Factor Receptor Gene Copy Number Research Articles

EGFR gene copy number decreases during anti-EGFR antibody therapy in colorectal cancer

Epidermal growth factor receptor (EGFR) gene copy number (GCN) increase is associated with a favorable anti-EGFR antibody treatment response in RAS wild-type metastatic colorectal cancer. However, there are limited and comparative data regarding the EGFR GCN in primary colorectal cancer tumors and corresponding metastases or the effect of anti-EGFR antibody treatment on EGFR GCN in recurrent disease. In addition, little is known about the potential EGFR GCN changes during anti-EGFR therapy in comparison with other treatment regimens. EGFR GCN was analyzed by EGFR immunohistochemistry-guided silver in situ hybridization in primary and corresponding recurrent local or metastatic tumors from 80 colorectal cancer patients. GCN levels were compared between KRAS wild-type patients having received anti-EGFR therapy and patients having received other forms of treatment after primary surgery. The EGFR GCN decrease between primary and recurrent tumors was more pronounced among the anti-EGFR-treated patients than among patients not treated with anti-EGFR therapy (P = .047). None of the patients experiencing an EGFR GCN increase of at least 1.0 between the primary and recurrent tumors were treated with anti-EGFR antibodies. When including only patients with distant metastases, an EGFR GCN decrease of at least 1.0 was more common among the anti-EGFR-treated patients than among patients not treated with anti-EGFR therapy (P = .028). Our results suggest that anti-EGFR antibody treatment is associated with EGFR GCN decrease between the primary and recurrent colorectal adenocarcinomas, whereas no GCN change is observed among patients receiving other forms of treatment after primary surgery.

Associations between epidermal growth factor receptor and topoisomerase II-alpha gene copy number variations, human papillomavirus positivity, and cytologic analysis in cervical cell lesions.

Cervical cancer is the second most common gynecologic cancer worldwide. Human papillomavirus (HPV) infection is a leading etiological factor in cervical carcinoma.The aim of this study was to compare HPV positivity, EGFR and TOP2A gene copy number variations and cervical cytologic findings. The study group comprised 100 female volunteers between 21-64 years old. Cytologic analysis was performed using the liquid-based cytology technique. HPV DNA testing was performed in all cases. Copy number variations that belong to EGFR and TOP2A genes evaluated by using FISH analysis. Cytologic analysis of the cervical samples revealed abnormal findings in 13 of the 100 study subjects. ASCUS , LSIL, HSIL were determined in 8, 2 and 3 cases respectively as the result of cytologic analysis on all cases. Forty-one (41%) of the 100 women were HPV-positive. Chi-square analysis confirmed that HPV-positive women showed significantly more abnormal cytology (P = 0.035). EGFR deletion, TOP2A deletion and both EGFR and TOP2A deletion were determined in 1, 8 and 1 cases respectively. We found no statistical difference in abnormal cytologic findings between subjects with these gene deletions and subjects with normal gene copy numbers (P > 0.05 for both). No cases of amplification were determined for either gene. As a result, HPV positivity and the determination of changes that may occur in the gene copy number in patients with abnormal cytology can be an important tool on account of prognosis. Research with more patients may be suggested.

EGFR gene copy number as a predictive/biomarker for patients with non-small-cell lung cancer receiving tyrosine kinase inhibitor treatment: a systematic review and meta-analysis

Epidermal growth factor receptor (EGFR) gene copy number has been proposed as a candidate biomarker for predicting treatment response to EGFR tyrosine kinase inhibitors (EGFR-TKIs) in patients with advanced non-small-cell...

Changes in Epidermal Growth Factor Receptor Gene Copy Number during Oral Carcinogenesis.

