乏氧激活HIF-1/ETS-1/CK2正反馈通路在肺癌细胞向肺癌内皮细胞转化中的作用及其介导抗血管治疗耐药的探讨

The endothelial cells are presumed to be genomically stable and thus less predisposed to the development of the therapy resistance commonly encountered in tumor cells. Despite this major theoretical advantage, there is still a confusing question remains in the clinic that patients will eventually develop resistance to anti-angiogenic therapies. We and others have recently described a new origin for the lung endothelial cell population, which arise directly from lung cancer epithelium under hypoxia. Like their ancestral tumor cells, these tumor-derived endothelial cells (TDECs) might possess inherent genomic instability (GI). In addition, the transformation process of TDECs and their angiogenic function were independent of VEGF. These findings suggest that TDECs'generation is a　crucial　phenomenon contributing to development of resistance to anti-angiogenic therapies. However, the molecular mechanism driven transformation of TDECs remains largely unexplored. Previous study from our group indicated that: CK2 and ETS-1 were both up-regulated significantly in lung cancer cells exposed to hypoxia. The enhancement of ETS-1 driven the transformation of lung cancer cells into lung TDECs. Whereas, the supression of CK2 kinase activity could block the process. In vitro transformation of lung TDECs were independent of VEGF. The current proposal aims: to compare the detailed properties, with regards to genotype, epigenetics and funtion, between lung TDECs and normal endothelial cells，as well as to further prove the role of lung TDECs in mediating resistance to anti-angiogenic therapy; to unreval the role of a positive feedback formed by Hypoxia/HIF-1/ETS-1/CK2 pathway on the transformation of lung TDECs, and to further investigate the associations and regulation mechanism within the pathway; to study the possibility of utilizing HIF-1/ETS-1/CK2 as predictive markers for early detecting of potential resistance case in lung cancer anti-VEGF therapies, and in the mean time to provide promising targets to resolve such resistance.

内皮细胞基因组稳定、不易耐药的理论与临床上抗肿瘤血管治疗耐药的现状相悖。我们及其它研究证实：肺癌内皮细胞可由肺癌细胞直接转化而来，故继承肺癌细胞的基因不稳定性，且转化过程不依赖VEGF途径，可能是介导抗血管治疗耐药的关键。但转化机制尚不清楚。前期工作发现：肺癌细胞中，乏氧可诱导CK2及ETS-1表达上调；ETS-1激动剂促进肺癌细胞向肺癌内皮细胞转化；而CK2激酶抑制剂起阻断作用；另外，贝伐单抗无法阻断该转化过程。因此提出假说：乏氧激活HIF-1/ETS-1/CK2正反馈通路促进肺癌细胞向肺癌内皮细胞转化，进而介导对抗血管治疗耐药。本项目拟比较肺癌内皮细胞与正常内皮细胞在基因稳定性、表型、功能和对贝伐单抗治疗反应上的差异，初步证实肺癌内皮细胞与耐药的相关性；并阐明HIF-1/ETS-1/CK2正反馈通路对肺癌内皮细胞转化的作用；为解决肺癌抗血管治疗耐药问题提供新的策略及靶点。

最新研究发现肿瘤血管中存在着一种特殊类型的内皮细胞，即肿瘤内皮样细胞，由肿瘤细胞本身转化而成。其不仅具备普通内皮细胞的一般特性，还惊奇地“继承”了肿瘤细胞的基因组不稳定性和容易耐药等特点。肿瘤内皮样细胞的产生、存活和发挥功能均不依赖 VEGF-VEGFR 信号。肿瘤内皮样细胞的上述特点可能是抗肿瘤血管治疗耐药的重要原因，且其转化机制尚不清楚。研究发现乏氧环境可以促进肿瘤细胞向肿瘤内皮样细胞转化，其关键效应因子 HIF-1α可激活 ETS-1 启动子，而后者调控下游多个血管新生相关基因，特异性促进肿瘤血管新生和内皮样细胞的发育。我们的研究发现，非小细胞肺癌组织中 HIF-1α 和 ETS-1 的表达水平呈正相关，且两者呈现高表达的肺癌组织血供丰富，瘤体体积大，疾病分期晚且预后不佳。进一步体外实验发现，肺癌细胞经乏氧诱导表达内皮标记物，具备内皮细胞生物学特征，且对贝伐单抗部分耐药。体内实验发现乏氧诱导后的肺癌细胞形成的移植瘤生长迅速，血供丰富，且对贝伐单抗治疗不敏感。我们进一步发现相比VEGF，ETS-1 表达水平对于乏氧环境更加敏感，乏氧关键效应因子 HIF-1α可以有效激活 ETS-1 启动子以及上调蛋白表达，常氧条件下上调肺癌细胞 ETS-1 表达可以诱导肺癌细胞内皮化，促进肺癌血管新生，导致贝伐单抗耐药；而下调乏氧肺癌细胞系中 ETS-1 表达可以逆转肺癌细胞内皮化，抑制肺癌血管新生，增强贝伐单抗治疗疗效。综上，HIF-1α/ETS-1 信号通路介导了肺癌细胞向肺癌内皮样细胞的转化，促进肺癌血管生成，进而诱导肺癌贝伐单抗治疗耐药。本研究不仅丰富了肺癌抗血管治疗耐药理论，而且为肺癌抗血管生成治疗提供了新靶点，最终为临床新药物上市提供新启示。