Clinicopathologic features and prognostic implications of epidermal growth factor receptor (EGFR) gene copy number and protein expression in non-small cell lung cancer

Increased epidermal growth factor receptor (EGFR) gene copy numbers and mutations predict sensitivity to EGFR tyrosine kinase inhibitor in non-small cell lung cancer (NSCLC). However, the clinicopathologic features of EGFR gene copy status in NSCLC remain unclear. We retrospectively analyzed 262 cases of NSCLC, including 135 squamous cell carcinomas (SCC) and 112 adenocarcinomas (ADC), for which paraffin blocks of the resected primary lung mass were available. None had received EGFR-targeted therapy. EGFR gene copy number was evaluated using fluorescence in situ hybridization (FISH), and EGFR expression was determined immunohistochemically using a tissue microarray. A high EGFR gene copy (EGFR FISH-positive) was found in 30.2% of the cases (amplification in 11.1% and high polysomy in 19.1%). There was no significant difference in EGFR FISH status with respect to SCC and ADC histology. The EGFR FISH-positive rate was higher in non-smokers than in smokers in the multivariate analysis (p=0.012). EGFR expression was significantly associated with a high EGFR gene copy and SCC histology (p=0.000). In the univariate survival analysis, EGFR FISH-positivity predicted worse survival in SCC (p=0.059), especially stage I SCC (p=0.04). EGFR amplification was associated with a shorter survival in node-positive SCC (p=0.015). However, the EGFR gene copy number or protein expression had no influence on the prognosis of ADC. In conclusion, the EGFR FISH-positive rate in Korean patients with NSCLC was similar to rates in Western populations, unlike the higher frequencies of EGFR mutation in East Asians. A high EGFR gene copy number was significantly more common in non-smokers, as were EGFR mutations. A high EGFR gene copy number predicted worse survival in SCC; therefore, the prognostic implications of the EGFR gene and protein should be analyzed in the context of histology and staging in NSCLC.     

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 between computed tomography findings and epidermal growth factor receptor and KRAS gene mutations in patients with pulmonary adenocarcinoma

We examined the correlation between computed tomography (CT) findings and the incidence of epidermal growth factor receptor (EGFR) and KRAS mutations in lung adenocarcinoma. We analyzed the tumors of 136 patients with surgically resected primary lung adenocarcinoma. CT scans were evaluated for the presence of ground grass opacity (GGO), spiculation and the maximum diameter of the tumor was measured. SMart Amplification Process (ver. 2) was used to detect the presence of EGFR and KRAS mutations. EGFR and KRAS mutations were found in 56 (41.1%) and 25 (18.4%) of the 136 cases, respectively. Although no significant association was found between GGO and EGFR mutations (p=0.07), the EGFR mutation occurred more frequently in male patients with GGO than in those without GGO (p=0.04). The KRAS mutation occurred more frequently in patients whose tumor diameter was ≥ 31 mm than in those whose tumor diameter was <30 mm (p=0.003). Evaluation of CT findings may be helpful for determining the presence of EGFR and KRAS mutations, particularly when it is not possible to obtain a tumor specimen.     