Oral squamous cell carcinoma (OSCC) is a global healthcare problem associated with poor clinical outcomes. Early detection is key to improving patient survival. OSCC may be preceded by clinically recognizable lesions, termed oral potentially malignant disorders (OPMD). As histologic assessment of OPMD does not accurately predict their clinical behavior, biomarkers are required to detect cases at risk of malignant transformation. Epidermal growth factor receptor gene copy number (EGFR GCN) is a validated biomarker in lung non-small cell carcinoma. We examined EGFR GCN in OPMD and OSCC to determine its potential as a biomarker in oral carcinogenesis. EGFR GCN was examined by in situ hybridization (ISH) in biopsies from 78 patients with OPMD and 92 patients with early-stage (stages I and II) OSCC. EGFR ISH signals were scored by two pathologists and a category assigned by consensus. The data were correlated with patient demographics and clinical outcomes. OPMD with abnormal EGFR GCN were more likely to undergo malignant transformation than diploid cases. EGFR genomic gain was detected in a quarter of early-stage OSCC, but did not correlate with clinical outcomes. These data suggest that abnormal EGFR GCN has clinical utility as a biomarker for the detection of OPMD destined to undergo malignant transformation. Prospective studies are required to verify this finding. It remains to be determined if EGFR GCN could be used to select patients for EGFR-targeted therapies. Abnormal EGFR GCN is a potential biomarker for identifying OPMD that are at risk of malignant transformation. Cancer Epidemiol Biomarkers Prev; 25(6); 927-35. ©2016 AACR.

Epidermal growth factor receptor expression and gene copy number analysis in gastric carcinoma samples from Chinese patients.

Epidermal growth factor receptor (EGFR) expression and gene copy number have been observed to be associated with a positive clinical response to EGFR inhibitors. The present study aimed to evaluate EGFR expression and gene copy number in samples of gastric carcinoma (GC) from Chinese patients. EGFR expression and gene copy number were detected using immunohistochemistry and fluorescence in situ hybridization, in tissue array slides containing 150 individual samples of GC tissue. The association between EGFR status, clinicopathological features and overall patient survival was analyzed. Out of the 150 cases of GC evaluated, 63 (42.00%) demonstrated weak EGFR expression and 20 (13.33%) demonstrated EGFR overexpression. EGFR expression was observed to be associated with tumor location (P<0.05). Out of 104 cases of GC, which produced a clear FISH signal, 6 (5.77%) exhibited EGFR gene amplification and 5 (4.80%) exhibited balanced polysomy. Patients exhibiting GC, who demonstrated weak EGFR expression, EGFR overexpression or increased EGFR gene copy number, possessed an unfavorable prognosis. Multivariate analysis revealed that EGFR expression, tumor/node/metastasis stage and tumor location were potential independent unfavorable prognostic factors for GC patients. In conclusion, EGFR overexpression, gene amplification and polysomy were observed in GC patients and were associated with an unfavorable prognosis. Evaluation of EGFR status may therefore facilitate the identification of a subset of GC patients sensitive to treatment with EGFR-targeted therapies.

Association between epidermal growth factor receptor gene copy number and ERCC1, BRCA1 protein expression in Chinese patients with non-small cell lung cancer

Mutation in epidermal growth factor receptor (EGFR) gene may predict response to chemotherapy in non-small cell lung cancer (NSCLC). However, the correlation between EGFR gene copy status and protein levels of drug-resistant genes, such as excision repair cross-complementing 1 (ERCC1) and breast cancer 1 (BRCA1), remains unclear. We retrospectively analyzed formalin-fixed, paraffin-embedded tumor tissues from 109 Chinese patients with NSCLC. EGFR gene copy number was evaluated by fluorescence in situ hybridization (FISH), and protein levels of platinum-resistance-associated genes, including ERCC1 and BRCA1, were determined by immunohistochemical staining. High EGFR gene copy (EGFR FISH-positive) was found in 21.1% of the patients (amplification in 7.3% and high polysomy in 13.8%, respectively). Immunohistochemical analysis revealed that ERCC1 protein expression was not associated with clinicopathological factors, whereas a significantly higher BRCA1 positive rate was found in poorly differentiated tumors (P=0.02). Further association studies demonstrated that EGFR gene copy number status was not correlated with protein levels of ERCC1 or BRCA1; however, expression of ERCC1 was significantly associated with that of BRCA1 in this set of Chinese patients with NSCLC (P<0.001, r=0.484). Our study demonstrated that EGFR gene copy number status was not correlated with ERCC1 or BRCA1 protein expression, but ERCC1 protein levels were significantly correlated to BRCA1 protein expression levels in tumor tissues from Chinese patients with NSCLC.

