Department of Pathology, Regina Elena National Cancer Institute, Via Elio Chianesi 53, 00144, Rome, Italy.
Department of Medical Sciences, University of Turin, Pathology Unit, Via Santena 7, 10126, Turin, Italy.
J Exp Clin Cancer Res. 2017 Oct 13;36(1):143. doi: 10.1186/s13046-017-0613-2.
This is a retrospective cross sectional study aimed to verify whether Multiplex Ligation-dependent Probe Amplification (MLPA), a quantitative molecular assay, may represent a valuable reflex test in breast cancer with equivocal HER2 expression by immunohistochemistry and HER2 gene signals/nucleus (s/n) ranging between 4.0 and 5.9 by in situ hybridization.
A series of 170 breast carcinomas scored as 2+ for HER2 expression by immunohistochemistry, were selected from our files and analyzed in parallel by silver in situ hybridization and by MLPA. According to ASCO-CAP 2013 guidelines, 54/170 tumors, displaying 4.0-5.9 HER2 gene s/n, were defined as low amplified (ratio ≥ 2) or equivocal (ratio < 2) on the basis of centromere enumeration probe 17 (CEP17) status. An independent set of 108 score 2+ breast cancers represented the external validation set. Concordance between the two techniques was assessed through the use of Cohen's K statistic.
A concordance rate of 78.2% (Cohen's K statistic: 0,548 95% CI:[0,419-0,677]) between in situ hybridization and MLPA was found in the whole series of 170 cases and of 55.5% (Cohen's K statistic: -0,043 95% CI:[-0,271-0,184]) in the 54 tumors presenting 4.0-5.9 HER2 gene s/n. By MLPA, we found HER2 amplification or gain in 14% of the 21 BC presenting a disomic status and in 18% of the 33 BC presenting a CEP17 > 2.0. These data were further confirmed in the external validation set. Interestingly, the 54 low amplified/equivocal breast carcinomas presented a frequency of hormonal receptor positivity significantly higher than that observed in the amplified tumors and similar to the non-amplified one (p = 0.016 for estrogen receptor and p = 0.001 for progesterone receptor).
To avoid to offer patients an ineffective therapy, HER2 status should be studied more thoroughly in low amplified and equivocal cases which can have lower response rates and shorter time to progression to trastuzumab. In this context, our data indicate that MLPA may be a reliable, objective supporting test in selecting HER2 positive breast cancer patients.
本研究为回顾性横断面研究，旨在验证多重连接依赖探针扩增（MLPA）这种定量分子检测手段是否可作为一种有用的辅助检测方法，用于对免疫组化结果为 2+且 HER2 基因信号/核（s/n）比值为 4.0-5.9 的乳腺癌患者进行检测。
本研究从我们的存档中选择了一系列免疫组化结果为 2+的乳腺癌患者，共 170 例，通过银原位杂交和 MLPA 进行平行分析。根据 ASCO-CAP 2013 指南，54/170 例肿瘤的 HER2 基因 s/n 比值为 4.0-5.9，根据着丝粒计数探针 17（CEP17）的状态，将其定义为低扩增（比值≥2）或不确定（比值<2）。另外还选择了 108 例免疫组化评分 2+的乳腺癌患者作为外部验证集。通过 Cohen K 统计量评估两种技术之间的一致性。
在整个 170 例患者中，原位杂交和 MLPA 之间的一致性率为 78.2%（Cohen K 统计量：0.548，95%CI：[0.419-0.677]），在 54 例 4.0-5.9 HER2 基因 s/n 比值的肿瘤中，一致性率为 55.5%（Cohen K 统计量：-0.043，95%CI：[-0.271-0.184]）。通过 MLPA，我们发现 21 例二倍体肿瘤中有 14%存在 HER2 扩增或增益，33 例 CEP17>2.0 的肿瘤中有 18%存在 HER2 扩增或增益。这些数据在外部验证集中得到了进一步证实。有趣的是，54 例低扩增/不确定的乳腺癌患者的激素受体阳性率显著高于扩增肿瘤，与非扩增肿瘤相似（雌激素受体 p=0.016，孕激素受体 p=0.001）。
为避免为患者提供无效的治疗，对于低扩增和不确定的病例，应更深入地研究 HER2 状态，因为这些病例的反应率可能较低，并且对曲妥珠单抗的进展时间也较短。在这种情况下，我们的数据表明 MLPA 可能是一种可靠的、客观的辅助检测方法，可用于选择 HER2 阳性乳腺癌患者。
