Differential Sensitivity of Glioma-versus Lung Cancer-Specific EGFR Mutations to EGFR Kinase Inhibitors

Human Oncology and Pathogenesis Program, Department of Neurology, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA.
Cancer Discovery (Impact Factor: 19.45). 05/2012; 2(5):458-71. DOI: 10.1158/2159-8290.CD-11-0284
Source: PubMed


Activation of the epidermal growth factor receptor (EGFR) in glioblastoma (GBM) occurs through mutations or deletions in the extracellular (EC) domain. Unlike lung cancers with EGFR kinase domain (KD) mutations, GBMs respond poorly to the EGFR inhibitor erlotinib. Using RNAi, we show that GBM cells carrying EGFR EC mutations display EGFR addiction. In contrast to KD mutants found in lung cancer, glioma-specific EGFR EC mutants are poorly inhibited by EGFR inhibitors that target the active kinase conformation (e.g., erlotinib). Inhibitors that bind to the inactive EGFR conformation, however, potently inhibit EGFR EC mutants and induce cell death in EGFR-mutant GBM cells. Our results provide first evidence for single kinase addiction in GBM and suggest that the disappointing clinical activity of first-generation EGFR inhibitors in GBM versus lung cancer may be attributed to the different conformational requirements of mutant EGFR in these 2 cancer types. SIGNIFICANCE: Approximately 40% of human glioblastomas harbor oncogenic EGFR alterations, but attempts to therapeutically target EGFR with first-generation EGFR kinase inhibitors have failed. Here, we demonstrate selective sensitivity of glioma-specific EGFR mutants to ATP-site competitive EGFR kinase inhibitors that target the inactive conformation of the catalytic domain.

