High Epidermal Growth Factor Receptor Expression Research Articles

Glioblastoma TCGA Mesenchymal and IGS 23 Tumors are Identifiable by IHC and have an Immune-phenotype Indicating a Potential Benefit from Immunotherapy.

Molecular subtype classifications in glioblastoma may detect therapy sensitivities. IHC would potentially allow the identification of molecular subtypes in routine clinical practice. Formalin-fixed, paraffin-embedded tumor samples of 124 uniformly treated, newly diagnosed patients with glioblastoma were submitted to RNA sequencing, IHC, and immune-phenotyping to identify differences in molecular subtypes associated with treatment sensitivities. We detected high molecular and IHC overlapping of the The Cancer Genome Atlas (TCGA) mesenchymal subtype with instrinsic glioma subtypes (IGS) cluster 23 and of the TCGA classical subtype with IGS cluster 18. IHC patterns, gene fusion profiles, and immune-phenotypes varied across subtypes. IHC revealed that the TCGA classical subtype was identified by high expression of EGFR and low expression of PTEN, while the mesenchymal subtype was identified by low expression of SOX2 and high expression of two antibodies, SHC1 and TCIRG1, selected on the basis of RNA differential transcriptomic expression. The proneural subtype was identified by frequent positive IDH1 expression and high Olig2 and Ki67 expression. Immune-phenotyping showed that mesenchymal and IGS 23 tumors exhibited a higher positive effector cell score, a higher negative suppressor cell score, and lower levels of immune checkpoint molecules. The cell-type deconvolution analysis revealed that these tumors are highly enriched in M2 macrophages, resting memory CD4+ T cells, and activated dendritic cells, indicating that they may be ideal candidates for immunotherapy, especially with anti-M2 and/or dendritic cell vaccination. There is a subset of tumors, frequently classified as mesenchymal or IGS cluster 23, that may be identified with IHC and could well be optimal candidates for immunotherapy.

EGFR and HER2 Expression in Primary Ovarian High-Grade Serous Carcinoma and Their Prognostic Value

Background: Human epidermal growth factors receptors such as EGFR and HER2 play an important role in tumorigenesis and are used as therapeutic targets. Their role in aggressive primary ovarian high-grade serous carcinoma (HGSC) is controversial. Aim: To study the expression of EGFR and HER2 in ovarian HGSC, to correlate their expression with other clinicopathological parameters and to study their prognostic value. Methods: Imunohistochemical staining of EGFR, HER2 and Ki-67 was done for 54 ovarian HGSC specimens. According to the used scoring methods, the expression of EGFR and HER2 was classified as high or low. Results: High expression of EGFR and HER2 was found in a minority of specimens; 39% and 15%, respectively. None of the studied clinicopathological parameters correlated significantly with the expression of EGFR and HER2, except for the carcinoembryonic antigen level which correlated positively with HER2 expression. Disease-free survival of patients did not differ significantly according to the level of expression of EGFR and HER2 (p =0.684 and 0.186, respectively). Similarly, overall survival did not differ significantly (p =0.911 and 0.346, respectively). Conclusion: The results of this study suggest that the prognostic value of EGFR and HER2 in ovarian HGSC is questionable.

Combined High Resistin and EGFR Expression Predicts a Poor Prognosis in Breast Cancer

Elevated levels of resistin and epidermal growth factor receptor (EGFR) facilitate the development of breast cancer, although there are no reports of any correlation between these proteins. This study analyzed 392 human breast cancer tissue specimens and 42 samples of adjacent normal tissue. Rates of positive and strongly positive resistin expression were significantly higher in breast cancer tissue than in the adjacent nontumor tissue (83.2% vs. 23.8% and 20.9% vs. 0.0%, respectively; P < 0.001 for both comparisons). Positive resistin expression was significantly associated with tumor size, grade, stage, estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2) status, and molecular classification; strongly positive resistin expression was associated with tumor grade, ER, PR, HER2 status, and molecular classification. Significantly positive correlations were observed between positive and strongly positive resistin expression and corresponding levels of EGFR expression. Relapse-free and overall survival was worse for patients with high levels of both proteins than for those with high levels of only one protein or normal levels of both proteins. Our evidence suggests that combined high levels of resistin and EGFR expression correlate with survival in patients with breast cancer.

