荧光原位杂交和免疫组织化学方法检测乳腺癌组织中人类表皮生长因子受体2基因状态差异及其特征分析

目的比较荧光原位杂交(fluorescence in situ hybridization,FISH)和免疫组织化学(immunohistochemistry,IHC)检测乳腺癌组织中人类表皮生长因子受体2(human epidermal growth factor receptor 2,HER-2)基因状态差异,分析其相关特征。方法回顾性收集51例中山大学附属第三医院2007年10月至2008年9月间乳腺浸润性导管癌组织标本和相关信息,分别采用IHC和FISH法检测HER-2蛋白表达情况和基因扩增状态,比较两种方法的结果差异,用Fisher精确概率检验分析这种差异的相关因素。结果在51例标本中,FISH检测乳腺癌组织中HER-2基因阴性32例(62.75%,32/51),阳性19例(37.25%,19/51);IHC检测乳腺癌组织中HER-2为(-)和(+)者的FISH检测均为阴性(21例);12例IHC检测HER-2为(++),其中3例FISH检测为阳性,其余9例FISH检测为阴性;18例IHC检测HER-2为(+++)者仍有2例FISH检测结果阴性。月经状态和雌激素受体(ER)表达与IHC检测HER-2阳性病例的FISH阳性结果具有显著联系(P值分别为0.023和0.007),即在IHC检测HER-2阳性[(++)和(+++)]的病例中,绝经前女性较绝经期女性以及ER阴性者较阳性者更可能为FISHHER-2阳性(有HER-2基因扩增)。结论本研究的结果有助于提高IHC判断HER-2基因扩增的准确性。     

乳腺浸润性导管癌中p53和表皮生长因子受体的表达及其意义

目的探讨p53和表皮生长因子受体(Epidermal growth factor receptor,EGFR)在乳腺浸润性导管癌中的表达及其临床病理意义。方法采用免疫组化SP法检测30例正常乳腺组织、160例浸润性乳腺癌组织中p53和EGFR的表达。结果乳腺浸润性导管癌中p53、EGFR的阳性表达与正常乳腺组织间差异有显著统计学意义(P<0.01)。p53、EGFR的阳性表达与浸润性乳腺导管癌的病理分级、临床分期及淋巴结转移有关(P<0.05),而与患者的年龄、肿块的大小无关。结论 p53和EGFR在乳腺浸润性导管癌的发生、发展过程中发挥了一定的作用,两者联合检测有助于预测患者的预后,并为临床进行靶向治疗提供依据。     

免疫组织化学法与荧光原位杂交技术检测861例乳腺癌HER-2表达的一致性分析

目的:对比免疫组织化学法(IHC)与荧光原位杂交技术(FISH)检测861例乳腺癌人类表皮生长因子受体2(HER-2)蛋白表达和基因状态的一致性。方法:参照《乳腺癌HER-2检测指南(2019版)》判读标准对乳腺癌石蜡组织采用IHC法及FISH法分别检测HER-2蛋白表达和HER-2基因状态,进行比较分析。结果:861例乳腺癌患者中,IHC HER-2（0/1+)和FISH(-)的一致率为97.1%（134/138）;IHC HER-2(2+)和FISH(+)的一致率为28.7%（188/655）;IHC HER-2(3+)和FISH(+)的一致率为98.5%（67/68）。两种检测方法的检测结果存在一致性,差异具有统计学意义(Kappa=0.137,P＜0.000 1）。结论:两种方法相结合,以便精准评价HER-2 检测结果指导靶向治疗。     

