Epidermal Growth Factor Receptor Variant III Expression Research Articles

Epidermal growth factor receptor variant III (EGFRvIII) vaccine (CDX-110) in GBM

2021 Background: Unlike conventional therapies for GBM, immunologic targeting of tumor-specific gene mutations allows precise eradication of neoplastic cells with reduced toxicity. EGFRvIII is a constitutively activated and immunogenic mutation not expressed in normal tissues, but widely expressed in GBM and other neoplasms. The cancer vaccine CDX-110 is comprised of an EGFRvIII-specific peptide sequence linked to keyhole limpet hemocyanin (KLH). Methods: A phase II multi-center trial assessed the immunogenicity and efficacy of CDX-110 in patients with newly-diagnosed, EGFRvIII+ GBM. After resection and radiation / TMZ, patients received CDX-110 vaccinations biweekly x 3, then monthly until tumor progression. Sequential cohorts received CDX-110 alone [ACTIVATE (n = 18)] or in combination with TMZ (200 mg/m2 x 5/28 days [ACT II A (n = 13)]) or (100 mg/m2 x 21/28 days [ACT II B (n=10)]). Results: Reversible systemic drug hypersensitivity reactions were seen in 1 ACTIVATE and 4 ACT II patients. Two patients had non-specific changes on MRI which were possibly due to the vaccine but which resolved. Despite grade 2 or 3 lymphopenia in all ACT II patients, EGFRvIII-specific immune responses were generated in all patients, and all immune responses were sustained or enhanced during subsequent TMZ cycles. Although ACT II B patients had more severe TMZ-induced lymphopenia, they developed greater EGFRvIII-specific immune responses (p = 0.028) when compared to ACT II A. EGFRvIII-specific IgG1 also increased in avidity with vaccination (Ka>>2x109M-1) in a randomly selected subset of 4 patients (p = 0.000068). Of the 23 recurrent tumors studied, 18 lost EGFRvIII expression (p = 0.001). There are no significant differences between ACT II A and B in estimated median TTP (18.5 vs. 14.9 months, p = 0.31) and OS (23.6 vs. 19.9 months, p = 0.75). ACTIVATE TTP (14.2 months) and OS (26.0 months) and ACT II TTP (15.2 months) and OS (23.6 months) compare favorably to a TMZ-treated, matched historical control group (TTP: 6.3 months; OS: 15.0 months). Conclusions: CDX-110 vaccination in patients with GBM appears very promising. TMZ enhances immune responses despite lymphodepletion. CDX-110 with simultaneous TMZ is under further investigation in a larger phase II trial. [Table: see text]

EGFRvIII and DNA double-strand break repair: a molecular mechanism for radioresistance in glioblastoma.

Glioblastoma multiforme (GBM) is the most lethal of brain tumors and is highly resistant to ionizing radiation (IR) and chemotherapy. Here, we report on a molecular mechanism by which a key glioma-specific mutation, epidermal growth factor receptor variant III (EGFRvIII), confers radiation resistance. Using Ink4a/Arf-deficient primary mouse astrocytes, primary astrocytes immortalized by p53/Rb suppression, as well as human U87 glioma cells, we show that EGFRvIII expression enhances clonogenic survival following IR. This enhanced radioresistance is due to accelerated repair of DNA double-strand breaks (DSB), the most lethal lesion inflicted by IR. The EGFR inhibitor gefitinib (Iressa) and the phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 attenuate the rate of DSB repair. Importantly, expression of constitutively active, myristylated Akt-1 accelerates repair, implicating the PI3K/Akt-1 pathway in radioresistance. Most notably, EGFRvIII-expressing U87 glioma cells show elevated activation of a key DSB repair enzyme, DNA-dependent protein kinase catalytic subunit (DNA-PKcs). Enhanced radioresistance is abrogated by the DNA-PKcs-specific inhibitor NU7026, and EGFRvIII fails to confer radioresistance in DNA-PKcs-deficient cells. In vivo, orthotopic U87-EGFRvIII-derived tumors display faster rates of DSB repair following whole-brain radiotherapy compared with U87-derived tumors. Consequently, EGFRvIII-expressing tumors are radioresistant and continue to grow following whole-brain radiotherapy with little effect on overall survival. These in vitro and in vivo data support our hypothesis that EGFRvIII expression promotes DNA-PKcs activation and DSB repair, perhaps as a consequence of hyperactivated PI3K/Akt-1 signaling. Taken together, our results raise the possibility that EGFR and/or DNA-PKcs inhibition concurrent with radiation may be an effective therapeutic strategy for radiosensitizing high-grade gliomas.

