CLEC5A DEFICIENCY INHIBITS INFLAMMATORY TUMOR MICROENVIRONMENT IN COLORECTAL CANCER

结直肠癌是目前全球致死率最高的癌症类型之一。靶向耗竭T细胞的免疫检查点阻断疗法如 PD-1/PD-L1阻断法在血液病等液体癌患者身上取得了良好的疗效, 但对实体肿瘤的治疗有明显的局限性。其中的主要原因是实体肿瘤内的肿瘤浸润髓系细胞塑造的免疫抑制性微环境限制了免疫治疗的效果。因此，寻找和靶向阻断肿瘤浸润髓系细胞的免疫抑制信号是进一步提升肿瘤免疫治疗疗效的一个重要方向。

本研究分析了实验室先前的结直肠癌单细胞转录组数据，发现CLEC5A在人肿瘤浸润髓系细胞中的表达显著高于癌旁组织。结直肠癌小鼠模型也显示Clec5a在肿瘤浸润髓系细胞中高表达。虽然有报道发现CLEC5A在感染应答中发挥着促炎的功能, 但CLEC5A在结直肠癌中的功能鲜有报道。于是，我们构建了MC38皮下移植瘤和AOM/DSS结直肠癌两个小鼠模型，结果都显示敲除Clec5a会抑制肿瘤生长。

为了进一步比较分析Clec5a对肿瘤微环境的影响，我们利用单细胞转录组技术对野生型和Clec5a^-/-小鼠的AOM/DSS诱导肿瘤进行单细胞转录组测序和细胞图谱绘制。我们发现Clec5a^-/-的小鼠的肿瘤浸润免疫细胞具有更低的炎症打分，这是由于Clec5a缺失抑制了NF-κB 和MAPK等促炎信号通路从而降低了炎症相关细胞因子的表达水平。Clec5a缺失会激活髓系细胞抗原呈递能力并提高糖代谢水平，从而与癌细胞争夺营养物质。

综上，我们的研究表明Clec5a的缺失减弱了炎性肿瘤微环境和增强了免疫应答来抑制肿瘤发展。我们也将借助实验，进一步探究Clec5a的缺失导致的促炎趋化因子水平降低是否限制了肿瘤转移，以及是否能通过髓系细胞来增强CD8⁺ T细胞的细胞毒性能力，从而揭示Clec5a在肿瘤微环境中的调节功能。

Colorectal cancer (CRC) is one of the most lethal cancers in the world. Immune checkpoint inhibitors including anti-PD-1/PD-L1, targeting exhausted T cells, have achieved efficacy in leukemia patients, but has obvious limitations in the treatment of solid tumors. The main reason is that the immunosuppressive tumor microenvironment created by tumor-infiltrated myeloid cells limits therapeutic efficacy. Therefore, finding and targeting the immunosuppressive signals of tumor infiltrating myeloid cells is important for further improving immunotherapy efficacy.

We analyzed the single cell RNA-seq (scRNA-seq) data of CRC generated by our labatory, and found that the expression of CLEC5A in human tumor-infiltrated myeloid cells is significantly higher than that in para-carcinoma samples. Besides, AOM/DSS mouse CRC model also showed Clec5a highly expressed in tumor-infiltrated myeloid cells. Although it has been reported that CLEC5A plays a pro-inflammatory role in response to infection, the function of CLEC5A in CRC is rarely reported. We constructed the MC38 subcutaneous tumor model and AOM/DSS-induced CRC mouse model, both of which showed that tumor was inhibited by Clec5a knockout.

To further analyze the effect of Clec5a on the tumor microenvironment, we generated the cell atlas of AOM/DSS-induced tumors in wild-type and Clec5a^-/- mice using scRNA-seq. We found that the tumor-infiltrated immune cells in Clec5a^-/- mice had a lower inflammatory score, which is caused by the inhibition of NF-κB and MAPK pro-inflammatory signaling after Clec5a deficiency, leading to decreased expression of inflammatory cytokines. Clec5a deficiency promotes the activation of antigen presenting and processing ability of myeloid cells. Moreover, Clec5a depletion would promote a high level of glucose metabolism in macrophages to compete for nutrients with cancer cells.

To sum up, our study suggested that Clec5a deficiency could weaken the inflammatory tumor microenvironment and activate immune response to inhibit cancer progression. Besides, we will further study whether decreasing level of pro-inflammatory chemokines caused by Clec5a deficiency restrained the tumor metastasis, and whether the cytotoxic ability of CD8⁺ T cells can be improved by myeloid cells, thus revealing the regulated function of Clec5a in the tumor microenvironment.

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