食管鳞癌肿瘤相关成纤维细胞分泌Wnt2调控髓源抑制细胞构筑淋巴结转移前微环境及机制研究

Myeloid-derived suppressor cells (MDSCs) are closely related to the metastasis and prognosis of esophageal squamous cell carcinoma (ESCC), but its underlying regulatory mechanism remains unclear. Preliminarily we found that the WNT2 gene showed increased expression in cancer-associated fibroblasts (CAFs). Furthermore, the secreted Wnt2 protein from CAFs promoted the development of ESCC. Our pilot study showed that the percentage and immunosuppression of MDSCs was reduced by neutralizing Wnt2 protein with anti-mouse Wnt2 monoclonal Antibody in the culture medium. Meanwhile Fzd7, a Wnt2 binding receptor, was highly expressed in MDSCs. Therefore, we proposed that Wnt2 secreted by CAFs might regulate MDSCs to promote ESCC metastasis through the signaling pathway of Fzd7/STAT3. .In this project, we would like to use flow cytometry, ChIP Seq, gene knockout and proteomics in the mouse lymph node metastasis model to clarify the molecular mechanism by which Wnt2 regulates MDSCs via the Fzd7/STAT3 pathway. Then, we will unveil the key protein and potential signaling pathway by which Wnt2 regulates MDSCs. Finally, we will evaluate the relationship between the level of Wnt2 protein and the percentage of MDSCs in peripheral blood, on patient prognosis with ESCC. This project will seek to demonstrate novel cell-to-cell communication in the tumor microenvironment, and how it participates in ESCC metastasis. Our work will provide a new target for ESCC diagnosis, treatment and prognosis monitoring.

髓源抑制细胞（MDSCs）与食管鳞癌转移和预后密切相关，然而其调控机制尚不明确。我们前期发现肿瘤相关成纤维细胞（CAFs）中WNT2基因显著表达；CAFs通过分泌Wnt2促进食管鳞癌进展。预实验结果显示，单抗中和Wnt2后可降低MDSCs比例及其免疫抑制功能，并且Wnt2结合受体Fzd7在MDSCs上高表达。据此我们提出：CAFs分泌的Wnt2可能通过Fzd7/STAT3非经典Wnt信号通路调控MDSCs促进食管鳞癌转移。本项目拟采用小鼠淋巴结转移模型，通过流式细胞术、ChIP-Seq、基因敲除和蛋白质组学等技术，明确Wnt2通过Fzd7/STAT3通路调控MDSCs促转移的分子机制；探索Wnt2调控MDSCs的关键蛋白和信号通路；阐明食管鳞癌患者外周血Wnt2和MDSCs比例与不良预后的关联性。本项目将揭示肿瘤微环境细胞对话参与食管鳞癌转移新途径，并为其诊疗及预后监测提供新的靶点和思路。

髓源抑制细胞（MDSCs）是一群具有免疫抑制能力、未成熟的异质性髓系细胞，是影响结直肠癌（CRC）治疗的主要因素之一。我们前期研究表明，肿瘤相关成纤维细胞（CAFs）是WNT2的主要来源。然而，CAFs分泌的WNT2是否能够通过调控MDSCs进而促进CRC的发生和发展，仍然未知。在这里，我们根据癌症基因组图谱（TCGA）数据集，在275名结肠癌患者和175名直肠癌患者中发现了WNT2表达与MDSCs比例之间存在潜在的正相关性，并在54例初诊未治疗和出现转移的CRC患者中进一步验证了二者的相关性。在体外模型中，WNT2处理后可显著促进MDSCs的比例，增强MDSCs的免疫抑制功能如促进Arg-1和CD38的表达。在小鼠CRC肿瘤模型中，中和WNT2可通过降低MDSCs的比例和免疫抑制功能恢复细胞毒性T细胞功能，抑制肿瘤进展。此外，我们通过RNA-seq分析确定MAPK信号通路在WNT2介导的MDSCs调控中起关键作用。综上，我们系统地证明了在肿瘤免疫抑制微环境（TIME）中，WNT2可显著促进MDSCs的增殖和功能，并提出了可能的分子机制。最后，我们提出联合外周血中WNT2浓度和MDSCs比例作为CRC患者早期诊断和预后监测的潜在应用价值，为CRC患者的诊疗和预后监测提供新思路和方法。