Download full-text


Available from: Dan Rohle, Oct 02, 2015
64 Reads
  • Source
    • "EGFRvV; a deletion of exons 25–28) and mutations affecting the tyrosine kinase domain (mainly exon 19 and codon L858) [3] [4] [5]. These mutations result in a constitutively activated isoform of the protein and contribute to oncogenic transformation [5] [6] [7] [8]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Epidermal growth factor receptor (EGFR) is frequently mutated in various types of cancer. Although all oncogenic mutations are considered activating, different tumour types have different mutation spectra. It is possible that functional differences underlie this tumour-type specific mutation spectrum. We have determined whether specific mutations in EGFR (EGFR, EGFRvIII and EGFR-L858R) have differences in binding partners, differences in downstream pathway activation (gene expression and phosphoproteins), and have functional consequences on cellular growth and migration. Using biotin pulldown and subsequent mass spectrometry we were able to detect mutation specific binding partners for EGFR. Differential binding was confirmed using a proximity ligation assay and/or Western Blot for the dedicator of cytokinesis 4 (DOCK4), UDP-glucose glycoprotein glucosyltransferase 1 (UGGT1), MYC binding protein 2 (MYCBP2) and Smoothelin (SMTN). We also demonstrate that each mutation induces the expression of a specific set of genes, and that each mutation is associated with specific phosphorylation patterns. Finally, we demonstrate using stably expressing cell lines that EGFRvIII and EGFL858R display reduced growth and migration compared to EGFR wildtype expressing cells. Our results indicate that there are distinct functional differences between different EGFR mutations. The functional differences between different mutations argue for the development of mutation specific targeted therapies. Copyright © 2015 Elsevier Ltd. All rights reserved.
    European journal of cancer (Oxford, England: 1990) 03/2015; 51(7). DOI:10.1016/j.ejca.2015.02.006 · 5.42 Impact Factor
  • Source
    • "Interestingly , the usefulness of certain irreversible EGFR inhibitors in EGFRvIII - positive GBM may obviate the need to combine c - MET and EGFR inhibitors ( Vivanco et al . , 2012 ) . Whether or not c - MET inhibitors are needed moving forward , SHP2 inhibitors might eventually offer an attractive alternative in the treatment of GBM where resistance to other inhibitors arises . Of course , our data support the potential usefulness of STAT3 inhibitors in treating GBM . STAT3 has previously been identified as a key"
    [Show abstract] [Hide abstract]
    ABSTRACT: Information from multiple signaling axes is integrated in the determination of cellular phenotypes. Here, we demonstrate this aspect of cellular decision making in glioblastoma multiforme (GBM) cells by investigating the multivariate signaling regulatory functions of the protein tyrosine phosphatase SHP2. Specifically, we demonstrate that SHP2's ability to simultaneously drive ERK and antagonize STAT3 pathway activities produces qualitatively different effects on the phenotypes of proliferation and resistance to EGFR and c-MET co-inhibition. While the ERK and STAT3 pathways independently promote proliferation and resistance to EGFR and c-MET co-inhibition, SHP2-driven ERK activity is dominant in driving cellular proliferation, and SHP2's antagonism of STAT3 phosphorylation prevails in promoting GBM cell death in response to EGFR and c-MET co-inhibition. Interestingly, the extent of these SHP2 signaling regulatory functions is diminished in glioblastoma cells expressing sufficiently high levels of the EGFR variant III (EGFRvIII) mutant, which is commonly expressed in GBM. In cells and tumors expressing EGFRvIII, SHP2 also antagonizes EGFRvIII and c-MET phosphorylation and drives expression of HIF-1/2α, adding complexity to the evolving understanding of SHP2's regulatory functions in GBM.
    Journal of Cell Science 06/2014; 127(16). DOI:10.1242/jcs.150862 · 5.43 Impact Factor
  • Source
    • "Cellular and molecular heterogeneity involving wild-type and mutant RTK composition, as we observed in this study, likely complicates strategies to effectively inhibit oncogenic signaling. Indeed, investigations carried out in vitro and in human patients indicate that the inhibitor sensitivity profiles of wild-type EGFR and EGFRvIII are distinct [46]. In this respect, our findings and those of others support the notion that a successful therapeutic strategy will require the effective inhibition of both mutant and wild-type receptor at concentrations achievable in the target tissue. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Intragenic deletion is the most common form of activating mutation among receptor tyrosine kinases (RTK) in glioblastoma. However, these events are not detected by conventional DNA sequencing methods commonly utilized for tumor genotyping. To comprehensively assess the frequency, distribution, and expression levels of common RTK deletion mutants in glioblastoma, we analyzed RNA from a set of 192 glioblastoma samples from The Cancer Genome Atlas for the expression of EGFRvIII, EGFRvII, EGFRvV (carboxyl-terminal deletion), and PDGFRAΔ8,9. These mutations were detected in 24, 1.6, 4.7, and 1.6 % of cases, respectively. Overall, 29 % (55/189) of glioblastomas expressed at least one RTK intragenic deletion transcript in this panel. For EGFRvIII, samples were analyzed by both quantitative real-time PCR (QRT-PCR) and single mRNA molecule counting on the Nanostring nCounter platform. Nanostring proved to be highly sensitive, specific, and linear, with sensitivity comparable or exceeding that of RNA seq. We evaluated the prognostic significance and molecular correlates of RTK rearrangements. EGFRvIII was only detectable in tumors with focal amplification of the gene. Moreover, we found that EGFRvIII expression was not prognostic of poor outcome and that neither recurrent copy number alterations nor global changes in gene expression differentiate EGFRvIII-positive tumors from tumors with amplification of wild-type EGFR. The wide range of expression of mutant alleles and co-expression of multiple EGFR variants suggests that quantitative RNA-based clinical assays will be important for assessing the relative expression of intragenic deletions as therapeutic targets and/or candidate biomarkers. To this end, we demonstrate the performance of the Nanostring assay in RNA derived from routinely collected formalin-fixed paraffin-embedded tissue.
    Acta Neuropathologica 11/2013; 127(5). DOI:10.1007/s00401-013-1217-3 · 10.76 Impact Factor
Show more