细胞外基质蛋白CTHRC1在HPV16/18型宫颈癌微环境中的调控作用机制研究

Microenvironment of cervical cancer through a variety of ways to promote tumor growth, proliferation, invasion and metastasis, but its targets and mechanisms of its occurrence is unclear. CTHRC1 assists with HPV E6/E7 in cervical cancer, promoting tumor cell growth, migration, invasion and metastasis, we demonstrated that CTHRC1 expression was significantly up-regulated in cervical cancer samples and was closely correlated with lower differentiation, higher invasion, positive lymph node, indicating a poor prognosis. Overexpression of CTHRC1 can promote cervical cancer cells migration and invasion, and this effect is related with inhibiting canonical Wnt pathway and activation of non-canonical Wnt pathway. In the previous study, we analyzed the expression of CTHRC1 in the tissue microarray of cancer samples and cell lines. Comprehensive investigations on detailed function and mechanism of CTHRC1 on tumorigenesis need future verification through multi-faceted, such as, analysis of clinical and pathological features, gene transfection (silence and over-expression) purification of recombinant proteins and monoclonal antibodies, in vitro co-culture and small animal imaging facility assay in vivo. Furthermore using the co-immunoprecipitation technique to find out the interaction receptors of CTHRC1 effected. The study is expected to clarify the molecular mechanisms of cancer microenvironment expression of CTHRC1 which will promote cancer cells invasion and indicate a potential therapeutic strategy for comprehensive treatment of cervical cancer.

宫颈癌微环境通过多种方式引起癌细胞促进肿瘤生长、增殖、侵袭达到远处转移，但其作用靶点和发生机制尚不清楚。CTHRC1具有在宫颈癌微环境中辅助HPVE6/E7引起宫颈癌发生、促进肿瘤细胞生长、迁移、侵袭和转移等重要功能，我们分析其在宫颈癌的低分化、高侵袭、有淋巴结转移的组织中高表达，高表达CTHRC1的宫颈癌细胞能促进肿瘤细胞的迁移侵袭，且通过抑制经典的Wnt通路激活非经典的Wnt通路在启动宫颈癌的侵袭转移过程中起重要作用。我们前期已通过芯片技术分析宫颈癌临床样本及细胞株中CTHRC1表达，本研究拟对其进行蛋白纯化和制备单克隆抗体，用体内外共培养、动物活体成像和免疫共沉淀等等技术寻找相互作用受体，多方位对CTHRC1诱导宫颈癌细胞侵袭转移机制进行深入研究。本研究可望阐明宫颈癌微环境中高表达CTHRC1促进宫颈癌细胞侵袭的分子机制，为临床宫颈癌的综合治疗提供新的研究思路和治疗靶点。

宫颈癌是世界范围内最常见的癌症之一，高危型HPV由E6和E7基因编码的致癌蛋白是导致宫颈癌的重要因子，但其作用机制尚不清楚。近年细胞外基质蛋白作为肿瘤微环境中的重要组成成分在肿瘤作用的研究倍受关注。.本研究首先特异性敲除宫颈癌细胞中E6/E7后通过全基因组表达谱芯片技术、GO和Pathway检索分析并在TCGA和GEO dataset等数据库中进行比对，发现CTHRC1在宫颈癌组织表达明显升高。免疫组化实验显示高表达CTHRC1与宫颈癌临床分期，淋巴结和脉管浸润，肿瘤大小、级别、基质浸润深度均显著相关，血清ELISA实验结果显示CTHRC1在宫颈鳞癌患者血清中高表达，CTHRC1联合SCC-Ag检测可能提高早期诊断的灵敏度和特异度。细胞功能实验结果显示过（高）表达CTHRC1在体内可以促进宫颈癌细胞的侵袭与迁移。体外研究还显示CTHRC1的单克隆抗体在体外能逆转宫颈癌迁移侵袭能力，在体内能抑制小鼠的宫颈癌肺转移水平。.在分子机制研究中发现，转录组表达谱芯片和转录因子预测到RORA、POU2F1两个转录因子可能与CTHRC1的启动区结合， CHIP验证了宫颈癌中转录因子POU2F1可以在转录水平上调控CTHRC1。同时发现P53可与POU2F1的启动区结合，CHIP验证了P53与POU2F1启动区有2个结合位点。荧光素酶报告基因和磷酸化信号通路芯片检测结果显示高表达CTHRC1促进了非经典通路Wnt/PCP，抑制了经典通路Wnt/β-catenin。.总之，研究表明CTHRC1是宫颈癌微环境中受HPVE6/E7调控的重要基质蛋白，有促进侵袭和转移的功能，具有评估宫颈癌预后的生物学标志物潜能，HPV E6/E7可能通过抑制P53上调转录因子POU2F1的表达来引起CTHRC1异常表达的。CTHRC1可能通过WNT非经典通路来促进癌细胞迁徙和侵袭的， CTHRC1的单克隆抗体治疗有望为宫颈癌临床治疗提供新策略。