非小细胞肺癌表皮生长因子受体基因拷贝数与核苷酸切除修复交叉互补基因1和乳腺癌易感基因1的表达及其相关性分析

目的 探讨表皮生长因子受体(EGFR)基因拷贝数与铂类耐药相关基因核苷酸切除修复交叉互补基因1(ERCC1)和乳腺癌易感基因1(BRCA1)在非小细胞肺癌(NSCLC)中的表达情况,以及三者之间的相关性.方法 用荧光原位杂交(FISH)技术检测EGFR基因的扩增情况,应用免疫组化SP法检测132例NSCLC患者标本ERCC1和BRCA1蛋白的表达,并进一步分析EGFR基因拷贝数、ERCC1和BRCA1的蛋白表达与患者临床病理特征的关系.结果 EGFR基因FISH阳性率为24.1% (40/166).女性的扩增阳性率高于男性(31.9%和18.6%,P=0.048);无吸烟史的患者扩增阳性率明显高于有吸烟史的患者(32.8%和16.7%,P=0.045);基因拷贝数增加与患者的年龄、临床分期、病理类型及有无淋巴结或远处转移均无关.ERCC1蛋白表达阳性率为45.5% (60/132),在不同的病理类型分布比较,差异有统计学意义(P=0.046),与患者的性别、年龄、临床分期、有无淋巴结或远处转移及吸烟状况均无关.BRCA1蛋白表达阳性率为35.1%(46/131),与患者的性别、年龄、临床分期、病理类型、淋巴结或远处转移及吸烟状况无关.ERCC1与BRCA1的蛋白表达呈中等相关(r=0.449,P＜0.001),与EGFR扩增无相关性(P=0.785);BRCA1表达与EGFR基因拷贝数状态无相关性(P=0.143).结论 在肺癌的个体化治疗中,检测EGFR基因拷贝数、ERCC1和BRCA1的表达对靶向治疗和铂类药物化疗的选择具有重要的临床指导意义.

Abstract：

Objective 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. Methods 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. Results 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. Conclusions 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.     

18F-FDG PET/CT对肺腺癌EGFR突变状态的预测价值研究*

目的 :本研究旨在阐明18F-FDG PET/CT对于肺腺癌EGFR突变状态的预测价值。 方法: 选取2016年6月1日至2017年7月31日于天津医科大学肿瘤医院行术前PET/CT扫描及术后基因检测的肺腺癌患者117例。分析患者临床特征、肺内原发病灶SU? Vmax、SUVmean、SUVpeak及肺内原发病灶与纵隔血池SUVmax比值与EGFR突变状态间的关系。采用受试者工作特征曲线分析不同PET代谢参数对EGFR突变状态的预测能力。 结果: 117例患者中,EGFR突变型患者65例。未吸烟患者EGFR突变率高于吸烟患者（62.5% vs. 40.5%,P=0.026）。EGFR突变型肺内原发病灶SUVmax、SUVmean、SUVpeak及肺内原发病灶与纵隔血池SU? Vmax比值均明显低于野生型（SUVmax:8.02±3.96 vs. 10.31±5.80,P=0.017;SUVmean:4.97±2.51 vs. 6.45±3.68,P=0.015;SUVpeak:6.03±3.22 vs. 8.06±5.01,P=0.013;T/N:5.08±3.01 vs. 6.91±4.40,P=0.012）。不同诊断标准间诊断效能无明显差异。 结论: EGFR突变型患者肺内原发病灶FDG摄取值低于野生型患者,因此,18F-FDG PET/CT对肺腺癌患者EGFR突变状态的预测具有一定指导意义。      

Evaluation of F-18 fluorodeoxyglucose (FDG) PET scanning for pulmonary nodules less than 3 cm in diameter, with special reference to the CT images

BACKGROUND: While pulmonary nodules can be substantially divided into solid and ground-glass opacity (GGO) ones on CT image, they have different biological natures which could cause false positive or false negative to diagnose malignancy on positron emission tomography with fluorodeoxyglucose (FDG-PET). To determine the effectiveness of PET for small pulmonary nodules, the nodules were classified into solid and GGO ones, of which results were compared with the data of PET scans. The lower limit size of nodules for PET imaging was also evaluated. METHODS: Prospective FDG-PET scans were undertaken for 136 non-calcified nodules less than 3 cm in diameter. CT density histograms were made for each nodule to classify into solid and GGO ones. RESULTS: Eighty-one nodules were malignant and 55 were benign. All of the 20 nodules less than 1 cm in diameter (n = 8 in malignant, n = 12 in benign), were negative on PET regardless of the histology. In the 116 nodules 1-3 cm in diameter (n = 73 in malignant, n = 43 in benign), there were 15 false negative and 15 false positive nodules, with a sensitivity of 79% and specificity of 65%. CT density histograms showed 101 solid nodules (n = 63 in malignant, n = 38 in benign) and 15 GGO nodules ( n = 10 in malignant, n = 5 in benign). All of the 10 malignant nodules with GGO images were histologically well-differentiated adenocarcinoma and 9 of them (90%) were false negative on PET. Four of the 5 (80%) benign nodules with GGO images were focal pneumonia with well-preserved air spaces, causing false positive on PET. Sensitivity and specificity for nodules with GGO images were 10 and 20%, respectively, which were significantly lower than 90 and 71% for nodules with solid images (P < 0.001). CONCLUSION: Pulmonary nodules which are less than 1cm in size or show GGO images on CT cannot be evaluated accurately by PET.     

