The Immune Landscape of Tumor-Associated Macrophage Reprogramming.

Tumor-associated macrophages (TAMs) generally acquire an immunosuppressive and tumor-promoting phenotype, which may contribute to tumor resistance to immunotherapy. We previously showed that suppression of microRNA activity through genetic Dicer1 inactivation rewires TAM's transcriptomes and prompts their immunostimulatory activation. This phenotypic switch enhanced recruitment and activation of CD8+ cytotoxic T cells (CTLs) and improved the efficacy of immunotherapy in mouse cancer models. Here, we performed single-cell RNA sequencing of whole tumors grown in either wild-type mice or mice with macrophage-specific Dicer1 deletion. The analysis of multiple cell populations, including several discrete monocyte and macrophage subsets, indicated broad and convergent immunostimulatory programming of the tumor microenvironment (TME), which was dependent on CTL-derived interferon-gamma (IFN?), in mice with DICER-deficient macrophages. Intriguingly, dynamic inferences on monocyte/macrophage ontogeny and differentiation by pseudotime analysis revealed trajectories associated with progression into cell cycle, monocyte-to-macrophage differentiation, and transition from an immunostimulatory to an immunosuppressive phenotype in tumors with DICER-proficient macrophages. Dicer1 deficiency interfered with this trajectory and stalled TAMs at an intermediate state between immature monocytes and macrophages with T cell-stimulatory capacity, thereby impeding immunosuppressive TAM development. This translated into enhanced response to antiangiogenic immunotherapy in an immunotherapy-resistant model of non-small cell lung cancer. Cycling/M2-like macrophages were conserved in human melanoma and hepatocellular carcinoma and should represent a more promising therapeutic target than the bulk of TAMs. Overall design: Comparative gene expression profiling of mouse MC38 tumors with Dicer know-out or wild type.

肿瘤相关巨噬细胞（Tumor-associated macrophages, TAMs）通常会获得免疫抑制性且促肿瘤的表型，这可能会促使肿瘤产生免疫治疗耐药性。我们此前的研究表明，通过遗传手段敲除Dicer1以抑制微RNA（microRNA, miRNA）活性，可重塑TAMs的转录组并促使其向免疫激活表型转化。这种表型转换可增强CD8+细胞毒性T淋巴细胞（CD8+ cytotoxic T cells, CTLs）的招募与活化，并在小鼠癌症模型中提升免疫治疗的疗效。本研究对野生型小鼠以及巨噬细胞特异性Dicer1敲除小鼠体内生长的完整肿瘤进行了单细胞RNA测序（single-cell RNA sequencing, scRNA-seq）。对包括多个单核细胞与巨噬细胞亚群在内的多种细胞群进行分析后发现，在DICER缺陷型巨噬细胞小鼠体内，肿瘤微环境（tumor microenvironment, TME）呈现出广泛且趋于一致的免疫激活编程模式，该过程依赖于CTL分泌的干扰素-γ（interferon-gamma, IFNγ）。有趣的是，通过拟时间分析对单核细胞/巨噬细胞的个体发育与分化进行动态推演后发现，在DICER功能正常的巨噬细胞小鼠肿瘤中，存在与细胞周期进程、单核细胞向巨噬细胞分化，以及从免疫激活表型向免疫抑制表型转换相关的分化轨迹。Dicer1缺陷会干扰这一分化轨迹，将TAMs阻滞在未成熟单核细胞与具备T细胞活化能力的巨噬细胞之间的中间状态，从而阻碍免疫抑制性TAMs的发育。这一效应在非小细胞肺癌（non-small cell lung cancer, NSCLC）的免疫治疗耐药模型中，转化为对抗血管生成免疫治疗的应答增强。增殖型/M2样巨噬细胞在人类黑色素瘤与肝细胞癌中均保守存在，相较于绝大多数TAMs，其或可成为更具潜力的治疗靶点。实验整体设计：对Dicer1敲除型与野生型小鼠MC38肿瘤进行比较性基因表达谱分析。