乳腺癌中HER-2基因及蛋白的检测和临床应用

目的:在蛋白及基因水平检测乳腺癌中HER-2的表达,比较免疫组化(IHC)及荧光原位杂交(FISH)法检测结果,分析其临床应用.方法: 应用免疫组化及荧光原位杂交法测定HER-2蛋白表达及基因扩增.结果: IHC检测HER-2蛋白,77例乳腺浸润性导管癌中 ,[(-)-(+)]占 35.06%,(++)占37.66%,(+++)占27.27%;FISH检测HER-2基因,扩增阳性率为51.95%.IHC强阳性组(+++),FISH阳性率为90.48%,IHC弱阳性组(++), FISH阳性率为 68.97%,IHC阴性组[(-)-(+)], FISH阳性率为 3.07%,P<0.01,具有显著统计学意义.以HER-2基因扩增为阳性标准,IHC 结果(+++)阳性预测值为90.48%,灵敏度为95.00%,特异度为92.86%;(++)阳性预测值为68.97%,灵敏度为95.24%,特异度为74.29%;[(-)-(+)]阴性预测值为96.30%.结论:HER-2蛋白过表达主要是基因扩增所致,IHC强阳性 (+++)及IHC阴性[(-)-(+)] 可直接指导临床用药,IHC弱阳性(++)需行FISH确定有无基因扩增.     

荧光原位杂交与免疫组化法检测乳腺癌HER-2表达的临床意义

[目的]探讨乳腺癌中荧光原位杂交(FISH)检测HER-2基因的扩增和免疫组织化学(IHC)检测HER-2蛋白表达在临床诊断及分子靶向治疗中的应用。[方法]采用FISH与IHC检测50例乳腺癌组织中HER-2基因的扩增与HER-2蛋白表达。[结果]50例浸润性乳腺癌中,FISH检测HER-2基因扩增阳性15例(30%),IHC检测HER-2蛋白表达(-～+)40例,HER-2蛋白表达(++)的2例,HER-2蛋白表达(+++)的8例。[结论]FISH检测HER-2基因的扩增与IHC检测HER-2蛋白表达(++～+++)有较高的一致性。对于IHC检测HER-2(-～++)表达时,必须进一步FISH检测。     

荧光原位杂交与免疫组织化学方法对乳腺癌Her-2检测的比较

[目的]探讨荧光原位杂交(FISH)与免疫组织化学方法(IHC)检测乳腺癌组织中人类表皮生长因子受体2(Her-2)表达结果的差异.[方法]采用FISH和IHC法分别检测508例乳腺癌标本中Her-2基因扩增和Her-2蛋白表达状况.[结果]在IHC法检测Her-2蛋白表达0/+的104例(20.47％)标本中,FISH检出13例(2.56％)Her-2基因扩增,89例(17.52％)为非扩增,2例(0.39％)为可疑;Her-2蛋白表达4++的245例(48.23％)标本中,FISH检出121例(23.82％)Her-2基因扩增,120例(23.62％)为非扩增,4例(0.79％)为可疑;IHC在Her-2蛋白表达+++的159例(31.30％)标本中,FISH检出137例(26.97％) Her-2基因扩增,22例(4.33％)为非扩增.[结论]对于IHC法初筛Her-2蛋白为++～+++的患者必须进行FISH检测,以明确Her-2基因的状态,从而指导临床医师诊断治疗.     

色素原位杂交法和免疫组织化学法检测乳腺癌HER-2基因扩增及蛋白表达

目的:对照色素原位杂交法(chromogenic in situ hybridization,CISH)和免疫组织化学法(immuno-histochemistry,IHC)检测乳腺癌组织中人表皮生长因子受体(human epidermal growth factor receptor-2,HER-2)基因扩增及其蛋白表达的状况.方法:采用CISH技术检测145例乳腺癌组织中HER-2的基因扩增情况,其中14例标本经过荧光原位杂交(fluorescence in si-tu hybridization,FISH)检测,随后分别用FISH和IHC方法检测的HER-2基因扩增及蛋白表达状况与之进行回顾性对照,并按照病理分级、淋巴结状态、绝经与否和雌激素受体(estrogen receptor,ER)/孕激素受体(progesterone receptor,PR)表达情况进行分层,分析HER-2表达与乳腺癌各高危因素之间的关系.结果:CISH检测发现,HER-2无扩增71例(50.0%),低扩增11例(7.6%),高扩增63例(43.4%).14例FISH与CISH检测结果比较,符合率为100.0%.145例IHC与CISH检测结果的符合率为84.8%(P<0.05).在IHC检测积分为0/ 以及 的病例中,HER-2基因扩增与蛋白表达情况基本一致,其符合率均在90%以上;而在IHC检测积分为 的标本中,HER-2基因扩增率仅为61.1%.CISH及IHC检测均显示ER/PR表达情况与HER-2阳性呈负相关,ER和PR均为阴性患者的HER-2基因扩增率和蛋白表达率明显高于ER/PR阳性的患者(CISH:68.3%vs 38.8%,P<0.01;IHC:71.7%vs 48.2%,P<0.01).HER-2状态与乳腺癌病理分级、腋淋巴结转移以及绝经与否无关(P>0.05).结论:CISH技术检测HER-2操作简便,准确性高,可以代替FISH技术,对IHC评分为 的病例应进一步确认HER-2状态.HER-2除了与ER/PR表达相关外,与其他乳腺癌高危因素无关,可作为独立指标进行检测.     