Increased epidermal growth factor receptor gene copy number detected by fluorescence in situ hybridization associates with increased sensitivity to gefitinib in patients with bronchioloalveolar carcinoma subtypes: a Southwest Oncology Group Study.

PURPOSE: Bronchioloalveolar carcinoma (BAC) and adenocarcinomas with BAC features seem to be increasing in incidence, particularly in younger, never-smoking women. Epidermal growth factor receptor (EGFR) inhibitors demonstrated response rates of 20% to 30% in patients with advanced BAC subtypes, but selection methods for patient therapy are not established. PATIENTS AND METHODS: EGFR and HER2 gene copy numbers were assessed by fluorescence in situ hybridization (FISH) in 81 patients treated with gefitinib 500 mg/d (Southwest Oncology Group protocol S0126) and were correlated to treatment outcome. Tumors were classified into two main strata: FISH-positive (high polysomy/gene amplification) and FISH-negative (disomy/low polysomy). RESULTS: In 81 patients, the median survival time for EGFR/FISH-negative patients was 8 months and not yet reached for FISH-positive patients (but approaching 18 months; hazard ratio [HR] = 2.02; P = .042). Median progression-free survival time for EGFR/FISH-positive patients was 9 months versus 4 months for the FISH-negative patients (HR = 1.67; P = .072). In multivariate analysis, EGFR copy number by FISH remained a significant predictive factor for survival after accounting for smoking status, sex, histology, and performance status. Fifty-five patients were evaluated for response using Response Evaluation Criteria in Solid Tumors Group, and 12 of 19 EGFR/FISH-positive patients (63%) demonstrated disease control versus 14 (39%) of 36 patients in the FISH-negative group (P = .087). No association was found between HER2 gene copy number and response (n = 39 patients) or survival (n = 56 patients; P > .10). CONCLUSION: Increased EGFR gene copy number detected by FISH is associated with improved survival after gefitinib therapy in patients with advanced BAC, suggesting FISH methodology can be used to assess survival potential in patients treated with EGFR tyrosine kinase inhibitors.     

Improvement in preoperative staging of gastric adenocarcinoma with positron emission tomography

BACKGROUND: Positron emission tomography (PET) with 18- fluorodeoxyglucose (FDG) has been used to both detect and stage a variety of malignancies. The current study examined the value of PET for preoperative staging of gastric adenocarcinoma. METHODS: Sixty-eight patients (49 males and 19 females) with gastric adenocarcinoma, who were referred for preoperative FDG-PET scans, were enrolled in this study. The patients underwent spiral-computed tomography (CT) within 1 week of referral. The final diagnosis in all patients was made by histologic and surgical findings. For quantitative PET analysis, the regional tumor FDG uptake was measured by the standardized uptake value (SUV). RESULTS: For the primary tumor of a gastric adenocarcinoma, PET demonstrated an increased uptake in 64 of 68 patients (sensitivity, 94%), with a mean SUV of 7.0 (range, 0.9-27.7). A comparison of FDG uptake and clinicopathologic features showed significant association between FDG uptake and macroscopic type, tumor size, lymph node metastasis, histologic type, and TNM stage. The PET scan had a similar accuracy with that of CT for diagnosing local and distant lymph node metastases as well as peritoneal status. In assessing local lymph node status, however, PET had a higher specificity than CT (92% vs. 62%, P = 0.000). Moreover, PET had additional diagnostic value in 10 (15%) of 68 patients by upstaging 4 (6%) and downstaging 6 (9%) patients. PET combined with CT was more accurate for preoperative staging than either modality alone (66% vs. 51%, 66% vs. 47%, respectively; P = 0.002). CONCLUSIONS: FDG-PET improves the preoperative TNM staging of gastric adenocarcinoma. Based on its superior specificity, FDG-PET can facilitate the selection of patients for a curative resection by confirming a nodal status identified by CT.     