Expression of Epidermal Growth Factor Receptor variant III in laryngeal carcinoma tissues

Objectives Epidermal Growth Factor Receptor variant III (EGFRvIII) has been believed to be an attractive tumor-specific candidate for molecular targeting therapy. However, there is little literature dealing with this variant of EGFR expressed in laryngeal carcinomas. In the present study, we try to evaluate the expression of EGFRvIII, as well as EGFR, in laryngeal carcinoma tissues and its correlation with clinicopathological features. Methods Real-time polymerase chain reaction (real-time PCR) with TaqMan probes was applied to detect the expression for EGFR and EGFRvIII mRNA in the 39 pairs of samples of laryngeal carcinoma tissues and microscopically normal laryngeal mucosal tissues adjacent to the tumor. 2 −ΔΔCT method was used to obtain the relative quantity of target mRNA expression. The correlation between EGFRvIII expression and its clinicopathological features was analyzed by Pearson's chi-squared test. Results Among the 39 pairs of samples of laryngeal carcinoma tissues and microscopically normal laryngeal mucosal tissues adjacent to the tumor, the level of EGFR mRNA of the former (0.030 ± 0.076) was higher than that of the latter (0.011 ± 0.046) ( P < 0.01). EGFRvIII mRNA was detected only in six samples of laryngeal carcinoma tissues. While, as control, in 39 samples of microscopically normal laryngeal mucosal tissues, EGFRvIII mRNA was hardly detected. As analyzing the correlation between expression of EGFRvIII and EGFR, we found the positive rate of EGFRvIII expression was higher in samples with relative EGFR mRNA value ≥0.025 than those of EGFR mRNA <0.025. The difference between them was statistically significant ( P < 0.05). Conclusions Expression of EGFRvIII in laryngeal carcinoma was confirmed in this study. It is tumor-specific and tends to be more frequent in EGFR-over expressing tumor tissues and poorly differentiated ones, which may in part contribute to the malignant phenotype.

Pro-survival AKT and ERK signaling from EGFR and mutant EGFRvIII enhances DNA double-strand break repair in human glioma cells

The Epidermal growth factor receptor (EGFR) is frequently dysregulated in malignant glioma that leads to increased resistance to cancer therapy. Up-regulation of wild type or expression of mutants such as EGFR variant III (EGFRvIII), the most common EGFR mutation in malignant glioma, is associated with tumor radioresistance and poor clinical outcome. This radioresistance is thought to be the result of a strong cytoprotective response fueled by signaling via AKT and ERK that is heightened by radiation in the clinical dose range. Several groups including ours have shown that this response may modulate DNA repair. Herein, we show that expression of EGFRvIII promoted Pro-survival AKT and ERK signaling from EGFR and mutant EGFRvIII enhances DNA double-strand break repair in human glioma cells γ-H2AX foci resolution, a surrogate for double-strand break (DSB) repair, and thus enhanced DNA repair. Conversely, small molecule inhibitors targeting EGFR, MEK, and the expression of dominant-negative EGFR (EGFR-CD533) significantly reduced the resolution of γ-H2AX foci. When homologous recombination repair (HRR) and non-homologous end joining (NHEJ) were specifically examined, we found that EGFRvIII stimulated and CD533 compromised HRR and NHEJ, respectively. Furthermore, NHEJ was significantly blocked by inhibitors of AKT and ERK signaling pathways. Moreover, expression of EGFRvIII and CD533 increased and reduced, respectively, the formation of phospho-DNA-PKcs and -ATM repair foci, and RAD51 expression levels, indicating that DSB repair is regulated at multiple levels. Altogether, signaling from EGFR and EGFRvIII promotes both HRR and NHEJ that is likely one contributing factor towards the radioresistance of malignant gliomas.

