Tumor Stroma Interactions, Department of Oncology, Luxembourg Institute of Health, Luxembourg, Luxembourg.
Faculty of Science, Technology and Medicine, University of Luxembourg, Esch-sur-Alzette, Luxembourg.
Front Immunol. 2020 Oct 20;11:578176. doi: 10.3389/fimmu.2020.578176. eCollection 2020.
In the past 20 years, the interest for the tumor microenvironment (TME) has exponentially increased. Indeed, it is now commonly admitted that the TME plays a crucial role in cancer development, maintenance, immune escape and resistance to therapy. This stands true for hematological malignancies as well. A considerable amount of newly developed therapies are directed against the cancer-supporting TME instead of targeting tumor cells themselves. However, the TME is often not clearly defined. In addition, the unique phenotype of each tumor and the variability among patients limit the success of such therapies. Recently, our group took advantage of the mass cytometry technology to unveil the specific TME in the context of chronic lymphocytic leukemia (CLL) in mice. We found the enrichment of LAG3 and PD1, two immune checkpoints. We tested an antibody-based immunotherapy, targeting these two molecules. This combination of antibodies was successful in the treatment of murine CLL. In this methods article, we provide a detailed protocol for the staining of CLL TME cells aiming at their characterization using mass cytometry. We include panel design and validation, sample preparation and acquisition, machine set-up, quality control, and analysis. Additionally, we discuss different advantages and pitfalls of this technique.
在过去的 20 年中，人们对肿瘤微环境（TME）的兴趣呈指数级增长。事实上，现在人们普遍认为 TME 在癌症的发生、发展、免疫逃逸和耐药性方面起着至关重要的作用。血液系统恶性肿瘤也是如此。相当多新开发的疗法是针对支持癌症的 TME 的，而不是针对肿瘤细胞本身。然而，TME 通常没有明确定义。此外，每个肿瘤的独特表型和患者之间的可变性限制了这些疗法的成功。最近，我们小组利用液质联用技术揭示了小鼠慢性淋巴细胞白血病（CLL）背景下特定的 TME。我们发现了两种免疫检查点 LAG3 和 PD1 的富集。我们测试了一种针对这两种分子的基于抗体的免疫疗法。这种抗体组合在治疗小鼠 CLL 方面取得了成功。在本方法文章中，我们提供了一个详细的方案，用于对 CLL TME 细胞进行染色，旨在使用液质联用技术对其进行特征描述。我们包括面板设计和验证、样本准备和采集、仪器设置、质量控制和分析。此外，我们还讨论了这项技术的不同优点和缺点。