The standardized uptake value for F-18 fluorodeoxyglucose is a sensitive predictive biomarker for cervical cancer treatment response and survival

BACKGROUND: The objective of this study was to evaluate cervical tumor uptake of F-18 fluorodeoxyglucose (FDG) measured as the maximal standardized uptake value (SUV(max)) by positron emission tomography (PET) and its association with treatment response and prognosis in patients with cervical cancer. METHODS: The study population consisted of 287 patients with stage IA2 through IVB cervical cancer who underwent pretreatment FDG-PET studies. SUV(max), tumor volume, and sites of lymph node metastasis were recorded. Therapy included surgery, chemoradiation, or palliation. RESULTS: The mean SUV(max) was 11.4 (range, 1-50.4). The mean tumor volume by stage was 42.1 cm(3) for stage I tumors (using International Federation of Gynecology and Obstetrics [FIGO] staging criteria), 63.7 cm(3) for stage II tumors, 129.2 cm(3) for stage III tumors, and 166.2 cm(3) for stage IV tumors. There was no correlation between tumor volume and SUV(max) (correlation coefficient [R(2)] = 0.01). No significant difference in SUV(max) was observed between squamous histology (n = 247 patients) and nonsquamous histology (n = 40 patients; P = .089). Higher SUV(max) was associated with an increased risk of lymph node metastasis at diagnosis (P = .0009). A Cox proportional-hazards model for death from cervical cancer was used to evaluate tumor histology, lymph node metastasis, tumor volume, and SUV(max). The results indicated that SUV(max) was the only significant independent factor (P = .0027). Three prognostic groups were established using SUV(max). The overall survival rates at 5 years were 95% for an SUV(max) 5.2 and 13.3 (P < .0001). Increasing SUV(max) was associated with persistent abnormal FDG uptake in the cervix on 3-month FDG-PET studies in 238 patients who received curative chemoradiation (P = .04). CONCLUSIONS: The SUV(max) of the cervical tumor at diagnosis was a sensitive biomarker of treatment response and prognosis for patients with cervical cancer.     

Prognostic significance of [(18)F]fluorodeoxyglucose uptake on positron emission tomography in patients with pathologic stage I lung adenocarcinoma

BACKGROUND: [(18)F]Fluoro-2-deoxyglucose uptake on positron emission tomography (FDG-PET) has been frequently used for diagnosis and staging of lung cancer. The prognostic significance of FDG uptake on PET was evaluated in patients with pathologic Stage I lung adenocarcinoma (tumor stages were based on the TNM classification of the International Union Against Cancer). METHODS: Disease-free survival of 98 patients with pathologic Stage I lung adenocarcinoma who were treated by curative resection was examined in relation to sex, age, histologic grade of differentiation, surgical procedure, tumor stage, and FDG uptake measured as the maximum standardized uptake value (SUV). RESULTS: Sixty-three patients were had Stage IA disease and 35 patients had Stage IB disease. Six patients each with Stage IA and Stage IB disease developed disease recurrence after a mean postsurgical follow-up period of 31 months. Ten (23%) of the 43 patients with SUV > or = 3.3 developed a recurrence compared with 2 (4%) of the 55 patients with SUV < 3.3 (P = .020). Ten (20%) of the 51 patients with moderately or poorly differentiated adenocarcinoma developed disease recurrence, compared with 2 (4%) of the 47 patients with well-differentiated adenocarcinoma (P = .056). Multivariate analysis demonstrated that histologic grade of differentiation was not correlated with the frequency of tumor recurrence (P = .286), whereas SUV was found to be marginally correlated (P = .079). CONCLUSIONS: FDG uptake appears to be predictive of disease-free survival in patients with Stage I lung adenocarcinoma. FDG uptake could yield important information for determining the likely value of postoperative adjuvant chemotherapy in such patients.     