Identification of prognostic aging-related genes associated with immunosuppression and inflammation in head and neck squamous cell carcinoma.

Aging is regarded as a dominant risk factor for cancer. Additionally, inflammation and asthenic immune surveillance with aging may facilitate tumor formation and development. However, few studies have comprehensively analyzed the relationship between aging-related genes (AGs) and the prognosis, inflammation and tumor immunity of head and neck squamous cell carcinoma (HNSCC). Here, we initially screened 41 differentially expressed AGs from The Cancer Genome Atlas (TCGA) database. In the training set, a prognosis risk model with seven AGs (APP, CDKN2A, EGFR, HSPD1, IL2RG, PLAU and VEGFA) was constructed and validated in the TCGA test set and the GEO set (P < 0.05). Using univariate and multivariate Cox regression analyses, we confirmed that risk score was an independent prognostic factor of HNSCC patients. In addition, a high risk score was significantly correlated with immunosuppression, and high expression of PLAU, APP and EGFR was the main factor. Furthermore, we confirmed that a high risk score was significantly associated with levels of proinflammatory factors (IL-1α, IL-1β, IL-6 and IL-8) in HNSCC samples. Thus, this risk model may serve as a prognostic signature and provide clues for individualized immunotherapy for HNSCC patients.

Growth Factor Receptor Expression in Oropharyngeal Squamous Cell Cancer: Her1-4 and c-Met in Conjunction with the Clinical Features and Human Papillomavirus (p16) Status.

Simple SummaryGrowth factor expression is a negative prognostic factor in head and neck squamous cell carcinoma (HNSCC). Targeted therapy has a limited effect on the treatment of advanced stages due to evolving resistance mechanisms. The aim of this study was to assess the distribution of growth factor receptors in oropharyngeal squamous cell cancer (OPSCC) and evaluate their role in the context of the human papillomavirus status, prognosis and possible relevance for targeted therapy. Tissue microarrays of 78 primary OPSCC, 35 related lymph node metastasis, 6 distant metastasis and 9 recurrent tumors were manufactured to evaluate the expression of human epidermal growth factor receptor (EGFR/erbB/Her)1–4 and c-Met by immunohistochemistry. EGFR and c-Met are relevant negative prognostic factors especially in noxae-induced OPSCC. Thus, dual targeting of EGFR and c-Met could be a promising prospective target in OPSCC treatment. Frequent coexpression of assessed receptors represents a possible intrinsic resistance mechanism in targeted therapy.This study aimed to assess the distribution of growth factor receptors in oropharyngeal squamous cell cancer (OPSCC) and evaluate their role in the context of human papillomavirus (HPV) status, prognosis and potential relevance for targeted therapy. The protein expression of human epidermal growth factor receptor (Her)1–4 and c-Met were retrospectively assessed using semiquantitative immunohistochemistry on tissue microarrays and analyzed for correlations as well as differences in the clinicopathological criteria. Her1–4 and c-met were overexpressed compared to normal mucosa in 46%, 4%, 17%, 27% and 23%, respectively. Interestingly, most receptors were coexpressed. Her1 and c-Met were inversely correlated with p16 (p = 0.04; p = 0.02). Her2 and c-Met were associated with high tobacco consumption (p = 0.016; p = 0.04). High EGFR, Her3, Her4 and c-Met expression were associated with worse overall and disease-free survival (p ≤ 0.05). Furthermore, EGFR and c-Met expression showed raised hazard ratios of 2.53 (p = 0.02; 95% CI 1.24–5.18) and 2.45 (p = 0.02; 95% CI 1.13–5.35), respectively. Her4 was expressed less in distant metastases than in corresponding primary tumors and was correlated to a higher T category. EGFR and c-Met are relevant negative prognostic factors in OPSCC, independent of known clinicopathological parameters. We suggest dual targeting of EGFR and c-Met as a promising strategy for OPSCC treatment.

Prognostic Worth of Epidermal Growth Factor Receptor (EGFR) in Patients with Head and Neck Tumors.

