The recent technical and computational advances in single-cell sequencing technologies have significantly broaden our toolkit to study tumor microenvironment (TME) directly from human specimens. The TME is the complex and dynamic ecosystem composed of multiple cell types, including tumor cells, immune cells, stromal cells, endothelial cells, and other non-cellular components such as the extracellular matrix and secreted signaling molecules. The great success on immune checkpoint blockade therapy has highlighted the importance of TME on anti-tumor immunity and has made it a prime target for further immunotherapy strategies. Applications of single-cell transcriptomics on studying TME has yielded unprecedented resolution of the cellular and molecular complexity of the TME, accelerating our understanding of the heterogeneity, plasticity, and complex cross-interaction between different cell types within the TME. In this review, we discuss the recent advances by single-cell sequencing on understanding the diversity of TME and its functional impact on tumor progression and immunotherapy response driven by single-cell sequencing. We primarily focus on the major immune cell types infiltrated in the human TME, including T cells, dendritic cells, and macrophages. We further discuss the limitations of the existing methodologies and the prospects on future studies utilizing single-cell multi-omics technologies. Since immune cells undergo continuous activation and differentiation within the TME in response to various environmental cues, we highlight the importance of integrating multimodal datasets to enable retrospective lineage tracing and epigenetic profiling of the tumor infiltrating immune cells. These novel technologies enable better characterization of the developmental lineages and differentiation states that are critical for the understanding of the underlying mechanisms driving the functional diversity of immune cells within the TME. We envision that with the continued accumulation of single-cell omics datasets, single-cell sequencing will become an indispensable aspect of the immune-oncology experimental toolkit. It will continue to drive the scientific innovations in precision immunotherapy and will be ultimately adopted by routine clinical practice in the foreseeable future.

单细胞测序技术的最新技术和计算进展显着拓宽了我们直接从人体样本研究肿瘤微环境 (TME) 的工具包。 TME是由多种细胞类型组成的复杂而动态的生态系统，包括肿瘤细胞、免疫细胞、基质细胞、内皮细胞以及其他非细胞成分，如细胞外基质和分泌的信号分子。免疫检查点阻断疗法的巨大成功凸显了TME在抗肿瘤免疫方面的重要性，并使其成为进一步免疫治疗策略的首要目标。单细胞转录组学在 TME 研究中的应用已经对 TME 的细胞和分子复杂性产生了前所未有的分辨率，加速了我们对 TME 内不同细胞类型之间的异质性、可塑性和复杂交叉相互作用的理解。在这篇综述中，我们讨论了单细胞测序在了解 TME 多样性及其对单细胞测序驱动的肿瘤进展和免疫治疗反应的功能影响方面的最新进展。我们主要关注渗透到人类 TME 中的主要免疫细胞类型，包括 T 细胞、树突状细胞和巨噬细胞。我们进一步讨论了现有方法的局限性以及利用单细胞多组学技术进行未来研究的前景。由于免疫细胞在 TME 内不断激活和分化以响应各种环境线索，因此我们强调整合多模式数据集以实现肿瘤浸润免疫细胞的回顾性谱系追踪和表观遗传分析的重要性。 这些新技术能够更好地表征发育谱系和分化状态，这对于理解驱动 TME 内免疫细胞功能多样性的潜在机制至关重要。我们预计，随着单细胞组学数据集的不断积累，单细胞测序将成为免疫肿瘤学实验工具包中不可或缺的一部分。它将继续推动精准免疫治疗的科学创新，并在可预见的未来最终被常规临床实践所采用。