Impact of epidermal growth factor receptor (EGFR) kinase mutations, EGFR gene amplifications, and KRAS mutations on survival of pancreatic adenocarcinoma

BACKGROUND: Erlotinib, in combination with gemcitabine, has shown clinical benefits in pancreatic adenocarcinoma patients. The presence of EGFR mutations and increased EGFR copy numbers in pancreatic adenocarcinoma was explored. METHODS: Sixty-six pancreatic cancer patients were included in the analysis. The EGFR mutation was analyzed by DNA sequencing of exons 18-21 in the tyrosine kinase domain. KRAS mutation was analyzed by sequencing codons 12, 13, and 61. Quantitative real-time polymerase chain reaction was performed to analyze the copy number of EGFR. RESULTS: In the current study the EGFR mutation was harbored in only 1 (1.5%) of the 66 inoperable or metastatic pancreatic adenocarcinoma patients. Amino acid substitution was detected in exon 20 of the EGFR gene. Increased EGFR copy numbers (> or =3.0 per cell) were detected in 26 (41%) patients. There was only 1 patient, who had a highly increased EGFR copy number (> or =6.0 per cell), who died, 2.1 months from the date of diagnosis. The EGFR amplification did not significantly influence survival in pancreatic adenocarcinoma patients (P = .935). Thirty-two (49%) of the 65 pancreatic adenocarcinomas examined harbored a point mutation in codons 12 (n = 31) and 61 (n = 1) of the KRAS gene. The presence of a point mutation in codon 12 adversely influenced survival of pancreatic cancer patients (P = .030). CONCLUSIONS: The incidence of somatic mutations in the tyrosine kinase domains of EGFR was very low and the increased gene copy number of EGFR did not significantly influence survival.     

非小细胞肺癌组织MET和EGFR基因扩增与预后相关性分析

目的：探讨非小细胞肺癌（non-smfllleelllungcancer,NSCLC）患者中肝细胞生长因子受体（MET）基因和表皮生长因子受体（epidermalgrowthfactorreceptor,EGFR）基因扩增与临床病理特征及预后的关系。方法：回顾分析唐山市协和医院（48例）和唐山市人民医院（109例）2001—01—2007—01手术切除的NSCLC石蜡包埋标本157例。应用荧光原位杂交（fluorescenceinsituhybridization,FISH）检测NSCLCMET、EGFR基因扩增情况,并结合临床病理资料进行统计分析。应用SPSS16．0进行统计分析,Kaplan-Meier模型分析中住生存期（overallsurvival,0S）,Log—rank检验比较生存曲线,多因素分析采用Cox回归模型。结果：157例NSCLC患者标本中,EGFR基因扩增70例（44．6％）。EGFR基因扩增与年龄、性别、吸烟状态、组织类型和TNM分期无关,P〉0．05。157例NSCLC患者标本中,MET基因扩增25例（15．9％）。EGFR扩增患者MET扩增率为22．9％,高于无EGFR扩增患者MET扩增率10．3％,P=0．033。MET基因扩增与年龄、性别、吸烟状态、组织类型和TNM分期无关,P〉0．05。Kaplan-Meier生存分析显示,I＋Ⅱ期中位生存期为51个月,明显高于Ⅲ＋Ⅳ期中位生存期29个月,P=0．001。EGFRFISH阳性患者中位0s为33个月与EGFRFISH阴性患者中位0s39个月比较,差异无统计学意义,P=0．495。METFISH阳性患者中位0S为29个月,低于METFISH阴性患者37个月,P=0．044。患者0s与病理类型、年龄、性别和吸烟状态无相关性,P〉0．05。多因素分析显示,临床分期、MET基因扩增与OS有关（相对危险度为12．573、6．892,P值分别为0．015、0．018）。结论：临床Ⅲ＋Ⅳ期和MET基因扩增NSCLC患者预后不良,EGFR基因扩增与NSCLC患者预后无关。     