Expression and significance of epidermal growth factor receptor variant III in primary gallbladder carcinoma

Objective To detect expression of epidermal growth factor receptor variant Ⅲ(EGFRvⅢ)and investigate its significance in development of primary gallbladder carcinoma. Methods Sixty-eight patients with primary gallbladder carcinoma entered this study. Immunohistoehemistry (SABC assay) and image analysis were used to semiquantitatively detect the expression of EGFRvⅢprotein. Results Positive expression rate of EGFRvⅢ was 53% in primary gallbladder carcinoma and 51% in adenocarcinoma. As cancer pathologic grading advanced, the expression of EGFRvⅢ increased with statistical deviation (q=2.49, P=0.02, q=3.63, P<0.01, q=5.69, P<0.01). EGFRv Ⅲexpression level showed no statistical deviation between male and female subjects(t=1.29, P=0.20).No significant relationship was observed between EGFRvⅢ expression and age(r=0.03, P=0.88).Conclusion EGFRvⅢ is overexpressed in primary gallbladder carcinoma and may play certain role in development of primary gallbladder carcinoma. Key words: Gallbladder neoplasms; Epidermal growth factor receptor variant Ⅲ; Immunohistochemistry

The ErbB signalling pathway: protein expression and prognostic value in epithelial ovarian cancer

Ovarian cancer is the most frequent cause of death from gynaecological cancer in the Western world. Current prognostic factors do not allow reliable prediction of response to chemotherapy and survival for individual ovarian cancer patients. Epidermal growth factor receptor (EGFR) and HER-2/neu are frequently expressed in ovarian cancer but their prognostic value remains unclear. In this study, we investigated the expression and prognostic value of EGFR, EGFR variant III (EGFRvIII), HER-2/neu and important downstream signalling components in a large series of epithelial ovarian cancer patients. Immunohistochemical staining of EGFR, pEGFR, EGFRvIII, Her-2/neu, PTEN (phosphatase and tensin homologue deleted on chromosome 10), total and phosphorylated AKT (pAKT) and phosphorylated ERK (pERK) was performed in 232 primary tumours using the tissue microarray platform and related to clinicopathological characteristics and survival. In addition, EGFRvIII expression was determined in 45 tumours by RT–PCR. Our results show that negative PTEN immunostaining was associated with stage I/II disease (P=0.006), non-serous tumour type (P=0.042) and in multivariate analysis with a longer progression-free survival (P=0.015). Negative PTEN staining also predicted improved progression-free survival in patients with grade III or undifferentiated serous carcinomas (P=0.011). Positive pAKT staining was associated with advanced-stage disease (P=0.006). Other proteins were expressed only at low levels, and were not associated with any clinicopathological parameter or survival. None of the tumours were positive for EGFRvIII. In conclusion, our results indicate that tumours showing negative PTEN staining could represent a subgroup of ovarian carcinomas with a relatively favourable prognosis.

RETRACTED ARTICLE: Targeting EGFR in bladder cancer

Expression and overexpression of the epidermal growth factor receptor (EGFR) have been described in several solid tumors including bladder, breast, colorectal, NSCLC, prostate, and ovarian cancers. In addition to gene amplification, point mutations within the kinase domain also occur. Previous reports indicate that the patient's response to gefitinib depends on either the presence of mutations within the kinase domain of EGFR or the expression of the most frequent alteration, the truncated EGFR variant III (EGFRvIII). Therefore, it is important to determine if these EGFR alterations are present in urothelial carcinoma. The kinase domain of EGFR (exons 18-21) from 11 bladder cancer cell lines as well as from 75 patient tumors was analyzed by automated sequencing. No mutations were detected in all samples tested. Furthermore, analysis of EGFRvIII by immunohistochemistry revealed that almost half of all the patient samples expressed this truncation in a urothelial carcinoma tissue microarray. However, there have been previous reports of inconsistencies in detecting EGFRvIII by immunohistochemistry owing to the specificity of the antibodies and the methodologies utilized. Therefore, these results were validated by reverse transcription PCR, real-time PCR and western blot analysis. In these assays, none of the samples tested positive for EGFRvIII. Taken together, these results indicate that mutations within the tyrosine kinase domain of EGFR and expression of EGFRvIII are rare events in bladder cancer and therefore do not contribute to the malignant phenotype of this tumor. These results have clinical implications in selecting tyrosine kinase inhibitors for the therapy of urothelial carcinoma.

