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化检测乳腺癌组织HER-2基因扩增或蛋白表达情况的比较分析

目的 评估免疫组织化学(IHC)法检测乳腺癌组织中HER-2蛋白表达和荧光原位杂交(FISH)法检测其HER-2基因扩增在乳腺癌分子靶向治疗中的临床意义.方法 采用IHC法及FISH法分别检测47例乳腺癌组织中HER-2蛋白表达及HER-2基因扩增情况.结果 47例乳腺癌石蜡标本中,HER-2蛋白表达(+++)20例(42.55%),(++)10例(21.28%),(+)17例(36.17%);HER-2基因扩增22例(46.81%),无扩增25例(53.19%),其中17号染色体多体5例(10.64%).两种检测结果呈正相关(rs=0.415,P<0.05).结论 IHC法检测乳腺癌组织中HER-2蛋白表达町以作为是否应用赫赛汀的初筛;HER-2蛋白表达阳性的病例有必要进一步进行HER-2基凶扩增检测,来确定临床治疗是否应用赫赛汀.     

非小细胞肺癌EGFR荧光原位杂交与免疫组化检测的比较

目的探讨荧光原位杂交（FISH）技术和免疫组化（IHC）法检测石蜡标本非小细胞肺癌（NSCLC）EGFR基因扩增及蛋白表达水平的差异性。方法采用FISH和IHC分别检测27例NSCLC患者石蜡标本EGFR基因和蛋白表达,对2种方法的检测结果进行对比分析。结果 14例IHC法EGFR表达（3＋）的标本中有9例FISH显示阳性（64.29%）,其中5例为EGFR基因高多体性扩增（55.56%）,4例为EGFR基因扩增（44.44%）;6例IHC（2＋）的标本中仅1例为高多体性扩增（16.67%）;2例IHC（1＋）及5例IHC（-）标本均无EGFR基因扩增。结论 IHC法初筛（3＋）、（2＋）的标本与FISH检测的符合率较低,提示对IHC检测EGFR表达为（3＋）及（2＋）并选择靶向药物治疗的病例,应采用FISH法对EGFR基因表达作进一步检测。     

荧光原位杂交和免疫组织化学法检测乳腺浸润性导管癌中HER-2基因状态的研究

目的 比较荧光原位杂交（FISH）和免疫组织化学（IHC）两种方法检测乳腺浸润性导管癌人类表皮生长因子受体2（HER-2）基因扩增及与C-erbB-2蛋白表达结果的一致性。方法 分别采用FISH和IHC法检测346例乳腺浸润性导管癌组织HER-2基因扩增和C-erbB-2蛋白表达,并对两种方法的结果进行统计学分析。结果 346例乳腺浸润性导管癌中,FISH检测HER-2基因扩增145例（41.9％）,无扩增201例（58.1％）。IHC检测显示,C-erbB-2蛋白（-）7例,（+）30例,（++）227例,（+++）82例。按乳腺癌HER-2检测指南,（-）和（+）为阴性结果,（+++）为阳性结果,（++）为不确定病例。全组IHC检测C-erbB-2蛋白阳性表达率为23.7％（82／346）。IHC检测C-erbB-2蛋白（-）的7例患者经FISH检测无HER-2基因扩增,一致率为100.0％。IHC检测C-erbB-2蛋白（+）的30例经FISH检测25例无基因扩增,一致率为83.3％;227例（++）中有65例基因扩增,一致率为28.6％;82例（+++）中有基因扩增75例,一致率为91.5％。IHC和FISH检测HER-2状态的一致率为89.9％,具有高度一致性（Kappa值=0.768,P＜0.001）。HER-2基因扩增与年龄、肿瘤大小、组织学分级及淋巴结转移均无关（P＞0.05）。结论IHC和FISH法检测乳腺浸润性导管癌HER-2表达状态有高度一致性。IHC可以作为初步筛查乳腺浸润性导管癌HER-2基因状态的首选检测方法,对于IHC检测结果为（++）的标本建议采用FISH法进一步明确HER-2基因扩增状态。     