肺腺癌中表皮生长因子受体基因突变与拷贝数的检测分析

目的检测肺腺癌组织中表皮生长因子受体（EGFR）19、21外显子基因突变和拷贝数,分析EGFR基因突变和拷贝数变化的相关性。方法应用荧光定量PCR技术和荧光原位杂交（FISH）技术分别检测58例肺腺癌患者肿瘤组织中的EGFR基因突变和基因扩增,用X^2检验进行数据分析。结果58例肺腺癌患者组织中,EGFR19、21外显子突变率为43．1％（25／58）,其中2例存在两种类型的突变。EGFR基因拷贝数增加阳性率为51．7％（30／58）,包括8例扩增,22例高多体扩增。不同分化程度的肺腺癌组织间,扩增阳性率差异无统计学意义（P〉0．05）,低分化癌的突变率低于高、中分化癌（P〈O．05）。EGFR基因突变与EGFR基因拷贝数之间显著相关（P〈0．01）。结论肺腺癌组织具有较高的EGFR基因突变率和扩增阳性率;联合检测EGFR基因拷贝数和基因突变,更有利于靶向药物的筛选。     

Relationships between EGFR Mutation Status of Lung Cancer and Preoperative Factors - Are they Predictive?

Background: The epidermal growth factor receptor (EGFR) mutation status of lung cancer is importantbecause it means that EGFR-tyrosine kinase inhibitor treatment is indicated. The purpose of this prospectivestudy is to determine whether EGFR mutation status could be identified with reference to preoperative factors.Materials and Methods: One hundred-forty eight patients with lung cancer (111 adenocarcinomas, 25 squamouscell carcinomas and 12 other cell types) were enrolled in this study. The EGFR mutation status of each lungcancer was analyzed postoperatively. Results: There were 58 patients with mutant EGFR lung cancers (mutantLC) and 90 patients with wild-type EGFR lung cancers (wild-type LC). There were significant differences ingender, smoking status, maximum tumor diameter in chest CT, type of tumor shadow, clinical stage betweenmutant LC and wild-type LC. EGFR mutations were detected only in adenocarcinomas. Maximum standardizeduptake value (SUVmax:3.66±4.53) in positron emission tomography-computed tomography of mutant LC wassignificantly lower than that (8.26±6.11) of wild-type LC (p<0.0001). Concerning type of tumor shadow, thepercentage of mutant LC was 85.7% (6/7) in lung cancers with pure ground glass opacity (GGO), 65.3%(32/49)in lung cancers with mixed GGO and 21.7%(20/92) in lung cancers with solid shadow (p<0.0001). For the resultsof discriminant analysis, type of tumor shadow (p=0.00036) was most significantly associated with mutant EGFR.Tumor histology (p=0.0028), smoking status (p=0.0051) and maximum diameter of tumor shadow in chest CT(p=0.047) were also significantly associated with mutant EGFR. The accuracy for evaluating EGFR mutationstatus by discriminant analysis was 77.0% (114/148). Conclusions: Mutant EGFR is significantly associatedwith lung cancer with pure or mixed GGO, adenocarcinoma, never-smoker, smaller tumor diameter in chestCT. Preoperatively, EGFR mutation status can be identified correctly in about 77 % of lung cancers.     

A phase II pharmacodynamic study of erlotinib in patients with advanced non-small cell lung cancer previously treated with platinum-based chemotherapy

PURPOSE: To examine potential markers of clinical benefit and the effects of erlotinib on the epidermal growth factor receptor (EGFR) signaling pathway in advanced non-small cell lung cancer patients refractory to platinum-based chemotherapy. EXPERIMENTAL DESIGN: Patients were given erlotinib (150 mg/d). Tumor biopsies were done immediately before treatment and in a subgroup of patients after 6 weeks' treatment. RESULTS: Of 73 evaluable patients, 7 (10%) had partial response and 28 (38%) had stable disease. In 53 patients with baseline tumor samples, no relationship was observed between pretreatment levels of EGFR, phosphorylated (p)-EGFR, p-AKT, p-mitogen-activated protein kinase (MAPK), or p27 and clinical benefit (i.e., response, or stable disease >/=12 weeks). Tumors from 15 of 57 patients had high EGFR gene copy number, assessed using fluorescence in situ hybridization (FISH positive), 10 of whom had clinical benefit, compared with 5 of 42 FISH-negative patients. FISH-positive patients had longer median progression-free [137 versus 43 days, P = 0.002; hazard ratio (HR), 0.37] and overall (226 versus 106 days, P = 0.267; HR, 0.70) survival than FISH-negative patients. In paired biopsy samples from 14 patients, p-EGFR (P = 0.002), p-MAPK (P = 0.001), and Ki-67 (P = 0.025) levels were significantly reduced after 6 weeks' treatment. Apoptosis was significantly increased in patients with clinical benefit (P = 0.029), and may be a marker of clinical benefit. CONCLUSION: In this study, EGFR FISH-positive status was associated with improved outcome after erlotinib therapy. Erlotinib led to reduced levels of p-EGFR, p-MAPK, and Ki-67, and stimulated apoptosis in tumor samples from patients with clinical benefit.     