EGFR Gene Copy Number as a Prognostic Marker in Colorectal Cancer Patients Treated with Cetuximab or Panitumumab: A Systematic Review and Meta Analysis

BackgroundThe epidermal growth factor receptor (EGFR) gene copy number (GCN) has been previously demonstrated to correlate with the clinical outcome of colorectal cancer (CRC) treated with anti-EGFR monoclonal antibodies (mAbs), although it remains controversial. We conducted a systematic review and meta-analysis to assess EGFR GCN as a potential biomarker of survival for patients with advanced CRC receiving treatment with anti-EGFR mAbs.MethodsWe systematically identified articles investigating EGFR GCN by fluorescent or chromogenic in situ hybridization or other detection techniques in patients with metastatic CRC treated with panitumumab or cetuximab, (last search: 10 August 2012). Eligible studies had to report on overall survival (OS), progression-free survival (PFS) or time-toprogression (TTP), stratified by EGFR GCN. Summary hazard ratios (HRs) were calculated using random-effects models.ResultsAmong 13 identified studies, 10 (776 patients, 302 with increased GCN), 8 (893 patients, 282 with increased GCN) and 3 (149 patients, 66 with increased GCN) were eligible for the OS, PFS and TTP meta-analyses, respectively. Increased EGFR GCN was associated with increased OS (HR = 0.62; 95% CI 0.50–0.77; P<0.001), PFS (HR = 0.65; 95% CI 0.47–0.89; P = 0.008) but not TTP (HR = 0.71; 95% CI 0.44–1.14; P = 0.157). It was also shown that EGFR GCN is independent of other factors such as KRAS status. Among those populations received second-line or higher treatment, increased EGFR GCN was strongly associated with improved survival (for OS, HR = 0.60; 95% CI 0.47–0.75; P<0.001; for PFS, HR = 0.59; 95% CI 0.47–0.75; P<0.001), whereas it did not influence survival in patients that received first-line therapy.ConclusionAmong the anti-EGFR-treated patients, increased EGFR GCN appears to be associated with improved survival outcomes. The effect on survival appears to be related to patients receiving the line of treatment.

Epidermal growth factor receptor (EGFR) gene copy number in colorectal adenoma-carcinoma progression

Adenomas are the easily identifiable precursors of the vast majority of colorectal cancers. Some of their morphological features, such as dysplasia, are predictive of their biological evolution toward adenocarcinomas. A large body of evidence has demonstrated that the epidermal growth factor receptor (EGFR) signaling pathway is commonly activated in colorectal cancer and EGFR-target therapies have improved the outcome for colorectal cancer patients. Nevertheless, the mechanisms underlying the role of EGFR in the adenoma-carcinoma sequence are not entirely clear. We retrospectively analyzed EGFR gene copy number by fluorescence in situ hybridization (FISH) in paraffin-embedded tissue from 215 patients recruited through a prospective colorectal cancer screening procedure and undergoing surgical colectomy. We observed that in human colorectal carcinogenesis, EGFR copy number increases progressively, from adenomas with high-grade dysplasia to locally advanced adenocarcinomas, through early invasive adenocarcinomas, suggesting that deregulation of EGFR may correlate with the malignant progression.

EGFR gene copy number as a predictive biomarker for the treatment of metastatic colorectal cancer with anti-EGFR monoclonal antibodies: a meta-analysis