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。     

172例乳腺浸润性癌 HER -2免疫组化与 FISH 技术检验对比研究

目的：比较免疫组化（IHC）和荧光原位杂交（FISH）技术在检测乳腺浸润性癌患者中 HER -2蛋白表达和基因扩增的一致性。方法：分别用 IHC 和 FISH 技术对172例乳腺浸润性癌 HER -2进行蛋白表达和基因扩增检测,比较两者检测的结果和相关性。结果：172例浸润性乳腺癌标本行 IHC 检测结果30例为（1＋）,88例为（2＋）,42例为（3＋）,12例为（-）。172例乳腺癌标本进行 FISH 检测结果71例为阳性,101例为阴性。其中 FISH 结果阳性标本中 IHC 检测有2例（1＋）,30例（2＋）,39例（3＋）。IHC 检测 HER -2为（3＋）的病例 FISH 检测中阳性符合率92.9%（39/42）,且检测 HER -2（-）的病例 FISH 检测均为阴性, IHC 检测 HER -2（2＋）的88例患者中有58例经 FISH 检测证实 HER -2呈阴性,30例呈阳性。FISH 检测共发现47例17号染色体多体,其发生率为27.3%。用 IHC 检测 HER -2发现有25例标本存在肿瘤之间的不同表达,而在这25例标本的 FISH 检测结果中有11例存在 HER -2基因瘤内遗传异质性。结论：HER -2蛋白表达和基因扩增 IHC 和 FISH 检测在免疫组化强阳性的标本中具有较好的一致性,且免疫组化染色强度与HER -2基因扩增呈正相关。理解和判断 HER -2基因遗传异质性对肿瘤药物应用及 HER -2基因检测方法具有指导意义。     

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。     

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。     

荧光原位杂交法检测乳腺癌HER-2表达与腋淋巴结转移相关性的研究

目的:探讨用荧光原位杂交法(FISH)检测乳腺癌HER-2基因的表达与腋淋巴结转移的相关性.方法:采用免疫组化方法检测乳腺癌组织中HER-2的表达,再用FISH进一步检测阳性病例中HER-2基因的扩增情况,比较两者结果的差异,并且分析HER-2的表达与腋淋巴结转移的相关性.结果:免疫组化(IHC)法检测HER-2阳性42例,FISH检测阳性9例,FISH法检测HER-2基因与HER-2蛋白过表达之间相关(P＜0.05),并且IHC和FISH两种方法检测HER-2表达均与腋淋巴结转移有相关性(P＜0.05).结论:FISH可稳定检测HER-2基因的扩增,其表达与腋淋巴结转移相关.     

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.     

HER-2/neu（2+）腔面型乳腺癌的HER-2基因扩增与TopⅡα蛋白及临床特征的关联性分析

目的：分析HER-2基因扩增与腔面型HER-2/neu（2+）乳腺浸润性导管癌的临床病理特征及TopⅡα蛋白的关系。方法运用FISH检测手段对68例雌激素受体（ER）/孕激素受体（PR）阳性的腔面型HER-2/neu（2+）乳腺浸润性导管癌标本进行检测;应用免疫组化法（IHC）对68例标本进行TopⅡα蛋白表达检测。结果 FISH检测结果显示,68例标本中,HER-2基因扩增为13例（19.1%）;HER-2基因扩增在不同年龄、术后病理分期、淋巴结转移数目、Ki-67表达的患者之间差异无统计学意义;HER-2基因扩增病例中,TopⅡαⅠ级6例（46.2%）,Ⅱ级6例（46.2%）,Ⅲ级1例（7.7%）,Ⅳ级0例。TopⅡα蛋白表达情况在有无HER-2基因扩增的乳腺癌间差异无统计学意义（Z=1.353,P=0.176）。结论在HER-2/neu（2+）腔面型乳腺癌中,HER-2基因扩增与年龄、术后病理分期、淋巴结转移数目、Ki-67表达和TopⅡα蛋白表达无关。     