环氧化酶2和血管内皮生长因子在乳腺浸润性导管癌中的表达及其临床意义

目的 探讨环氧化酶2（COX-2）、血管内皮生长因子（VEGF）mRNA及其蛋白在乳腺浸润性导管癌中的表达及其与临床病理特征的关系.方法 采用原位杂交和免疫组织化学技术检测70例乳腺浸润性导管癌和30例乳腺纤维腺瘤中COX-2、VEGF mRNA及蛋白的表达.计数资料的分析运用χ2检验,相关性分析采用Spearman检验.结果 乳腺浸润性导管癌COX-2 mRNA阳性率明显高于乳腺纤维腺瘤[74.2%（52/70）比30.0%（9/30）,χ2=17.31,P=0.00],COX-2 蛋白阳性率也明显高于乳腺纤维腺瘤[72.8%（51/70）比23.3% （7/30）,χ2=21.14,P=0.00].乳腺浸润性导管癌COX-2 mRNA及蛋白的表达均与淋巴结转移有关（χ2=7.54,P=0.00;χ2=6.36,P=0.01）,与年龄、肿瘤大小和组织学分级无关（P〉0.05）.乳腺浸润性导管癌VEGF mRNA阳性率明显高于乳腺纤维腺瘤[71.4%（50/70）比33.3% （10/30）,χ2 =12.70,P=0.00],其VEGF蛋白阳性率也明显高于乳腺纤维腺瘤[65.7%（46/70）比26.7%（8/30）,χ2 =12.89,P=0.00].乳腺浸润性导管癌VEGF mRNA表达与淋巴结转移有关（χ2=8.33,P=0.00）,与年龄、肿瘤大小及组织学分级无关（P〉0.05）;而VEGF蛋白表达与组织学分级、淋巴结转移有关（P〈0.05）,与年龄及肿瘤大小无关（P〉0.05）.在乳腺浸润性导管癌中,COX-2 mRNA表达与VEGF mRNA表达之间以及COX-2蛋白表达与VEGF蛋白表达之间均呈正相关关系（r=0.64,P=0.00;r=0.44,P=0.00）.结论 COX-2和VEGF的mRNA及蛋白在乳腺浸润性导管癌中表达均上调且与淋巴结转移有关,因此,COX-2、VEGF可作为判断乳腺浸润性导管癌预后的生物学标志物.     

乳腺浸润性导管癌组织表皮生长因子受体基因检测方法的比较及应用

目的分析免疫组织化学（IHC）、色素原位杂交（CISH）、荧光原位杂交（FISH）方法在乳腺浸润性导管癌组织表皮生长因子受体（Her-2）基因检测中的选择应用。方法分别采用IHC、CISH、FISH方法对100例乳腺浸润性导管癌组织的Her-2基因进行检测。结果IHC（-）、（＋）、（＋＋）、（＋＋＋）者与CISH结果符合率分为100％（6／6）、7．1％（1／14）、32％（16／50）、80％（24／30）;CISH和FISH符合率为100％（20／20）。结论IHC、CISH、FISH方法各有优劣。CISH是一准确、可靠且较易普及开展的Her-2基因检测方法。     