BackgroundEpidermal growth factor receptor gene copy number (EGFR GCN) has been heavily investigated as a potential predictive biomarker for the treatment of metastatic colorectal cancer (mCRC) with anti-EGFR monoclonal antibodies (MAbs). The objective of this study was to systematically review current evidences on this issue.MethodsPubMed, EMBASE, The Cochrane Library, Chinese Biomedical Literature Database, Wanfang Data, and the conference abstracts of American Society of Clinical Oncology and European Society of Medical Oncology were comprehensively searched. Studies that reported the objective response rate (ORR), progression-free survival, and/or overall survival of mCRC patients treated with anti-EGFR MAbs, stratified by EGFR GCN status, were included. The effect measures for binary outcome (response) and time-to-event outcomes (progression-free survival and overall survival) were risk difference and hazard ratio, respectively. Statistical heterogeneity among the studies was assessed by the Cochran’s Q-test and the I2 statistic. If appropriate, a quantitative synthesis of data from different studies would be conducted with a random-effects model.ResultsNineteen eligible studies were identified. The criteria for increased EGFR GCN (GCN+) were highly inconsistent across different studies. The prevalence of GCN + ranged from 6.9% to 88.9%, and the difference in ORR between patients with GCN + and those with non-increased EGFR GCN (GCN-) varied from −28% to 84%. Because of the significant heterogeneity, no quantitative synthesis of data was performed. There was a general trend towards higher ORR in patients with GCN+. The difference in ORRs between patients with GCN + and those with GCN- was even greater in KRAS wild-type patients, while in KRAS mutated patients the difference often did not exist. Almost all patients with EGFR amplification responded to the treatment. However, the prevalence of EGFR amplification was generally low. Incomplete data on progression-free survival and overall survival seemingly supported the findings on ORR.ConclusionsAlthough increased EGFR GCN is generally associated with a better outcome of anti-EGFR MAbs treatment, especially among patients with wild-type KRAS, the clinical utility of this biomarker for selecting recipients of anti-EGFR MAbs would be severely limited by the heterogeneous scoring system and the poor reproducibility of EGFR GCN enumeration due to technical reasons.

Analysis of HER-3, insulin growth factor-1, nuclear factor-kB and epidermal growth factor receptor gene copy number in the prediction of clinical outcome for K-RAS wild-type colorectal cancer patients receiving irinotecan–cetuximab

A large proportion of colorectal cancer patients does not benefit from the use of anti-epidermal growth factor receptor (EGFR) treatment although in the absence of a mutation of the K-RAS gene. Preliminary observations suggested that HER-3, insulin-like growth factor-1 (IGF-1), nuclear factor-kB (NF-kB) and EGFR gene copy number (GCN) might identify patients not likely to benefit from anti-EGFR therapy. We tested the interaction between HER-3, IGF-1, NF-kB, EGFR GCN and K-RAS mutational analysis to verify the relative ability of these variables to identify a subgroup of patients more likely to benefit from EGFR-targeted treatment among those harbouring a K-RAS wild-type status. We retrospectively collected tumours from 168 patients with metastatic colorectal cancer treated with irinotecan-cetuximab. K-RAS was assessed with direct sequencing, EGFR amplification was assessed by chromogenic in situ hybridisation (CISH) and HER-3, IGF-1 and NF-kB were assessed by immunohistochemistry. In patients with K-RAS wild-type tumours, the following molecular factors resulted independently associated with response rate: HER-3 [odds ratio (OR)=4.6, 95% confidence interval (CI) 1.8-13.6, P=0.02], IGF-1 (OR=4.2, 95% CI 2-10.2, P=0.003) and EGFR GCN (OR=4.1, 95% CI 1.9-26.2, P=0.04). These factors also independently correlated with overall survival as follows: HER-3 [hazard ratio (HR)=0.4, 95% CI 0.28-0.85, P=0.008], IGF-1 (HR=0.47, 95% CI 0.24-0.76, P<0.0001) and EGFR GCN (HR=0.59, 95% CI 0.22-0.89, P=0.04). We believe that our data may help further composing the molecular mosaic of EGFR-resistant tumours. The role of HER-3, IGF-1 and CISH EGFR GCN should be prospectively validated in clinical trials investigating anti-EGFR treatment strategies in colorectal cancer patients.

Relationship between epidermal growth factor receptor gene mutation and copy number in Chinese patients with non-small cell lung cancer

Epidermal growth factor receptor (EGFR) gene mutation and copy number are useful predictive markers that guide the selection of non-small cell lung cancer (NSCLC) patients for EGFR-targeting therapy. This study aimed to investigate the correlation between EGFR gene mutation and copy number and clinicopathologic characteristics of Chinese patients with NSCLC. NSCLC specimens collected from 205 patients between November 2009 and January 2011 were selected to detect EGFR gene mutations with real-time polymerase chain reaction (RT-PCR) and to detect EGFR gene copy number with fluorescence in situ hybridization (FISH). EGFR mutations primarily occurred in females, non-smokers, and patients with adenocarinomas (all P < 0.001). Tissues from 128 (62%) patients were FISH-positive for EGFR, including 37 (18%) with gene amplification and 91 (44%) with high polysomy. EGFR gene mutation was correlated with FISH-positive status (R = 0.340, P < 0.001). Multivariate analysis showed that not smoking (OR = 5.910, 95% CI = 2.363–14.779, P < 0.001) and having adenocarcinoma (OR = 0.122, 95% CI = 0.026–0.581, P = 0.008) were favorable factors for EGFR gene mutation. These results show a high frequency of EGFR FISH positivity in NSCLC tissues from Chinese patients and a significant relevance between EGFR gene mutations and FISH-positive status. Among the FISH-positive samples, EGFR gene mutation occurred more frequently in samples with gene amplification compared to those with high polysomy, suggesting that EGFR mutation and gene amplification should be used as clinical decision parameters to predict response to EGFR-targeting therapy.

