Guerriero Jennifer L, Lin Jia-Ren, Pastorello Ricardo G, Du Ziming, Chen Yu-An, Townsend Madeline G, Shimada Kenichi, Hughes Melissa E, Ren Siyang, Tayob Nabihah, Zheng Kelly, Mei Shaolin, Patterson Alyssa, Taneja Krishan L, Metzger Otto, Tolaney Sara M, Lin Nancy U, Dillon Deborah A, Schnitt Stuart J, Sorger Peter K, Mittendorf Elizabeth A, Santagata Sandro
Division of Breast Surgery, Department of Surgery, Brigham and Women's Hospital, Boston, MA, 02115, USA.
Breast Tumor Immunology Laboratory, Dana-Farber Cancer Institute, Boston, MA, 02215, USA.
NPJ Breast Cancer. 2024 Jan 2;10(1):2. doi: 10.1038/s41523-023-00605-3.
Emerging data suggests that HER2 intratumoral heterogeneity (ITH) is associated with therapy resistance, highlighting the need for new strategies to assess HER2 ITH. A promising approach is leveraging multiplexed tissue analysis techniques such as cyclic immunofluorescence (CyCIF), which enable visualization and quantification of 10-60 antigens at single-cell resolution from individual tissue sections. In this study, we qualified a breast cancer-specific antibody panel, including HER2, ER, and PR, for multiplexed tissue imaging. We then compared the performance of these antibodies against established clinical standards using pixel-, cell- and tissue-level analyses, utilizing 866 tissue cores (representing 294 patients). To ensure reliability, the CyCIF antibodies were qualified against HER2 immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) data from the same samples. Our findings demonstrate the successful qualification of a breast cancer antibody panel for CyCIF, showing high concordance with established clinical antibodies. Subsequently, we employed the qualified antibodies, along with antibodies for CD45, CD68, PD-L1, p53, Ki67, pRB, and AR, to characterize 567 HER2+ invasive breast cancer samples from 189 patients. Through single-cell analysis, we identified four distinct cell clusters within HER2+ breast cancer exhibiting heterogeneous HER2 expression. Furthermore, these clusters displayed variations in ER, PR, p53, AR, and PD-L1 expression. To quantify the extent of heterogeneity, we calculated heterogeneity scores based on the diversity among these clusters. Our analysis revealed expression patterns that are relevant to breast cancer biology, with correlations to HER2 ITH and potential relevance to clinical outcomes.
新出现的数据表明，人表皮生长因子受体2（HER2）肿瘤内异质性（ITH）与治疗耐药性相关，这凸显了评估HER2 ITH新策略的必要性。一种有前景的方法是利用多重组织分析技术，如循环免疫荧光（CyCIF），该技术能够在单细胞分辨率下对单个组织切片中的10 - 60种抗原进行可视化和定量分析。在本研究中，我们鉴定了一组乳腺癌特异性抗体，包括HER2、雌激素受体（ER）和孕激素受体（PR），用于多重组织成像。然后，我们利用866个组织芯（代表294名患者），通过像素、细胞和组织水平分析，将这些抗体的性能与既定的临床标准进行比较。为确保可靠性，针对来自相同样本的HER2免疫组织化学（IHC）和荧光原位杂交（FISH）数据对CyCIF抗体进行了鉴定。我们的研究结果表明，用于CyCIF的乳腺癌抗体组成功鉴定，与既定的临床抗体显示出高度一致性。随后，我们使用经过鉴定的抗体以及针对CD45、CD68、程序性死亡受体1配体（PD-L1）、p53、Ki67、视网膜母细胞瘤蛋白（pRB）和雄激素受体（AR）的抗体，对来自189名患者的567个HER2阳性浸润性乳腺癌样本进行特征分析。通过单细胞分析，我们在HER2阳性乳腺癌中鉴定出四个不同的细胞簇，它们表现出HER2表达的异质性。此外，这些簇在ER、PR、p53、AR和PD-L1表达上存在差异。为了量化异质性程度，我们根据这些簇之间的多样性计算了异质性分数。我们的分析揭示了与乳腺癌生物学相关的表达模式，与HER2 ITH相关且可能与临床结果相关。