Introduction Head and neck tumors (HNT) are tumors that normally occur at the head and neck region of the body. Epidermal growth factor receptor (EGFR) has been found to be highly expressed in breast and other tumors; therefore, there is the need to investigate the level of EGFR expression among patients with head and neck tumors in Ghana. Method The level of EGFR expression was determined in head and neck tumor and control head and neck tissues with quantitative real-time PCR and immunohistochemistry analysis. Results The level of EGFR expressions was high in tumor tissues than in the control tissues. There was a significant difference of p value 0.025 among the ages >40 and ≤ 40 when the high and low level of EGFR was compared in the head and neck malignant tumor. The area under the curve for the high expression of EGFR among the malignant head and neck tumors was 0.901 with a specificity of 86.4%. Conclusion EGFR can serve as a prognostic marker in monitoring patients with HNT as well as a molecular therapeutic target.

Interactions between anti-EGFR therapies and cytotoxic chemotherapy in oesophageal squamous cell carcinoma: why clinical trials might have failed and how they could succeed

PurposeOesophageal squamous cell carcinoma (ESCC) has a poor prognosis. Advanced tumours are treated with fluoropyrimidine/platinum chemotherapy followed by irinotecan or taxane monotherapy, but resistance is common and new treatments are needed. Approximately 20% of ESCCs carry copy number gain (CNG) of the epidermal growth factor receptor (EGFR) gene. Previous trials show that while anti-EGFR monotherapy benefits biomarker-selected patients with EGFR CNG and/or high EGFR expression, combining anti-EGFR therapies with platinum fluoropyrimidine chemotherapies is not effective, and uncertainty remains regarding the optimal cytotoxic chemotherapy partner for anti-EGFR therapies in ESCC.MethodsThe effects of EGFR CNG on fluoropyrimidine/platinum chemotherapy sensitivity in a cohort of gastroesophageal cancer patients (n = 302) was evaluated. Drug combination studies using the EGFR inhibitor gefitinib with cytotoxic chemotherapies, docetaxel, cisplatin, oxaliplatin and irinotecan, on cell proliferation and cell death of EGFR CNG ESCC cell lines were assessed.ResultsEGFR CNG in gastroesophageal cancer patients was associated with improved overall survival following fluoropyrimidine/platinum chemotherapy. However, co-administration of gefitinib and oxaliplatin or cisplatin was frequently antagonistic in cell-based assays in EGFR CNG ESCC, whereas the combination of gefitinib with docetaxel or irinotecan was more efficacious. Co-administration of gefitinib/docetaxel and sequential administration of docetaxel before gefitinib showed synergy, but docetaxel given after gefitinib was antagonistic.ConclusionGefitinib/platinum co-administration demonstrated antagonism suggesting a possible explanation for the lack of benefit from addition of anti-EGFR therapies to fluoropyrimidine/platinum chemotherapy in trials. Gefitinib/docetaxel co-administration demonstrated synergy suggesting taxanes could be the most effective cytotoxic partner for anti-EGFR therapies in EGFR CNG-positive ESCC, but careful consideration of drug scheduling is required.

BIOM-22. EPIDERMAL GROWTH FACTOR RECEPTOR (EGFR) AS A MOLECULAR DETERMINANT OF GLIOBLASTOMA RESPONSE TO DOPAMINE RECEPTOR 2 (DRD2) INHIBITORS

Abstract BACKGROUND There are ongoing clinical trials exploring the efficacy of dopamine receptor 2 (DRD2) inhibition against glioblastomas, the most common primary brain tumor. Here we examine potential molecular determinants of this efficacy. METHODS The Cancer Genome Atlas (TCGA) glioblastoma database and other published mRNA profiles were used to analyze the DRD2 and EGFR expression pattern. In vitro and in vivo responses to DRD2 inhibitors were determined using patient derived xenograft (PDX) glioblastoma models. Immunohistochemical studies were performed on clinically annotated glioblastoma samples derived from patients treated with ONC201, a DRD2 inhibitor. RESULTS Analysis of clinical glioblastoma specimens derived from independent patient cohorts revealed an inverse correlation between EGFR and DRD mRNA expression, with implication that signaling mediated by these proteins shares overlapping functions. In independent panels of PDX glioblastoma lines, high EGFR expression was associated with poor in vitro and in vivo response to DRD2 inhibitors, including haloperidol and ONC201. Moreover, ectopic expression of a constitutively active EGFR, EGFRvIII, suppressed glioblastoma sensitivity to ONC201. DRD2 expression positively correlated with expression of rate-limiting enzymes for dopamine synthesis as well as dopamine secretion, suggesting contribution of autocrine DRD2 signaling. Analysis of specimens from patients treated with ONC201 (n = 15) showed an inverse correlation between the intensity of EGFR staining and clinical response. The median overall survival for patients with high and low EGFR staining was 162 and 373 days, respectively (p = 0.037). CONCLUSIONS High EGFR expression is a determinant of poor glioblastoma response to DRD2. This finding should inform future clinical trial designs.