Correlation of epithelial-mesenchymal transition markers with clinicopathologic parameters in adenocarcinomas and squamous cell carcinoma of the lung.

Epithelial-mesenchymal transition (EMT) is characterized by the loss of epithelial cell junction proteins and the gain of mesenchymal markers. The aim of this study was to analyze the associations between the EMT-related markers vimentin, E-cadherin, β-catenin, slug, snail, and twist1 and clinicopathologic parameters as well as epidermal growth factor receptor (EGFR) gene copy number and protein expression in non-small cell lung carcinoma (NSCLC). Fifty-nine squamous cell carcinomas (SCC) and 43 adenocarcinomas (AD) were immunohistochemically examined for respective EMT markers and for EGFR, using the EGFR PharmDx kit (Dako) for protein expression and automated silver enhanced in situ hybridization (SISH) for copy number. Vimentin expression in tumor epithelia was significantly higher in AD samples than in SCC samples (P=0.015). Among AD samples, vimentin expression was positively correlated with histologic grade (2 vs. 3; P=0.021) and exhibited a tendency toward a positive correlation with pTNM stage (I vs. II-IV; P=0.052). EGFR gene copy number was positively correlated with EGFR protein expression among both AD samples (P=0.008) and SCC samples (P=0.042), with EGFR protein expression being significantly higher in SCC samples compared with AD (P=0.038). Among AD samples, EGFR protein expression was associated with higher cytoplasmic expression of β-catenin (P=0.031). Among SCC samples, EGFR protein expression was negatively correlated with nuclear expression of β-catenin (P=0.033) but positively with nuclear slug (P=0.021). The expression pattern of EMT markers in AD suggests that vimentin is a possible immunohistochemical predictor of tumor progression.

A Phase II Study of Gefitinib for Aggressive Cutaneous Squamous Cell Carcinoma of the Head and Neck

To determine the disease control rate and toxicity of treating patients with aggressive cutaneous squamous cell carcinoma (CSCC) with neoadjuvant gefitinib. A prospective phase II clinical trial evaluating neoadjuvant gefitinib given prior to standard treatment with surgery and/or radiotherapy. Patients with stable disease after one cycle received escalated doses. Patients who responded were given gefitinib during radiation therapy, as well as maintenance therapy after definitive treatment. We analyzed the correlation between epidermal growth factor receptor (EGFR) expression, mutation status, and gene copy number on available tissue samples and clinical response. Twenty-three patients were accrued and 22 patients were evaluable for response prior to definitive local treatment; complete responses were attained by 18.2% of patients and partial responses by 27.3%. Grades 2 to 3 toxicities were observed in 59.1% of patients experiencing class-specific effects during induction therapy. After induction, 11.8% underwent surgery alone, 17.6% had definitive radiation, 11.8% were treated with radiation and concurrent gefitinib, and 47% had surgery with postoperative radiation and concurrent gefitinib. Median follow-up for the censored observations was 32 months. Two-year overall, disease-specific, and progression-free survival rates were 72.1%, 72.1%, and 63.6%, respectively. No EGFR-activating mutations were identified in tumor samples available from 10 patients. No associations between EGFR correlative studies and patient outcomes were identified. Gefitinib, in the neoadjuvant setting, was active and well tolerated in patients with aggressive CSCC and did not interfere with definitive treatment. In view of the 18% complete response rate we observed, EGFR tyrosine kinase inhibitors should be further explored in the treatment of aggressive CSCC.