非小细胞肺癌组织中表皮生长因子受体基因突变与拷贝数之间的相关性以及与患者临床病理特征之间的关系

目的 研究中国非小细胞肺癌(NSCLC)患者组织中表皮生长因子受体(EGFR)基因状态,分析EGFR基因突变与拷贝数之间的相关性以及与NSCLC患者临床病理特征之间的关系。方法 应用实时荧光定量PCR法检测118例NSCLC组织中EGFR基因的突变状态,同时采用荧光原位杂交(FISH)技术检测EGFR基因拷贝数。分析EGFR基因突变与拷贝数之间的相关性,探讨EGFR基因状态与NSCLC患者临床病理特征之间的关系。结果118例NSCLC组织中,EGFR基因的突变率为41.5％,其中腺癌为50.0％,鳞癌为5.0％。EGFR基因FISH阳性率为70.3％,其中29例为基因扩增,54例为高度多体性;腺癌FISH阳性率为78.1％,鳞癌为35.0％。EGFR基因突变与EGFR基因高拷贝数主要存在于腺癌、晚期、女性和不吸烟的患者,EGFR基因突变与EGFR基因拷贝数之间有显著的相关性。结论在中国NSCLC患者中,腺癌、晚期、女性、不吸烟的患者EGFR基因突变率和FISH阳性率较高,且二者具有显著的相关性;联合检测EGFR基因拷贝数和基因突变可能更有利于NSCLC靶向药物的筛选。     

联合检测EGFR、CEA对恶性胸腔积液的诊断价值

目的:评价联合检测胸腔积液患者的胸水细胞块表皮生长因子受体（epidermal growth factor receptor,EGFR）基因拷贝数,以及胸水、血清CEA水平对良恶性胸腔积液鉴别诊断的价值。方法:应用荧光原位杂交技术（Fish法）检测恶性胸腔积液（n=35）、良性胸腔积液（n=30)组患者胸水细胞块EGFR基因拷贝数水平。采用电化学发光全自动生化分析仪检测胸水及血清中CEA水平,根据受试者工作特性曲线（ROC）选取最佳灵敏性和特异性的点作为临界值,评价CEA及联合检测EGFR基因拷贝数对良恶性胸腔积液的诊断价值。结果:35例恶性胸腔积液完成Fish检测。恶性胸腔积液中15例阴性,20例阳性,阳性率为57.1％。其中13例为EGFR基因高度多体性,7例为EGFR基因扩增。肺腺癌16例中,EGFR基因高度多体性、扩增14例,肺腺癌扩增率为87.5％;肺鳞癌14例中,EGFR基因簇状扩增3例(21.4％),点状扩增3例(21.4％),无扩增8例(57.1％),肺鳞癌扩增率为42.9％。腺癌Fish阳性率(87.5％)高于鳞癌(42.9％）,P<0．01。30例良性胸腔积液中有1例脓胸患者EGFR Fish检测阳性,阳性率为3.3%,余检测结果均阴性。恶性胸腔积液患者胸水及血清CEA分别为（220.9±71.65）ng/ml、(18.11±11.38）ng/ml,显著高于良性胸腔积液组（2.31±1.29)ng/ml、(1.67±1.06）ng/ml,差异有统计学意义（P<0.01）。其中,恶性胸腔积液胸水CEA明显高于血清CEA,而良性胸腔积液组中,胸水CEA与血清CEA无明显差异。肺腺癌所致胸水及血清CEA分别为（441.02±102.65)ng/ml、(32.87±28.66）ng/ml,鳞癌所致胸水及血清CEA分别为（28.75±21.39）ng/ml、（5.99±5.32）ng/ml,腺癌显著高于鳞癌,差异有统计学意义（P<0.01）。比较胸水EGFR、CEA对良恶性胸腔积液诊断的效能,两者之间无明显差异（P=0.453＞0.05）。Spearman相关性分析胸水EGFR同CEA之间存在显著正相关。结论:EGFR在恶性胸腔积液的形成中起重要作用,通过Fish技术检测胸水细胞块EGFR基因拷贝数可行,其敏感性为57.1%。对肺腺癌导致恶性胸腔积液的诊断敏感性为87.5%。CEA（临界值5.0ng/ml）在恶性胸腔积液及血清中显著高于良性,其中胸水CEA检测诊断敏感性为65.7%,而在腺癌中为87.5%,其在胸水及血清中的比值＞1.5有助于恶性胸腔积液的诊断。EGFR基因突变阳性与肿瘤标记物CEA阳性表达呈正相关,尤见于肺腺癌患者,两者联合检测可提高诊断性试验的准确性。     

