HER2免疫组织化学结果不确定的乳腺癌原位mRNA表达特征

HER2 mRNA expression in breast cancers with equivocal results using in situ mRNA hybridization

目的了解HER2免疫组织化学结果不确定的乳腺癌荧光原位杂交（FISH）阳性和阴性两组患者的原位mRNA表达特征，探索mRNA原位杂交（RNAscope）用于乳腺癌组织HER2检测的潜在可行性。方法收集北京协和医院2010年6月至2013年6月问手术获得的69例乳腺浸润性导管癌组织HER2免疫组织化学结果不确定（IHC2＋）的石蜡标本，并制作组织芯片，分别采用FISH和RNAscope技术检测其HER2基因状态和原位mRNA表达，计数100个癌细胞，按mRNA表达量从低到高的5个等级（0、1、2、3、4分）判读mRNA结果。分析FISH阳性和阴性两组患者HER2mRNA的表达特征。结果在69例标本中，FISH阳性共23例，其中RNAscope16例（70％，16／23）4分、6例（26％，6／23）3分、1例（4％，1／23）2分。96％的HER2FISH阳性标本为原位mRNA高表达（RNAscope≥3分），RNAscope4分的标本全部为FISH阳性。在FISH阴性的46例标本中，RNAscope17例（37％，17／46）3分、25例（54％，25／46）2分、4例（9％，4／46）1分。结论根据原位mRNA表达结果，可将IHC2＋乳腺癌病例进一步分类，其中以RNAscope4分为mRNA高表达的判读标准可能成为确认IHC2＋结果的检测技术。HER2原位mRNA检测可作为提示乳腺癌HER2基因扩增和蛋白质表达的候选检测手段，具有潜在的临床应用价值。

Objective To investigate in situ mRNA expression of HER2 oncogene in breast cancers with equivocal immunohistochemical results, and to explore the potential feasibility of RNAscope technique in evaluating HER2 status in breast cancers. Methods Sixty-nine FFPE samples of invasive ductal breast cancer with equivocal HER2 immunohistochemistry results （ IHC 2 ＋ ） were collected from surgical excisions from Peking Union Medical College Hospital between June 2010 and June 2013. HER2 status and in situ mRNA expression were tested by fluorescence in situ hybridization （FISH） and RNAscope respectively using tissue microarray constructed from tumor paraffin blocks. The results of HER2 mRNA expression were scored 0 to 4 （ from low to high levels） according to mRNA expression in 100 cancer cells. HER2 mRNA expression was evaluated in two groups of patients, with positive and negative FISH results. Results Twenty-three of the 69 samples were FISH positive, including 16 samples that were scored 4 by RNAscope （70%, 16/23）, 6 samples were scored 3 （26% ,6/23） and one sample was scored 2 （4%, 1/23）. High in situ mRNA expression （ score 4 or 3） were observed in 96% of HER2 FISH positive samples. All of samples that were scored 4 by RNAscope were FISH positive. Forty-six samples were FISH negative, including 17 samples that were scored 3 by RNAscope （37%, 17/46）, 25 samples were scored 2 （54% ,25/46）, and 4 samples were scored 1 （9%, 4/46 ）. Conclusions Breast cancer with HER2 IHC 2 ＋ could be further classified according to in situ mRNA expression status. Among them, RNAseope score of 4 could be one of the interpretation criteria for re-testing IHC 2 ＋ samples. In situ detection of HER2 mRNA may be an additional candidate method of confirmation for HER2 gene amplification or protein overexpression, and has potential clinical utility.