Epidermal Growth Factor Receptor Variant III Status Defines Clinically Distinct Subtypes of Glioblastoma

The clinical significance of epidermal growth factor receptor variant III (EGFRvIII) expression in glioblastoma multiforme (GBM) and its relationship with other key molecular markers are not clear. We sought to evaluate the clinical significance of GBM subtypes as defined by EGFRvIII status. The expression of EGFRvIII was assessed by immunohistochemistry in 649 patients with newly diagnosed GBM. These data were then examined in conjunction with the expression of phospho-intermediates (in a subset of these patients) of downstream AKT and Ras pathways and YKL-40 as well as with known clinical risk factors, including the Radiation Therapy Oncology Group's recursive partitioning analysis (RTOG-RPA) class. The RTOG-RPA class was highly predictive of survival in EGFRvIII-negative patients but much less predictive in EGFRvIII-positive patients. These findings were seen in both an initial test set (n = 268) and a larger validation set (n = 381). Similarly, activation of the AKT/MAPK pathways and YKL-40 positivity were predictive of poor outcome in EGFRvIII-negative patients but not in EGFRvIII-positive patients. Pair-wise combinations of markers identified EGFRvIII and YKL-40 as prognostically important. In particular, outcome in patients with EGFRvIII-negative/YKL-40-negative tumors was significantly better than the outcome in patients with the other three combinations of these two markers. Established prognostic factors in GBM were not predictive of outcome in the EGFRvIII-positive subset, although this requires confirmation in independent data sets. GBMs negative for both EGFRvIII and YKL-40 show less aggressive behavior.

Mutations Within the Kinase Domain and Truncations of the Epidermal Growth Factor Receptor Are Rare Events in Bladder Cancer: Implications for Therapy

It has previously been reported that the patient response to gefitinib depends on the presence of mutations within the kinase domain of epidermal growth factor receptor (EGFR) or the expression of its truncated form, EGFR variant III (EGFRvIII). The focus of this study was to determine if these alterations are present within the tyrosine kinase and ligand-binding domain of EGFR in urothelial carcinoma. The kinase domain found within exons 18 to 21 of the EGFR from 11 bladder cancer cell lines and 75 patient tumors were subjected to automated sequencing. EGFRvIII expression was determined by immunohistochemistry using a urothelial carcinoma tissue microarray, and its expression was subsequently verified by reverse transcription PCR, real-time PCR, and Western blot analysis, using an EGFRvIII-transfected glioblastoma cell line and glioblastoma tumors as positive controls. Our analysis failed to detect mutations within the tyrosine kinase domain of EGFR in the 11 cell lines and 75 patients tested. The initial analysis of EGFRvIII expression by immunohistochemistry revealed that at least 50% of the patient tumors expressed EGFRvIII in a urothelial carcinoma tissue microarray. Conflicting reports exist, however, regarding the extent of EGFRvIII expression in tissues owing to the specificity of the antibodies and the methodologies used. Therefore, we sought to validate this observation by reverse transcription PCR, real-time PCR, and Western blot analysis. In these assays, none of the samples were positive for EGFRvIII except for control transfectants and glioblastomas. When our results are taken together, we conclude that alterations within the tyrosine kinase domain and expression of EGFRvIII are rare events in bladder cancer. The present study has clinical implications in selecting tyrosine kinase inhibitors for the therapy of urothelial carcinoma.