Caveolin-1在乳腺浸润性导管癌间质内癌相关成纤维细胞中的表达水平与乳腺癌分子亚型的相关性

目的 检测Caveolin-1（Cav-1）在乳腺浸润性导管癌间质内癌相关成纤维细胞（CAFs）中的表达,分析Cav-1与乳腺癌分子亚型、HER-2基因状态的相关性及其与预后的关系。方法应用免疫组织化学法检测168例乳腺癌组织Cav-1在CAFs中的表达情况。用荧光原位杂交（FISH）方法检测乳腺癌组织中HER-2基因的扩增状态。结果 Cav-1在HER-2过表达型和Luminal B型的阳性表达率均为83.3％,显著高于Luminal A型（58.1％）和Basal like型（35.3％）,差异有统计学意义（P＜0.05）。乳腺癌HER 2基因扩增占39.3％（66／168）,与HER-2蛋白表达呈正相关（r=0.625,P＜0.05）。HER-2基因扩增组的Cav-1阳性表达率为83.3％,显著高于未扩增组的55.9％（P＜0.05）。Cav-1表达水平与HER-2蛋白（r=0.253）及基因（r=0.305）表达均呈正相关（P＜005）,与预后亦明显相关（P=0.041）。结论 乳腺癌间质内Cav-1表达与分子分型相关,HER 2蛋白表达水平增高及基因扩增与间质Cav-1表达水平增高呈正相关（P＜0.05）,乳腺癌间质CAFs中Cav-1高表达提示患者预后较好。     

Reproducible Immunohistochemical Criteria Based on Multiple Raters’ Judgments for Expression of Thymidine Phosphorylase in Breast Cancer Tissue

The role of HER-2/ neu in male breast cancer is not well defined. The purpose of the current study was to measure the frequency of HER-2/ neu expression in primary male breast cancer, to demonstrate HER-2/ neu gene amplification in cases found to be positive for protein overexpression, and to correlate HER-2/ neu positivity with clinicopathological variables. Formalin-fixed, paraffin-embedded archival material from 99 primary male breast carcinomas was evaluated by immunohistochemistry (IHC) using the HercepTest (DAKO Corp., Hamburg, Germany). Scoring was performed according to established guidelines. All cases demonstrating HER-2/ neu staining by IHC (1+/2+ and 3+) were analyzed for HER-2/ neu gene amplification by fluorescence in situ hybridization (FISH) utilizing the PathVysion assay (Vysis Corp., Downers Grove, Illinois) to assess HER-2/ neu amplification status. The immunohistochemical staining of the HER-2/ neu protein revealed HER-2/ neu positivity in 15/99 (15.1%) cases, eight tumors showed 2+ and 7 tumors 3+ staining. HER-2/ neu gene amplification was observed in 11/99 cases (11,1%), and all of the 3+ and 4/8 from the 2+ cases were amplified. HER-2/ neu gene amplification/protein overexpression did not correlate with tumor state, histological grade or estrogen/progesterone receptor status nor the axillary lymph node status. This is the first comprehensive study of HER-2/ neu gene amplification by FISH analysis in primary male breast cancer. Compared to female primary breast cancer the percentage of HER-2/ neu positivity in our study was lower. Our data provide first evidence for HER-2/ neu gene amplification in male breast cancer. Further studies should be addressed on the potential application of the monoclonal rhuMAB HER-2/ neu antibody for treatment of HER-2/ neu positive male breast cancer.     

Detection of HER-2/neu gene amplification in breast carcinomas using quantitative real-time PCR — A comparison with immunohistochemical and FISH results

The aim of our study was to evaluate the value of quantitative real-time-PCR (qPCR) in the determination of HER-2/neu amplification status of human breast carcinomas by comparing qPCR, FISH and immunohistochemistry results from the same samples. A total of 210 breast carcinomas were examined. Ready-to-use CB11 antibody was applied to detect HER-2/neu oncoprotein expression. In 76 out of 210 cases FISH was performed, and 162 cases were investigated with qPCR. Seventy-five tumors were 2+ or 3+ positive with immunohistochemistry, while 135 samples were either completely negative or 1+. In 45 cases results from all three methods were available. Out of these, in twenty negative and sixteen positive cases both FISH and qPCR led to similar results. The mean qPCR amplification ratio in the concordant positive cases was 5.424 while in the qPCR+/FISH- group the mean ratio was 2.765. Out of 121 samples with scores of 0 or 1+ immunohistochemical result, analyzed also with qPCR, 26 showed HER-2/neu gene amplification. In these cases the mean amplification ratio was 2.53. Comparison of FISH and qPCR together with immunohistochemistry shows that qPCR is more sensitive to detect HER-2/neu gene amplification in tumors scored as 2+ with immunohistochemistry, but the diagnostic cut-off ratio should be defined above 2.7 to avoid high number of false positive cases. Amongst the immunohistochemistry score 2+ cases, 10 of 18 showed gene amplification by qPCR while 10 of 26 by FISH. In conclusion, a well calibrated HER-2/neu qPCR assay may serve as useful alternative to FISH in breast cancer patients.     

