Mutant EGFR is required for maintenance of glioma growth in vivo, and its ablation leads to escape from receptor dependence

Abstract

Epidermal growth factor receptor (EGFR) gene amplification is the most common genetic alteration in high-grade glioma, and ≈50% of EGFR-amplified tumors also harbor a constitutively active mutant form of the receptor, ΔEGFR. Although ΔEGFR greatly enhances tumor growth and is thus an attractive target for anti-glioma therapies, recent clinical experiences with EGFR kinase inhibitors have been disappointing, because resistance is common and tumors eventually recur. Interestingly, it has not been established whether ΔEGFR is required for maintenance of glioma growth in vivo, and, by extension, if it truly represents a rational therapeutic target. Here, we demonstrate that in vivo silencing of regulatable ΔEGFR with doxycycline attenuates glioma growth and, therefore, that it is crucial for maintenance of enhanced tumorigenicity. Similar to the clinical experience, tumors eventually regained aggressive growth after a period of stasis, but interestingly, without re-expression of ΔEGFR. To determine how tumors acquired this ability, we found that a unique gene, KLHDC8, herein referred to as SΔE (Substitute for ΔEGFR Expression)-1, is highly expressed in these tumors, which have escaped dependence on ΔEGFR. SΔE-1 is also expressed in human gliomas and knockdown of its expression in ΔEGFR-independent "escaper" tumors suppressed tumor growth. Taken together, we conclude that ΔEGFR is required for both glioma establishment and maintenance, and that gliomas undergo selective pressure in vivo to employ alternative compensatory pathways to maintain aggressiveness in the event of EGFR silencing. Such alternative pathways function as substitutes for ΔEGFR signaling and should therefore be considered as potential targets for additional therapy.tumorigenicity | gliomblastoma | epidermal growth factor receptor

Crosstalk between the urokinase-type plasminogen activator receptor and EGF receptor variant III supports survival and growth of glioblastoma cells

“…Thus, we examined the effects of Dox on the activity of cell-signaling factors known to be regulated by both EGFRvIII and uPAR. In Dox-treated U373MG cells, phospho-ERK (P-ERK) was slightly decreased and P-STAT3 was more substantially decreased, as anticipated (15).…”

“…Failure of EGFR-targeting therapeutics to induce sustained remission may reflect mutations in gene products downstream of the EGFR, such as the tumor suppressor PTEN, or activation of alternative receptors with overlapping cell-signaling activity (10,(12)(13)(14). Mukasa et al (15) developed a derivative of the U373MG GBM cell line in which EGFRvIII is expressed under the control of a doxycycline (Dox)-repressible promoter system. These cells establish xenografts in mice only if EGFRvIII is expressed.…”

“…When the mice are treated with Dox to suppress EGFRvIII expression, the tumors enter a state of dormancy; however, many tumors eventually reestablish aggressive growth. Tumors that escape dependency on EGFRvIII in mice may represent a model for phenotypes that emerge in patients with GBM who are treated with EGFR-targeting therapeutics (15).…”

Distribution of Gefitinib to the Brain Is Limited by P-glycoprotein (ABCB1) and Breast Cancer Resistance Protein (ABCG2)-Mediated Active Efflux

“…Moreover, recent studies have demonstrated that EGFRvIII is required for tumor maintenance in glioma (Mukasa et al, 2010). The EGFR tyrosine kinase inhibitor gefitinib has been evaluated in a number of clinical trials for GBM; however, results have been disappointing (Rich et al, 2004;Lieberman et al, 2004).…”

Soluble Urokinase Receptor Is Released Selectively by Glioblastoma Cells That Express Epidermal Growth Factor Receptor Variant III and Promotes Tumor Cell Migration and Invasion*

“…Parental U87MG cells and U87MG cells that overexpress wild-type EGFR (WT-EGFR) or express EGFRvIII were described previously (43). Parental U373MG cells, U373MG cells that overexpress WT-EGFR, and U373MG cells that express EGFRvIII under the control of a doxycycline-repressible promoter also were previously described (44). These cells were cultured in the presence of puromycin (1 g/ml) and geneticin (200 g/ml), with or without doxycycline (1 g/ml).…”