肿瘤相关成纤维细胞分泌的WNT2作为食管鳞癌治疗新靶向的研究

前期研究发现，肿瘤相关成纤维细胞 (TF)分泌的WNT2蛋白通过激活食管鳞癌细胞Wnt/ β-catenin信号通路以及促进EMT过程的发生，进而参与食管鳞癌的恶性发展进程。预实验结果显示，含有WNT2的条件培养基可诱导食管鳞癌细胞中干细胞标志物表达增强，提示肿瘤微环境因子WNT2与食管肿瘤干细胞的相关性。此外，当鼠源WNT2特异性单克隆抗体中和条件培养基内的WNT2之后，与对照组相比，肿瘤细胞β-catenin的核转移被显著抑制。更重要的是，初步实验结果显示WNT2单抗可有效抑制裸鼠皮下移植瘤的生长，因而成功建立WNT2单抗治疗的体内动物模型。为了进一步了解肿瘤微环境因子WNT2是否可以作为单抗治疗食管癌新的分子靶向，本课题拟采用体外体内实验相结合的方法，详细阐述WNT2单抗通过拮抗旁分泌WNT2对食管鳞癌细胞产生的生物学影响及机制，为发展新型食管癌的治疗方法提供坚实的理论基础。

Esophageal squamous cell carcinoma (ESCC) is one of the most common fatal malignances of the digestive tract. Despite advances in diagnosis and multimodality therapies, ESCC continues to carry a grim prognosis. The major reason for this poor survival is the fact that ESCC frequently metastasizes to regional and distant lymph nodes, even at initial diagnosis. Earlier studies from our group found that the tumor fibroblast secreted WNT2 plays a critical role in the malignant progression of esophageal cancer cells via Wnt/β-catenin signaling pathway, supporting the importance of the cancer microenvironment in the development and progression of esophageal cancer. Our preliminary data have demonstrated that conditioned medium (CM) from WNT2 transfected CHO (Chinese Hamster Ovary) cells induces stemness markers (NANOG, SOX2, OCT4 and ALDH1A1) in KYSE30 cells, indicating that stromal signal WNT2 may promote cancer cell metastasis by inducing stem-like cells. Next, we treated the KYSE30 cells cultured in CHO-WNT2-CM with or without anti-WNT2 mouse monoclonal antibody (mAb). The results showed that the anti-WNT2 mouse mAb inhibits the β-catenin nuclear translocation in KYSE30 cells. Importantly, the therapeutic potential of the anti-WNT2 mouse mAb has been demonstrated in a subcutaneous mouse xenograft model showing the paracrine WNT2 bioactivity. In the present research proposal, we extend our studies to investigate therapeutic potential of the microenvironment factor WNT2 in ESCC. Hence, the objectives in the present proposal will be achieved through the following interrelated specific aims: 1) to investigate the biological effects of anti-WNT2 mouse mAb that block secreted WNT2 in ESCC; 2) to investigate the mechanism underlying the biological effects of anti-WNT2 mouse mAb in ESCC; and 3) to investigate the role of anti-WNT2 mouse mAb in mouse xenograft model. These research objectives will be tackled with a multidisciplinary approach combining biochemistry, molecular and cell biology, and animal model. This proposed study will help elucidate the future clinical implications of the WNT2 therapeutic antibody in esophageal cancer.