Digital Biology Center, Bio-Rad Laboratories Inc., 7068 Koll Center Pkwy, Ste. 401, Pleasanton, CA 94566, USA.
Methods. 2013 Jan;59(1):S20-3. doi: 10.1016/j.ymeth.2012.09.012. Epub 2012 Oct 2.
The human epidermal growth factor receptor 2 (HER2, also known as erbB2) gene is involved in signal transduction for cell growth and differentiation. It is a cell surface receptor tyrosine kinase and a proto-oncogene. Overexpression of HER2 is of clinical relevance in breast cancer due to its prognostic value correlating elevated expression with worsening clinical outcome. At the same time, HER2 assessment is also of importance because successful anti-tumor treatment with Herceptin® is strongly correlated with HER2 overexpression in the tumor (approximately 30% of all breast tumors overexpress HER2). In a comprehensive national study, Wolff et al. [1] state that "Approximately 20% of current HER2 testing may be inaccurate" which underscores the importance of developing more accurate methods to determine HER2 status. Droplet Digital™ PCR (ddPCR™) has the potential to improve upon HER2 measurements due to its ability to quantitate DNA and RNA targets with high precision and accuracy. Here we present a study which investigates whether ddPCR can be used to assess HER2 transcript levels in formalin-fixed paraffin embedded (FFPE) human breast tumors and whether these ddPCR measurements agree with prior assessments of these same samples by pathologists using immunohistochemistry (IHC) and in some cases fluorescence in situ hybridization (FISH). We also determined the copy number of HER2 in these samples as compared to the CEP17 reference gene.
Clinical FFPE samples were successfully studied using ddPCR and compared to results from standard FISH and IHC methodology. The results demonstrate that ddPCR can rank order the samples in complete agreement with the current standard methods and that ddPCR has the added benefit of providing quantitative results, rather than relying on the expert skill of a seasoned pathologist for determination.
人表皮生长因子受体 2（HER2，也称为 erbB2）基因参与细胞生长和分化的信号转导。它是一种细胞表面受体酪氨酸激酶和原癌基因。由于其预后价值与临床结果恶化相关，HER2 的过表达在乳腺癌中具有临床相关性。同时，HER2 评估也很重要，因为曲妥珠单抗®的成功抗肿瘤治疗与肿瘤中 HER2 的过表达强烈相关（约 30%的所有乳腺癌肿瘤过表达 HER2）。在一项全面的全国性研究中，Wolff 等人[1]指出，“目前大约 20%的 HER2 检测可能不准确”，这突显了开发更准确的方法来确定 HER2 状态的重要性。由于其能够高精度和准确性地定量 DNA 和 RNA 靶标，液滴数字 PCR（ddPCR）有可能改善 HER2 测量。在这里，我们进行了一项研究，调查 ddPCR 是否可用于评估福尔马林固定石蜡包埋（FFPE）人乳腺肿瘤中的 HER2 转录水平，以及这些 ddPCR 测量值是否与病理学家使用免疫组织化学（IHC）和在某些情况下荧光原位杂交（FISH）对这些相同样本的先前评估一致。我们还确定了这些样本中 HER2 的拷贝数与 CEP17 参考基因的比较。
使用 ddPCR 成功研究了临床 FFPE 样本，并与标准 FISH 和 IHC 方法的结果进行了比较。结果表明，ddPCR 可以完全按照当前标准方法对样本进行排序，并且 ddPCR 具有提供定量结果的额外优势，而不是依赖经验丰富的病理学家的专业技能来进行确定。