EGFR FISH assay predicts for response to cetuximab in chemotherapy refractory colorectal cancer patients.

BACKGROUND: Standardized conditions to distinguish subpopulations of colorectal cancer (CRC) patients more and less sensitive to cetuximab therapy remain undefined. Materials and methods: We retrospectively analyzed epidermal growth factor receptor (EGFR) copy number by fluorescence in situ hybridization (FISH) in paraffin-embedded tumor blocks from 85 chemorefractory CRC patients treated with cetuximab. Results were analyzed according to different score systems previously reported in colorectal and lung cancers. The primary end point of the study was identification of the EGFR FISH score that best associates with response rate (RR). RESULTS: Using receiver operating characteristic (ROC) analysis, the cut-off that best discriminated responders versus nonresponders to cetuximab was a mean of 2.92 EGFR gene copies per cell. This model showed sensitivity of 58.6% [95% confidence interval (CI) = 47.1-70.1) and specificity of 93.3% (95% CI = 80.6-100). EGFR FISH-positive patients (N = 43, 50.6%) had significantly higher RR (P = 0.0001) and significantly longer time to disease progression (P = 0.02) than EGFR FISH negative (N = 42, 49.4%). Other scoring systems resulted less accurate in discriminating patients with the highest likelihood of response to cetuximab therapy. CONCLUSIONS: CRC patients with high EGFR gene copy number have an increased likelihood to respond to cetuximab therapy. Prospective clinical trials with a careful standardization of assay conditions and pattern interpretation are urgently needed.     

Comprehensive epidermal growth factor receptor gene analysis from cytological specimens of non-small-cell lung cancers

Epidermal growth factor receptor (EGFR) gene mutations and increased copy numbers are considered as predictors of response to EGFR tyrosine kinase inhibitors (EGFR-TKI) in non-small-cell lung cancer (NSCLC). Lung cancer diagnosis is often based on cytology alone. However, almost all published data on EGFR gene analyses were obtained from biopsies. This study tested the feasibility of EGFR gene analyses on cytological specimens. Eighty-four cytological specimens from NSCLCs were prospectively analysed for EGFR gene mutation in exons 18-21 and EGFR gene copy numbers were evaluated by fluorescence in situ hybridisation (FISH). A FISH-positive result was defined according to the criteria by Cappuzzo et al established for biopsies of NSCLCs. Fluorescence in situ hybridisation results of cytological specimens were compared to the FISH results on matching biopsies (n=33). Initial diagnosis of NSCLC was solely based on cytology in 37 out of 84 (44.0%) patients. Out of 80 NSCLCs, 6 (7.5%) showed EGFR gene mutations. Out of 67 cancers, 45 (67.2%) were FISH positive on cytological specimens. Comparison of FISH showed a FISH-positive result in 21 out of 33 (63.6%) cytological specimens but in only 8 out of 33 (24.2%) matched biopsies. Epidermal growth factor receptor gene analyses are well applicable to cytological specimens. The high FISH-positive rate of NSCLC on cytological specimens contrasts with the low rate on biopsies when previously suggested criteria are used. New criteria for a positive EGFR FISH status to predict response to therapy with EGFR-TKI need to be defined for cytological specimens.