Standardisation of EGFR FISH in colorectal cancer: results of an international interlaboratory reproducibility ring study

AimsEpidermal growth factor receptor (EGFR) gene copy number evaluated by fluorescence in situ hybridisation (FISH) can discriminate among KRAS wild-type patients those with better outcome to EGFR-targeted therapy in metastatic...

PP 22 Analysis of HER-3, insulin-growth factor-1 (IGF-1), nuclear factor k-B (NF-kB) and epidermal growth factor receptor (EGFR) gene copy number (GCN) in the prediction of clinical outcome for K-RAS wild type colorectal cancer patients receiving irinotecan–cetuximab

PP 22 Analysis of HER-3, insulin-growth factor-1 (IGF-1), nuclear factor k-B (NF-kB) and epidermal growth factor receptor (EGFR) gene copy number (GCN) in the prediction of clinical outcome for K-RAS wild type colorectal cancer patients receiving irinotecan–cetuximab

9132 POSTER Efficacy Outcomes in First-line Treatment of Advanced NSCLC With Gefitinib (G) vs Carboplatin/paclitaxel (C/P) by Epidermal Growth Factor Receptor (EGFR) Gene-copy Number Score and by Most Common EGFR Mutation Subtypes – Exploratory Data From IPASS

9132 POSTER Efficacy Outcomes in First-line Treatment of Advanced NSCLC With Gefitinib (G) vs Carboplatin/paclitaxel (C/P) by Epidermal Growth Factor Receptor (EGFR) Gene-copy Number Score and by Most Common EGFR Mutation Subtypes – Exploratory Data From IPASS

Relationship between epidermal growth factor receptor gene copy number and protein expression in oral cavity squamous cell carcinoma

This study was designed to explore the relationship between epidermal growth factor receptor (EGFR) copy number and EGFR protein expression in oral cavity squamous cell carcinoma (OSCCs) in Taiwan. A total of 160 oral cavity squamous cell carcinomas were examined for EGFR protein overexpression using immunohistochemistry and for copy number using a fluorescence in situ hybridization (FISH) assay. Overexpression and increased gene copy numbers of EGFR were found in 75 (46.88%) and 50 (31.25%) cases, respectively. The concordance rate for EGFR gene amplification and protein overexpression was 100%. EGFR overexpression was associated with a poor prognosis both in terms of disease-free survival (DFS) and overall survival (OS). On the other hand, the association between an increase in EGFR gene copies and DFS or OS was insignificant. This was despite the observed significant associations between gene copy number and tumor stage, depth of tumor invasion, lymph node metastasis, bone invasion and perineural invasion. EGFR protein overexpression is closely related to EGFR copy number. Standard methodological and interpretation criteria need to be established that allows EGFR copy number combined with EGFR protein expression to be determined in a manner that allows individualized EGFR targeted therapy in OSCC patients.

EGFR gene copy number, ERCC1 and BRCA1 protein expression and their relationship in non-small cell lung cancer

To evaluate the expression of epidermal growth factor receptor (EGFR) gene copy number and the expression of ERCC1 and BRCA1 proteins in patients with non-small-cell lung cancer (NSCLC) and the correlation between them. The status of EGFR gene copy number was determined by in situ hybridization (FISH), and the expression of ERCC1 and BRCC1 proteins was examined by immunohistochemistry (IHC). The relationship of EGFR gene copy number with the expression of ERCC1 and BRCA1 and the clinical pathologic features were analyzed. FISH-positive EGFR expression was identified in 40 of 166 samples (24.1%). More FISH-positive EGFR in the female than male patients (31.9% vs. 18.6%, P = 0.048), and non-smoker than smoker (32.8% vs. 16.7%, P = 0.045). FISH-positive EGFR was not associated with age, pathological type, clinical stage and metestasis status (P > 0.05). The expression of ERCC1 protein was identified in 60 of 132 samples (45.5%). The expression of ERCC1 protein varied significantly in tumors of different pathological types (P = 0.046), but not associated with age, gender, clinical stage, metestatic status and smoking status (P > 0.05). The expression of BRCA1 protein was identified in 46 of 131 samples (35.1%). The expression of BRCA1 was not associated with age gender, pathological type, clinical stage, metestatic ststus and smoking status (P > 0.05). There was a moderate correlation between the expressions of ERCC1 and BRCA1 (r = 0.449, P < 0.001), but EGFR gene copy number was not correlated with the expression of ERCC1 or BRCA1 protein. FISH-positive EGFR expression is associated with gender and smoking status, but not correlated with the expression of ERCC1 and BRCA1 proteins. There is a moderate correlation between the expressions of ERCC1 and BRCA1.