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.     

乳腺癌组织与转移淋巴结HER-2基因表达及检测方法的比较

目的 探讨乳腺癌组织与转移淋巴结中人表皮生长因子受体（human epidermal growth factor receptor-2, HER-2）基因表达的情况,对比荧光原位杂交法（Fluorescence in situ hybridization, FISH) 与免疫组织化学法（Immunohistochemistry,IHC）两种检测方法。方法 选取187例乳腺癌组织和76例转移淋巴结石蜡包埋标本,采用FISH和IHC方法检测HER-2基因表达,分析HER-2过表达与各临床病理因素的关系以及两种检测方法的差异,比较原发灶与转移灶HER-2表达的状况。结果 187例乳腺癌组织中,FISH检测显示HER-2阳性43例,阴性144例。IHC评分+++者22例、评分++者40例、评分-/+者125例。IHC评分+++者中,FISH阳性率为90.9%（20/22）;评分++者中,FISH阳性率为52.5%（21/40）;评分0/+者中,FISH阳性率为1.6%（2/125）。两种方法相比差异有统计学意义（P<0.001）。临床病理因素分析示HER-2基因扩增与淋巴结转移呈正相关 (P<0.05)。乳腺癌原发灶与其配对转移淋巴结相比,HER-2表达的不一致性为11.8%(9/76)。结论 FISH较之IHC检测HER-2表达准确性更高,尤适用于对IHC ++的患者判断HER-2的表达。乳腺癌原发灶与转移灶之间HER-2的表达存在一定的不一致性。     

HER-2 gene amplification by fluorescence in situ hybridization (FISH) compared with immunohistochemistry (IHC) in breast cancer: a study of 528 equivocal cases

The concordance rate between fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC) on evaluating HER-2/neu status is still controversial, especially for the IHC 2+/equivocal cases. In this study, we investigated the amplification of HER-2/neu gene by FISH in IHC (2+)/equivocal specimens to clarify the correlation between HER-2/neu and clinicopathologic features of breast cancers. HER-2/neu amplification was determined by FISH on 528 specimens of breast carcinomas with equivocal expression of HER-2/neu protein by IHC detection. 65.5 % of IHC 2+ patients were negative for HER-2/neu amplification, 29.0 % were positive and the remaining was equivocal. A statistically significant inverse association was found between hormone receptor expression and HER-2/neu amplification (P < 0.05). Furthermore, polysomy of CEP17 was detected in 60 % of breast carcinomas. The results highlight the necessity of FISH test for further categorization when breast carcinoma cases are scored 2+ by IHC.     

Different expression of HER2/neu oncogene in breast carcinoma and in liver metastasis. Description of a case

We report the clinical, morphological and molecular findings regarding a 37-year-old woman with breast cancer metastatic to the liver and describe the different expression of a tumor marker in the primary and secondary lesions and the singular responsiveness to treatment. The patient suffered from a carcinoma of the left breast with metastasis to the liver. High HER-2 protein expression assessed by immunohistochemistry and HER-2/neu amplification determined by FISH were present in the primary tumor, while the liver metastasis showed a lower value of HER-2 protein (2+) and absence of HER-2/neu amplification. The patient was treated with chemotherapy (epirubicin and paclitaxel) followed by trastuzumab and docetaxel. After 5 months, at the completion of chemo-immunotherapy, liver ultrasonography showed a further hepatic response. A second biopsy was performed on the residual liver nodule: immunohistochemistry revealed negative (1+) HER-2 expression and FISH confirmed that the HER-2/neu gene was not amplified. The different amplification of HER-2 and expression of its protein in primary and metastatic carcinoma could be important for planning adequate treatment.