Reversal effect of peptide-modified chitosan tetramethylpyrazine nanoparticles on multidrug resistance in tumor cells

To prepare peptide-modified chitosan tetramethylprazine nanoparticles(FGF-CS-TMP-NPS) and investigate its reversal effect on multidrug resistance in tumor cells. The pEGF-CS-TMP-NPs were prepared by ion crosslinking method, and their physicochemical properties were investigated. Western blot was used to detect the expression levels of epidermal growth factor receptor(EGFR)(MCF-7, MCF-7/ADR, K562 and K562/ADR) and drug-resistant related protein P-gp. MCF-7/ADR and K562/ADR were selected as cell models. The cytotoxicity of pEGF-CS-TMP-NPs, the multiple of cell resistance to adriamycin, the reversal resistance index of pEGF-CS-TMP-NPs to doxorubicin and the sensitization of pEGF-CS-TMP-NPs to doxorubicin were detected by MTT assay. After MCF-7/ADR and K562/ADR were treated with pEGF-CS-TMP-NPs, the expression changes of P-gp were detected by Western blot. The encapsulation efficiency and drug loading of pEGF-CS-TMP-NPs were 37.66%± 0.53% and 3.25%± 0.34% respectively in HPLC. The nanoparticles showed an average particle size of(150.50±9.3) nm, polymer dispersity index of(0.059±0.007) and Zeta potential of(19.30±2.02) mV as detected by laser particle size analyzer. The nanoparticles were spherical and well dispersed under transmission electron microscope. Western blot results showed that EGFR was positively expressed in MCF-7 and MCF-7/ADR cells, while negatively expressed in K562 and K562/ADR cells. P-gp was highly expressed in MCF-7/ADR and K562/ADR, while negatively expressed in MCF-7 and K562. pEGF-CS-TMP-NPs had a weak effect on MCF-7/ADR and K562/ADR. The adriamycin resistance of MCF-7/ADR cells was 108.36 times, and that of K562/ADR cells was more than 100 times. When IC_(85) of pEGF-CS-TMP-NPs was used as the administration concentration, the reversion index of MCF-7/ADR and K562/ADR cells was 3.68 and 1.87, respectively. pEGF-CS-TMP-NPs could enhance the sensitivity of adriamycin to MCF-7/ADR cells in a positive correlation with the concentration, and the sensitivity was significantly higher than that of K562/ADR cells. Western blot results showed that the expression level of P-gp in MCF-7/ADR cells decreased significantly after treatment with pEGF-CS-TMP-NPs, while the expression level of P-gp in K562/ADR cells did not change significantly. Experimental results show that pEGF-CS-TMP-NPs have an active targeting effect on MCF-7/ADR cells with high EGFR expression, and can effectively reverse the multidrug resistance of MCF-7/ADR cells. Active targeting effect is related to the peptides modification of nanoparticles, and the mechanism of reversing tumor MDR may be achieved by down-regulating the expression level of P-gp.

Ethnic disparities in demographic, clinicopathologic and biological behaviours and prognosis of gastric cancer in northwest China.

This retrospective study aimed to investigate ethnic disparities in demographic, clinicopathologic, and biological behaviours of gastric cancer (GC) in a high GC incidence area of China. There were 5022 GC patients, including 3987 Han (79.4%) and 987 Hui (14.4%) patients from Northwest China. All patient data were retrieved from 2009 to 2017. Median survival was estimated using the Kaplan‐Meier method and compared using the log‐rank test. A Cox proportional hazards model was used to assess the impact of covariates. Similarly, low 5‐year OS rates were observed in both the Hui and Han groups (23.8% and 24.2% respectively). Hui patients with stage T1 or N0 or with tumours <5 cm had 2.144‐fold, 1.426‐fold and 1.305‐fold increased risks of poor prognosis compared with Han patients with these characteristics respectively (all p < 0.05). Further, Hui patients had 1.265‐fold, 1.364‐fold and 1.401‐fold increased risks of poor prognosis compared with Han patients among those with high expression of Ki67, EGFR and VEGF respectively (all p < 0.05). There are ethnic disparities in the prognosis of GC patients in Northwest China. Understanding the effects of ethnicity on GC will guide reasonable evaluations of prognosis and future interventions to equalise access to high‐quality care for GC patients of different ethnicities in China.