双色FISH分析浸润性乳腺癌HER-2基因扩增模式

[目的]应用荧光原位杂交技术(FISH)分析乳腺癌HER-2基因状态。[方法]采用双色FISH技术检测100例浸润性乳腺癌HER-2基因,分析FISH和免疫组织化学(IHC)检测结果,分析17号染色体多倍体比率及其与HER-2基因扩增的关系。[结果]100例浸润性乳腺癌标本中,HER-2蛋白IHC3+21例者中FISH检测均为阳性;IHC2+69例者中FISH检测为阳性的占72%,IHC1+10例者FISH检测为阳性的占20%。17号染色体多倍体的比率为51%。17号染色体二倍体与多倍体标本中,HER-2基因扩增的比率分别是76%和69%,差异无统计学意义(P=0.330)。[结论]乳腺癌IHC检测HER-2为1+~2+者需进一步行FISH检测确定HER-2基因状态;应用双色FISH方法能准确、全面评估HER-2基因状态。     

HER2 status in lung adenocarcinoma: A comparison of immunohistochemistry,fluorescence in situ hybridization (FISH), dual-ISH,and gene mutations

Objectives

While novel anti-human epidermal growth factor receptor 2 (HER2) agents have recently been developed, no definite criteria have been proposed as indications for the use of these agents in patients with lung cancer. Here, we tested HER2 alterations by using four methods and explored the concordance of these methods to improve our understanding of the accuracy of HER2 testing methods.

Materials and methods

We analyzed HER2 protein expression by immunohistochemistry (IHC) and HER2 amplification by fluorescence in situ hybridization (FISH) and dual in situ hybridization (DISH) by using a tissue microarray comprising lung adenocarcinoma specimens from 243 consecutive patients. The presence of mutations in the EGFR, KRAS, and HER2 genes were also determined.

Results

Positive IHC (score 3+) was observed in six cases (2.5%). Amplification of HER2 was observed in five cases (2.1%) by FISH and in nine cases (3.7%) by DISH. HER2 expression by IHC and gene amplification by FISH were significantly associated (P < 0.001). The overall concordance between FISH and DISH by amplification status was 96.7% (P < 0.001). One hundred nine tumors (49.9%) had EGFR mutations, 25 (11.2%) had KRAS mutations, and six (2.7%) had HER2 mutations. All of these mutations were mutually exclusive. Cases having HER2 mutations were positively correlated with cases having HER2 amplification (P < 0.001). Two of six cases with HER2 mutations showed amplifications by FISH and DISH tests.

Conclusion

HER2 protein overexpression, gene amplification, and gene mutations appeared to be uncommon in lung adenocarcinoma. Cases with HER2 mutations tended to show HER2 gene amplification. The results indicated that HER2 gene amplification and mutations should be tested to determine whether patients are eligible for administration of new anti-HER2 agents. In addition, DISH was better than FISH for detection of cases with HER2 amplification.     

The HER-2/neu Oncogene in Tumors of the Gastrointestinal Tract

The HER-2/neu oncogene is localized to chromosome 17q and shares significant homology with the epidermal growth factor receptor. As a result of its potential role in the selection of therapy, HER-2/neu testing has reached near-standard-ofpractice status in breast cancer. There is considerable interest in HER-2/neu as a prognostic factor and target of therapy in tumors of the gastrointestinal tract. In this review of HER-2/neu expression in esophageal squamous cell carcinoma and adenocarcinomas of the esophagus, stomach, and colon, a wide range of expression of HER-2/neu from 0 to 83% likely reflects both differences in methods and reagents, as well as study bias associated with patient selection (i.e., early versus advanced disease). For esophageal squamous cell carcinoma, little information exists as to the prognostic significance of HER-2/neu expression. In adenocarcinoma associated with Barrett's esophagus there is contradictory data. However, most of the information available indicates that this marker has significant prognostic value. In gastric adenocarcinoma, the wide expression range may truly reflect patient selection because HER-2/neu positivity appears linked to advanced rather than early disease with limited invasion. The majority of studies favor a significant prognostic value of HER-2/neu status for this tumor. Finally, in colorectal cancer HER-2/neu overexpression also appears to be a significant adverse outcome indicator as judged by the current published literature. In conclusion, given that either HER-2/neu protein overexpression or gene amplification is associated with approximately one-fourth of all gastrointestinal tract malignancies, strategies designed to employ the marker in therapy selection appear warranted. During the next several years it will not be surprising to see these tumors treated with antiHER-2/neu modalities such as Herceptin^TM, likely in combination with other agents initially for patients with advanced disease, and possibly for individuals with high-risk lesions in an adjuvant setting.     

