基于单细胞测序技术的肿瘤相关巨噬细胞亚群的定义、功能和调控机制研究

The tumor microenvironment is a complex ecology of cells that evolves and interacts closely with the immune cells recruited to the tumor site. Macrophages in the tumor tissues are defined as ’tumor-associated macrophages, TAMs’, which are particularly abundant and generally play a protumoral role through multiple mechanisms, which can stimulate angiogenesis and enhance tumor cell invasion, motility, and intravasation. TAMs are also immunosuppressive, preventing anti-tumor T cell effector function and promoting tumor evasion. Typically, macrophages are classified as two subtypes in terms of their activation status and function, namely M1 or M2. However, this classification is oversimplified and is not applicable to TAM, which is a heterogeneous cell population under complex tumor microenvironment. In this grant, we proposed to use single-cell RNA sequencing to identify potential new TAM populations, to understand their temporal and spatial patterns in the process of tumor development, and to further dissect their specific function towards tumor growth and underlying molecular programs. Elucidation of these questions will provide new insights into how tumor microenvironment interacts with TAM. Precisely targeting this population may thus provide better therapeutic potentials or better combination therapy strategy in cancer treatment.

肿瘤间质中的巨噬细胞是一类被称之为肿瘤相关巨噬细(tumor-associated macrophages, TAMs)的重要炎性反应细胞，参与了肿瘤的发生、生长、侵袭及转移过程。TAM由于本身极大的可塑性，在肿瘤的复杂微环境中，其表型和功能研究也变得更加复杂。传统的M1或者M2的分型，已经很难应用于肿瘤微环境中TAM的分型。更精细的对于TAM的分类及其对应亚类群的功能研究，亟待探索和解决。本项目借助小鼠低免疫原性肿瘤模型，通过单细胞测序，单细胞基因表达谱聚类分析，了解探索肿瘤微环境中，TAM亚群的精细分类和潜在新亚群的定义。并阐明TAM新亚群的时空调控、发展轨迹、动态变化、不同亚群的关系，等等。同时进一步深入探索TAM新亚群的潜在功能、调控机制，以及对于肿瘤发生发展的作用。该项研究不仅为全面解析肿瘤微环境和TAM的相互作用提供线索，也为未来开发更为精准的靶向TAM的治疗策略提供依据。

肿瘤相关巨噬细(tumor-associated macrophages, TAMs)参与了肿瘤的发生、生长、侵袭及转移过程。TAM由于本身极大的可塑性，在肿瘤的复杂微环境中，其表型和功能研究也变得更加复杂。传统的M1或者M2的分型，已经很难应用于肿瘤微环境中TAM的分型。更精细的对于TAM的分类及其对应亚类群的功能研究，亟待探索和解决。本项通过单细胞测序，单细胞基因表达谱聚类分析，揭示肿瘤微环境中，巨噬细胞亚群的精细分类和潜在新亚群的定义，并确定新巨噬细胞亚群的表面膜蛋白标志基因 G7。并通过CRISPR/Cas9技术成功构建G7-EGFP-IRES-Cre小鼠和G7-DTR小鼠，通过流式、免疫荧光在细胞、组织层面明确G7+巨噬细胞亚群的分布特征。通过DT特定清除G7表达的巨噬细胞亚群，明确该群巨噬细胞对小鼠肿瘤表型的显著促进作用。并进一步通过RNA-seq profiling明确了G7+巨噬细胞亚群的特征基因表达和潜在调控机制。G7作为膜蛋白，目前正在筛选针对它的阻断型抗体，特异性地清除该群细胞。该项研究不仅为全面解析肿瘤微环境和TAM的相互作用提供线索，也为未来开发更为精准的靶向TAM的治疗策略提供依据。