Targeting and Efficacy of Novel mAb806-Antibody-Drug Conjugates in Malignant Mesothelioma.

Epidermal growth factor receptor (EGFR) is highly overexpressed in malignant mesothelioma (MM). MAb806 is a novel anti-EGFR antibody that selectively targets a tumor-selective epitope. MAb806-derived antibody drug conjugates (ADCs), ABT-414, ABBV-221 and ABBV-322, may represent a novel therapeutic strategy in MM. EGFR and mAb806 epitope expressions in mesothelioma cell lines were evaluated using an array of binding assays, and the in vitro cell effects of ABT-414 and ABBV-322 were determined. In vivo therapy studies were conducted in mesothelioma xenograft and patient-derived xenograft (PDX) tumor models. We also performed biodistribution and imaging studies to allow the quantitative targeting of MM by mAb806 using a 89Zr-labeled immunoconjugate—ch806. A high EGFR expression was present in all mesothelioma cell lines evaluated and mAb806 binding present in all cell lines, except NCIH-2452. ABT-414 and ABBV-322 resulted in significant tumor growth inhibition in MM models with high EGFR and mAb806 epitope expressions. In contrast, in an EGFR-expressing PDX model that was negative for the mAb806 epitope, no growth inhibition was observed. We demonstrated the specific targeting of the mAb806 epitope expressing MM tumors using 89Zr-based PET imaging. Our data suggest that targeting EGFR in MM using specific ADCs is a valid therapeutic strategy and supports further investigation of the mAb806 epitope expression as a predictive biomarker.

A simplified integrated molecular and immunohistochemistry-based algorithm allows high accuracy prediction of glioblastoma transcriptional subtypes

AbstractGlioblastomas (GBM) can be classified into three major transcriptional subgroups (proneural, mesenchymal, classical), underlying different molecular alterations, prognosis, and response to therapy. However, transcriptional analysis is not routinely feasible and assessment of a simplified method for glioblastoma subclassification is required. We propose an integrated molecular and immunohistochemical approach aimed at identifying GBM subtypes in routine paraffin-embedded material. RNA-sequencing analysis was performed on representative samples (n = 51) by means of a “glioblastoma transcriptional subtypes (GliTS) redux” custom gene signature including a restricted number (n = 90) of upregulated genes validated on the TCGA dataset. With this dataset, immunohistochemical profiles, based on expression of a restricted panel of gene classifiers, were integrated by a machine-learning approach to generate a GliTS based on protein quantification that allowed an efficient GliTS assignment when applied to an extended cohort (n = 197). GliTS redux maintained high levels of correspondence with the original GliTS classification using the TCGA dataset. The machine-learning approach designed an immunohistochemical (IHC)-based classification, whose concordance was 79.5% with the transcriptional- based classification, and reached 90% for the mesenchymal subgroup. Distribution and survival of GliTS were in line with reported data, with the mesenchymal subgroup given the worst prognosis. Notably, the algorithm allowed the identification of cases with comparable probability to be assigned to different GliTS, thus falling within overlapping regions and reflecting an extreme heterogeneous phenotype that mirrors the underlying genetic and biological tumor heterogeneity. Indeed, while mesenchymal and classical subgroups were well segregated, the proneural types frequently showed a mixed proneural/classical phenotype, predicted as proneural by the algorithm, but with comparable probability of being assigned to the classical subtype. These cases, characterized by concomitant high expression of EGFR and proneural biomarkers, showed lower survival. Collectively, these data indicate that a restricted panel of highly sensitive immunohistochemical markers can efficiently predict GliTS with high accuracy and significant association with different clinical outcomes.The authors developed a novel simplified assay for glioblastoma transcriptional classification on formalin-fixed-paraffin-embedded tissue samples. On such dataset, immunohistochemical profiles, based on expression of a restricted panel of gene classifiers, were integrated by machine learning approach to generate a glioblastoma transcriptional signature based on protein quantification that allowed to efficiently assign transcriptional subgroups to an extended cohort. Correlations with both histopathological features and clinical outcome have been also performed.