Relationship between Epidermal Growth Factor Receptor Status, p16INK4A, and Outcome in Head and Neck Squamous Cell Carcinoma

Human papilloma virus (HPV) infection is a powerful prognostic biomarker in head and neck squamous cell carcinoma (HNSCC). Increased epidermal growth factor receptor (EGFR) gene copy number and protein expression have been reported to be negative predictors of outcome. This study examined the relationship between HPV status, EGFR gene copy number, EGFR protein expression, and clinical outcome in HNSCC patients treated with chemoradiation. HPV status was determined using p16(INK4A) immunohistochemistry (IHC), EGFR gene copy number was evaluated with FISH, and EGFR protein expression by IHC in 212 subjects. EGFR FISH was positive in 41 of 204 (20%) patients and was negatively correlated with failure-free survival (FFS; HR = 1.84, P = 0.027) and overall survival (OS; HR = 1.78, P = 0.082). For p16(INK4A), 85 of 200 (42.5%) patients were found to be p16 positive, including 75 of 131 (57%) with oropharyngeal cancer. Patients with p16-positive oropharyngeal cancer had significantly improved FFS (HR = 0.28, P < 0.001) and OS (HR = 0.31, P = 0.002). Only 2 of 126 (1.6%) oropharyngeal cancer patients were found to be p16+/EGFR FISH+. EGFR IHC was positive in 81 of 93 (87%) of patients and was associated with poorer FFS (HR = 1.98, P = 0.35) and OS (HR = 2.52, P = 0.22). Increased EGFR gene copy number is largely restricted to p16(INK4A)-negative oropharyngeal cancer. Although p16(INK4A) and EGFR FISH are both predictive of outcome in univariate analyses, only p16(INK4A) remains independently predictive. Knowledge of HPV and EGFR status can have implications for treatment options and prognosis in HNSCC.

Standardization of EGFR FISH in colorectal cancer: Results of an international, interlaboratory reproducibility ring study.

3597 Background: Epidermal growth factor receptor (EGFR) gene copy number (GCN) evaluated by fluorescence in situ hybridization (FISH) has been shown to discriminate among KRAS wild type patients those with better outcome to EGFR-targeted therapy in metastatic colorectal cancer (CRC), further enhancing selection of patients. Nevertheless, enumeration of gene copies in CRC is challenging and the lack of analytical standardisation has limited incorporation of the test into the clinical practice. Methods: A ring study was designed in order to assess EGFR FISH inter-laboratory consensus among 5 pathology reference centres located in Belgium, Italy, Switzerland, and United States. A set of 12 CRC samples circulated among laboratories and samples were scored according to commonly agreed guidelines. Reproducibility was quantified using the standard error of measurement (SEM). Results: A SEM of 0.865 and a within-subject coefficient of variation (WSCV) of 26.8% for mean EGFR gene per nuclei and a SEM of 0.235 and a WSCV of 19.4% for the mean EGFR gene/CEP7 ratio were observed, indicating low level of agreement. Measurement of the fraction of cells displaying chromosome 7 polysomy within tumor specimens was also performed, showing WSCV of 46.6%, 34.0% and 51.0% for % of cells displaying ≤2, ≥3 and ≥4 EGFR signals, respectively. Among different slides of the same specimen, the WSCV was 6.1% for mean EGFR gene per nuclei and 3.9% for mean of EGFR gene/CEP7 ratios and 10.5%, 12.4% and 24.0% for the % of cells displaying ≤2, ≥3 and ≥4 EGFR signals, respectively. Conclusions: Molecular diagnosis of EGFR GCN by FISH varied largely among experienced international pathology centres, with fluctuations covering the whole range of proposed cut-offs of predictive usefulness from literature. Definition of a detailed scoring system and implementation of a comprehensive training program for molecular diagnostic laboratories are therefore necessary before including the test into clinical practice as a standard assay for predicting outcome to EGFR-targeted therapies in metastatic CRC.