyuridine triphosphate nick-end labeling/fluorescence-activated cell sorting, respectively. All p53 genes were completely and bidirectionally sequenced. Suppression of anchorage-independent growth by OSI-774 was inversely correlated to the induction of EGFR mRNA during relative serum starvation (r = Ϫ0.74) and was unrelated to p53 status. Overall, suppression of anchorage-independent growth was a considerably stronger effect of OSI-774 than inhibition of proliferation. The extent of OSI-774-induced apoptosis positively correlated with both proliferation and anchorage-independent growth of GBM cell lines (r = 0.75 and 0.79, respectively). In a single cell line derived from a secondary GBM, exposure to concentrations of greater than or equal to 1 mol/L resulted in a substantial net cell loss during proliferation studies. Conclusions. The induction of EGFR mRNA may constitute a cellular mechanism to counteract the inhibitory effect of OSI-774 on the anchorage-independent growth of GBM cells. In contrast, no considerable correlation could be established between baseline expression levels of EGFR (both mRNA and protein) in GBM cell lines and their biological response to OSI-774. The OSI-774 induced greater (p53-independent) apoptosis in more malignant GBM phenotypes and may be a promising therapeutic agent against secondary GBM. KEY WORDS • glioblastoma multiforme • epidermal growth factor receptor • tyrosine kinase inhibitor • proliferation • anchorage-independent growth • apoptosis Neurosurg. Focus / Volume 16 / February, 2004 1 Abbreviations used in this paper: cDNA = complementary DNA; DMSO = dimethyl sulfoxide; EGF = epidermal growth factor; EGFR = EGF receptor; FCS = fetal calf serum; GBM = glioblastoma multiforme; HBSS = Hanks balanced salt solution; mAb = monoclonal antibody; mRNA = messenger RNA; PBS = phosphate-buffered saline; PCR = polymerase chain reaction; PDGF = platelet-derived growth factor; PDGFR = PDGF receptor; RT = reverse transcription; TGF = transforming growth factor; TyK = tyrosine kinase; TNF = tumor necrosis factor; TUNEL = terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling.
doi:10.3171/foc.2004.16.2.12
fatcat:op7s3duaw5ecff3bcpav6flh6q