Epidermal growth factor receptor as a molecular determinant of glioblastoma response to dopamine receptor D2 inhibitors.

There are ongoing clinical trials exploring the efficacy of dopamine receptor D2 (DRD2) inhibition against glioblastomas, the most common primary brain tumor. Here we examine potential molecular determinants of this efficacy. The Cancer Genome Atlas glioblastoma database and other published mRNA profiles were used to analyze the DRD2 and epidermal growth factor receptor (EGFR) expression pattern. In vitro and in vivo responses to DRD2 inhibitors were determined using patient-derived xenograft (PDX) glioblastoma models. Immunohistochemical studies were performed on clinically annotated glioblastoma samples derived from patients treated with ONC201. Analysis of clinical glioblastoma specimens derived from independent patient cohorts revealed an inverse correlation between EGFR and DRD2 mRNA expression, with implication that signaling mediated by these proteins shares overlapping functions. In independent panels of PDX glioblastoma lines, high EGFR expression was associated with poor in vitro and in vivo response to DRD2 inhibitors, including haloperidol and ONC201. Moreover, ectopic expression of a constitutively active EGFR, variant (v)III, suppressed glioblastoma sensitivity to ONC201. DRD2 expression positively correlated with expression of rate-limiting enzymes for dopamine synthesis as well as dopamine secretion, suggesting contribution of autocrine DRD2 signaling. Analysis of specimens from patients treated with ONC201 (n = 15) showed an inverse correlation between the intensity of EGFR staining and clinical response. The median overall survival for patients with high and low EGFR staining was 162 and 373 days, respectively (0.037). High EGFR expression is a determinant of poor glioblastoma response to DRD2. This finding should inform future clinical trial designs.

MiR-450a-5p strengthens the drug sensitivity of gefitinib in glioma chemotherapy via regulating autophagy by targeting EGFR

Glioma reported to be refractory to EGFR tyrosine kinase inhibitor is the most common malignant tumor in central nervous system. Our research showed the low expression of miR-450a-5p and high expression of EGFR in glioma tissues. MiR-450a-5p was also observed to synergize with gefitinib to inhibit the proliferation, migration and invasion and induce the apoptosis and autophagy of glioma cells. Furthermore, miR-450a-5p was demonstrated to target 3′UTR of EGFR, and regulated EGFR-induced PI3K/AKT/mTOR signaling pathway. Moreover, the above effects induced by miR-450a-5p in glioma cells were reversed by WIPI1 silencing. The inhibition role of miR-450a-5p on glioma growth was also confirmed in vivo by subcutaneous and intracranial tumor xenografts. Therefore, we conclude that miR-450a-5p synergizes with gefitinib to inhibit the glioma tumorigenesis through inducing autophagy by regulating the EGFR-induced PI3K/AKT/mTOR signaling pathway, thereby enhancing the drug sensitivity of gefitinib.

Vandetanib inhibits cell growth in EGFR-expressing cutaneous squamous cell carcinoma

Advanced cutaneous squamous cell carcinoma (SCC) responds poorly to chemotherapy, leading to significant morbidity or death. Overexpression of epidermal growth factor receptor (EGFR) is frequently observed in advanced cutaneous SCC. Vandetanib is a multiple tyrosine kinase targeting vascular endothelial growth factor receptor-2 (VEGFR2), EGFR, and the rearranged during transfection (RET) proto-oncogene. Vandetanib has been reported to inhibit tumor growth in head and neck SCC. However, the efficacy of vandetanib against cutaneous SCC has not been thoroughly investigated. The aim of this study is to evaluate the efficacy of vandetanib against cutaneous SCC in vitro and in vivo. Vandetanib is found to inhibit the proliferation of cutaneous SCC cells as assessed by cell viability and clonogenic assay. Cell death analysis indicates that vandetanib induces cell death in SCC cells but not in normal human keratinocytes or fibroblasts. The in vivo anti-tumor effect of vandetanib is shown in xenograft tumor models using A431 SCC cells. Mechanistically, vandetanib suppresses the phosphorylation of EGFR in SCC cells. Clinically, EGFR expression levels are elevated in cutaneous SCC specimens, relative to normal epidermis. In conclusion, we identified vandetanib as a novel therapeutic option for cutaneous SCC, especially in tumors with high EGFR expression.