HER-2/neu and p53 in osteosarcoma: an immunohistochemical and fluorescence in situ hybridization analysis

The overexpression of HER-2/neu and p53 has been associated with poor outcome in many neoplasms. Their role in patients with osteosarcoma is unclear. We studied the expression of HER-2/neu and p53 in 22 osteosarcoma samples (from 20 patients—2 had locally recurrent disease) biopsied at the University of Medicine and Dentistry of New Jersey (UMDNJ) from 1996-2000 using both immunohistochemical (IHC) and fluorescence in situ hybridization (FISH) analysis. Fourteen patients (14 samples) presented with Stage II and 6 patients (8 samples) presented with Stage III disease. Median follow-up is two years (range one year to five years). Four of 22 (18%) samples showed focal membranous or cytoplasmic positivity for HER-2/neu and six of 22 samples (27%) showed nuclear positivity for p53 by IHC analysis. In contrast, none of 22 tested samples showed gene amplification for HER-2/neu by FISH analysis. Seven of 13 HER-2/neu and p53 negative patients (54%) are currently disease free (between one year to five years). In this sample of patients, the HER-2/neu oncogene is not overexpressed or amplified in osteosarcoma; six of 22 samples (27%) showed overexpression of p53 by IHC analysis. By FISH, none of the samples demonstrated deletion of p53. Neither HER2/neu nor p53 expression was important for the biology of osteosarcoma in this population.     

乳腺癌患者HER-2在血清和细胞学中表达的相关性分析

目的:探讨乳腺癌患者HER-2在血清和细胞学中表达的相关性.方法:纳入2015年1～10月住院治疗乳腺癌患者188例,进行血清和细胞学HER-2检查,血清HER-2应用酶免方法(ELISA检测,细胞学HER-2应用免疫组化和荧光原位杂交(fluorescence in situ hybridization,FISH)方法检测,应用统计学检验方法对检查结果进行分析.结果:188例乳腺癌患者血清中HER-2检查阳性率为35.1％ (66/188),四分位数(P25、P5o、P75)分别为6.9ng/ml、11.2ng/ml、15.4ng/ml;细胞学HER-2检查的阳性率为28.2％ (53/188),应用计量资料相关性的Spearman分析,t=0.1432(P ＞0.05),认为血清HER-2和细胞学HER-2没有相关性.结论:乳腺癌患者的血清HER-2和细胞学HER-2检查是不同的分析方法,没有相关性,临床应用可以参考,不能等同.     

乳腺导管原位癌与浸润性导管癌激素受体及HER-2表达的比较

目的研究乳腺导管原位癌（DCIS）与浸润性导管癌（IDC）患者的激素受体和HER-2表达及分子分型的差异。方法收集2006年3月至2012年9月本院治疗的52例DCIS和224例IDC患者的手术标本,应用免疫组化方法检测其ER、PR、HER-2的表达,并用×2检验综合分析其分子分型构成等特点。结果IDC组ER阳性表达率明显高于DCIS组[67．86％（152／224）比48．08％（25／52）,）（X^2=7．18,P=0．01],IDC组HER-2阳性表达率明显低于DCIS组[54．02％（121／224）比73．08％（38／52）,）（X^2=6．28,P=0．01],而两组的PR表达差异无统计学意义（X^2=1．39,P=0．24）。DCIS组以HER-2阳性型为主要亚型,占44．23％（23／52）,而IDC组以luminalA型为主要亚型,占43．75％（98／224）,两组患者的分子分型构成比差异有统计学意义（X^2=13．11,P=0．00）。结论检测乳腺导管原位癌和浸润性导管癌患者的ER、HER-2表达及其分子分型,可指导临床诊疗及制定个体化综合治疗方案。     

HER-2/neu amplification detected by fluorescence in situ hybridization in fine needle aspirates from primary breast cancer.