Epidermal growth factor receptor expression as a molecular determinant of glioblastoma response to the dopamine receptor D2 antagonist, ONC201.

e14552 Background: Durable response in glioblastoma patients have been reported in phase I/II clinical trials for the blood-brain penetrant dopamine receptor D2 (DRD2) antagonist, ONC201. Here we examine potential molecular determinants of response to DRD2 inhibition. Methods: The Cancer Genome Atlas (TCGA) glioblastoma database and other published mRNA profiles were used to analyze the DRD2 expression pattern. In vitro and in vivo responses to ONC201 were determined using patient derived xenograft glioblastoma models. Immunohistochemical studies were performed on clinically annotated glioblastoma samples derived from phase I/II clinical trials involving ONC201. Results: For the majority of clinical glioblastoma specimens in both the TCGA and non-TCGA dataset, epidermal growth factor receptor (EGFR) expression was inversely correlated with DRD2. This observation was recapitulated in a panel of patient-derived glioblastoma lines. In this panel of DRD2 expressing lines, high EGFR expression was associated with poor response to ONC201 in vitro and in vivo. Moreover, ectopic expression of EGFR reduced DRD2 expression and ONC201 sensitivity, suggesting functional redundancy between DRD2 and EGFR. In cell lines and clinical glioblastoma samples, DRD2 expression closely associated with the expression of rate-limiting enzymes for dopamine synthesis, suggesting dependency of a subset of glioblastomas on autocrine DRD2 signaling. Analysis of specimens from patients treated with ONC201 (n = 15) showed an inverse correlation between the intensity of EGFR staining and clinical response. The median overall survival for patients with high and low EGFR staining was 162 and 373 days, respectively (p = 0.037). All patients who exhibited progression free survival beyond 200 days showed low to no EGFR expression. Conclusions: Our results suggest EGFR expression as a determinant of response to ONC201 in glioblastoma patients and should inform the design of future clinical trials.

CRISPR-mediated ablation of overexpressed EGFR in combination with sunitinib significantly suppresses renal cell carcinoma proliferation.

Receptor tyrosine kinases, such as VEGFR, PDGFR and EGFR, play important roles in renal cancer. In this study, we investigated EGFR knockout as a therapeutic approach in renal cell carcinoma (RCC). We showed that a renal cell carcinoma cell line (RC21) has higher expression of EGFR as compared to other frequently used cell lines such as HEK293, A549, Hela and DLD1. Ablation of EGFR by CRISPR/Cas9 significantly restrained tumor cell growth and activated the MAPK (pERK1/2) pathway. The VEGFR and PDGFR inhibitor, sunitinib, attenuated the expression of MAPK (pERK1/2) and pAKT induced by EGFR loss and further inhibited EGFR-/- cell proliferation. We showed that loss of EGFR eventually leads to resistance to SAHA and cisplatin. Furthermore, EGFR loss induced G2/M phase arrest and resulted in an increased resistance to TNF-related apoptosis-inducing ligand (TRAIL) in renal cell carcinoma. Thus, ablation of overexpressed EGFR by CRISPR/Cas9 alone or in combination with sunitinib may be a new treatment option for renal cell carcinoma.

Image-based ex vivo drug screen to assess targeted therapies in recurrent thymoma

ObjectivesThymoma is a rare malignancy derived from the thymic epithelial cells. No standard salvage treatments are available for recurrent thymoma and due to the low number of cases, alternative treatment regimens have been assessed only in small case series with varying success. The aim of this study was to use an image-based ex vivo drug screening strategy to assess efficacy of a large panel of anti-cancer agents for thymoma using patient derived tumor cells. Materials and MethodsVital tumor and tumor associated cells were used to assess the efficacy of 147 anti-cancer drugs including approved and experimental agents. Drug efficacy was analyzed at single cell resolution using image-based high content drug screening to assess tumor cell specific responses. Molecular profiling and histopathology was used to confirm the drug targets identified by the screen. ResultsThe ex vivo drug screen identified selective sensitivity of the cancerous epithelial thymoma cells to EGFR-, HDAC- and mTOR-inhibition. Histopathology confirmed high protein level expression of EGFR in the patient’s tumor. Patient was initiated treatment with Cetuximab resulting in stable disease after relapse on five different chemotherapy regimens. ConclusionThe results show that the image-based ex vivo therapy efficacy screening strategy can be used to identify patient and tumor relevant drug sensitivity patterns in thymoma. The results also warrant continued research on EGFR as a biomarker and therapy target in recurrent thymomas.

The role of epidermal growth factor receptor (EGFR) in the efficacy of neoadjuvant chemotherapy in triple-negative breast cancer patients

Background. Identification of predictive molecular markers of triple-negative breast cancer (TNBC) will enable the evaluation of the efficacy of neoadjuvant chemotherapy (NACT) and define optimum approaches for the prognosis of the disease course in TNBC patients. The aim of the study was to examine the correlation between the expression of the epidermal growth factor receptor (EGFR), its gene’s polymorphic variants and the neoadjuvant chemotherapy (NACT) efficacy in triple-negative breast cancer (TNBC) patients. Materials and methods. The study included 70 patients with triple-negative breast cancer, who had received 2-4 cycles of FAC and CAX regimens. The efficacy of the neoadjuvant chemotherapy was assessed according to the RECIST scale. The EGFR expression level in tumors before and after the NACT was evaluated with the help of immunohistochemistry. Genotypes for EGFR (rs2227983 and rs1468727) were detected by a real-time PCR. Results. It was found that NCT significantly decreases the EGFR expression level in the tumor ( p = 0.000). The research associates the objective clinical response as well as the pathological complete response with the low EGFR expression level ( p = 0.007 and p = 0.000 respectively). Patients carrying the EGFR CC mutant genotype of rs1468727 did not achieve a pathological complete response ( p = 0.042). In addition, patients with EGFRCC mutant genotype are more likely to have tumors with a high EGFR expression compared to EGFRTT wild-type genotype patients ( p = 0.047). Conclusion . The EGFR expression level in tumor tissue and the polymorphic variants of its gene in the rs1468727 locus can be considered as potential molecular markers with predictive significance in relation to the NACT efficacy in triple-negative breast cancer patients.

Cellular internalization of targeted and non-targeted delivery systems for contrast agents based on polyamidoamine dendrimers

New high-molecular-weight contrast agents based on polyamidoamine (PAMAM) dendrimers for targeted imaging of malignant tumors characterized by overexpression of human epidermal growth factor receptor (EGFR) and human alpha-fetoprotein receptor (RECAF) were designed. Conjugates of second (G2) and third (G3) generation polyamidoamine dendrimers with 1,4,7,10-tetraazocyclodecane-1,4,7,10-tetraacetic acid (DOTA) were obtained. The quantitative composition of the conjugates was determined by 1HNMR spectroscopy. It was shown that four out of the 16 terminal NH2 groups in G2-DOTA and nine out of the 32 groups in G3-DOTA were modified with DOTA. The morphology, size, and charge of the synthesized macromolecules were characterized by dynamic light scattering and electrophoresis. Gadolinium(III) was loaded into the conjugates and the Gd content was determined by atomic emission spectroscopy. For increasing the selectivity of accumulation in the tumor cells, two recombinant proteins able to bind selectively to EGFR and RECAF, namely, human recombinant epidermal growth factor (rEGF) and human recombinant 3rd domain of alpha-fetoprotein (3dAFPpG), were conjugated with G2 and G3 dendrimers. The conjugates containing vector molecules were mainly accumulated via clathrin-dependent endocytosis, whereas G2-DOTA and G3-DOTA were absorbed via caveolin-dependent endocytosis and macropinocytosis. The dendrimer conjugates with vector molecules were intensely accumulated in A549 cells characterized by high expression of EGFR (Herl) and RECAF, whereas the accumulation of conjugates in the control K562 cells (with low expression of Her1) and in the CD14− population of human unstimulated mononuclear white blood cells was insignificant. The 3dAFPpG-conjugated dendrimers were partly recycled. All synthesized conjugates had a rather low toxicity in the range of 350–450 µmol L−1 (IC50).