BACKGROUND: The HER-2/neu gene is amplified in 20-30% of human breast cancers and has been shown to have prognostic and predictive value for treatment with chemotherapy, hormone therapy and antibodies against the HER-2/neu domain (trastuzumab). The aim of our study was to evaluate the reliability of HER-2/neu determination by fluorescence in situ hybridization (FISH) on fine-needle aspirates (FNAs) from primary breast cancer patients by comparison with the results obtained by FISH and immunohistochemistry (IHC) on the corresponding histological sections. MATERIALS AND METHODS: HER-2/neu amplification was determined by FISH on 66 breast cancer FNAs. Twenty-three and 36 corresponding formalin-fixed, paraffin-embedded sections were assayed by FISH and by IHC, respectively, in order to detect HER-2/neu amplification and HER-2/neu protein expression. RESULTS: Twenty-seven per cent (18/66) of breast cancer FNAs showed amplification of HER-2/neu by FISH. Paired results by FISH cytology and FISH histology were available in 22 cases. Concordance was 91% (20/22). Paired results by FISH cytology and IHC were available in 36 cases. Concordance was 92% (33/36). Eighteen of 66 breast cancer FNAs were also submitted to flow cytometric DNA analysis. None of the diploid cases showed HER-2/neu amplification by FISH. Six out of the eight aneuploid cases were amplified and two were polysomic. CONCLUSIONS: HER-2/neu gene amplification can be reliably estimated by FISH on breast cancer FNAs and a good correlation has been found between FISH and IHC results from the corresponding histological sections.     

HER-2 (c-erbB-2) test update: Present status and problems

HER-2 tests are routinely used for the identification of patients with metastatic breast cancer that is potentially responsive to trastuzumab (herceptin) therapy. Recently, convincing data have been published with regard to the efficacy of trastuzumab as a drug for neoadjuvant therapy or adjuvant therapy for operable primary breast cancer that overexpresses HER-2. It is also noteworthy that a the St. Gallen International Consensus Conference 2005, HER-2 protein overexpression or HER-2 gene amplification has been included as an indicator for higher risk of recurrence for both node-negative and node-positive breast cancers. To measure the HER-2 level, the worldwide consensus appears to be that immunohistochemistry (IHC) should be performed first and, if the results of IHC are uncertain, fluorescence in situ hybridization (FISH) should be performed later, although some investigators argue that FISH should be performed first. These tests should be performed in strict adherence to existing instructions. Quality control is of utmost importance when performing HER-2 tests, both internal and external, for routine diagnosis and in clinical protocol studies.     

微阵列技术用于检测乳腺癌人表皮生长因子受体2基因状态

目的探讨细胞核微阵列技术及组织微阵列技术用于检测乳腺癌组织中HER-2基因扩增和蛋白表达状态。方法将248例乳腺癌普通组织蜡块制成组织微阵列组,应用免疫组织化学法（IHC）和荧光原位杂交法（FISH）分别检测HER-2基因和蛋白表达。两种检测方法的-致性采用检验,并以FISH检测结果为金标准,绘制IHC检测乳腺癌HER-2表达的ROC曲线图。结果248例乳腺癌FISH与IHC检测结果-致性分析显示：Kappa=0．711,P=0．000,两种检测方法的-致性尚可。IHC检测乳腺癌HER-2ROC曲线下面积为0．888,P=0．000,约登指数为0．700,准确率为87．9％。IHC（＋＋）中4例为17号染色体单体型,占FISH阳性病例总数的5．26％（4／76）。IHC（＋＋＋）中5例为17号染色体多倍体型,FISH检测均为阴性,占FISH阴性总例数的2．9％（5／172）。IHC的检测可以较好地反映出FISH的结果。结论细胞核微阵列及组织微阵列技术用于检测乳腺癌HER．2基因状态有着节约实验成本、同一性好、结果可靠的优点。