肿瘤-睾丸抗原GAGE2B通过STAT3调控骨肉瘤干性介导转移耐药的作用机制研究

Metastasis and drug resistance are key reasons for poor prognosis of tumor. The existence of cancer stem cells may be the main factor of metastasis and resistance, while the activation mechanism of osteosarcoma stem cell is not clear. In this project, we found that tumor-testis antigen GAGE2B was highly expressed in metastatic/drug-resistant osteosarcoma by RNA-seq and tissue microarray analysis. It is known that GAGE family plays an important role in maintaining stemness of spermatogonial stem cell. We found that the metastatic potency, drug resistance and stemness of osteosarcoma altered with the expression change of GAGE2B, and GAGE2B has a co-expressing relationship with stem cell marker CD133. Stem cell transcription factors were further screened, and we found that STAT3 and its downstream pathway is closely related to GAGE2B. Double luciferase assay showed that GAGE2B can regulate STAT3 transcription. Therefore, we hypothesize that GAGE2B regulates osteosarcoma metastasis and resistance through STAT3 activated stem like features. This project aims to confirm the role of GAGE2B in regulating osteosarcoma metastasis, resistance and stemness by detecting clinical samples and cell lines of osteosarcoma, using classical molecular biological methods such as gene silencing/overexpression, IP and ChIP, and to clarify the regulational mechanism of GAGE2B on STAT3 and its abnormal activation mechanism, so as to provide new ideas for stem cell therapy of osteosarcoma.

转移和耐药是肿瘤预后不良的重要原因，肿瘤干细胞存在可能是转移耐药的主要因素，而骨肉瘤干细胞活化的机制还不明确。本项目通过RNA-seq及组织芯片发现肿瘤-睾丸抗原GAGE2B在转移/耐药骨肉瘤中高表达，已知GAGE家族对精原干细胞干性维持有重要作用，前期研究发现GAGE2B表达改变后骨肉瘤转移耐药及干性相应改变，且其与干细胞标志物CD133存在共表达，进一步筛选干细胞转录因子发现STAT3及其下游通路与GAGE2B密切相关，双荧光素酶实验表明GAGE2B可转录调控STAT3。据此我们提出假说：骨肉瘤中GAGE2B通过STAT3调控骨肉瘤干性介导转移耐药。本课题拟通过骨肉瘤临床样本和细胞系检测，利用基因沉默/过表达、IP、ChIP等经典分子生物学方法，证实GAGE2B调控骨肉瘤转移耐药及干性的作用，明确GAGE2B对STAT3的调控机制及其异常活化的机制，为靶向骨肉瘤干细胞治疗提供新思路。

转移和耐药是肿瘤预后不良的重要原因，肿瘤干细胞存在可能是转移耐药的主要因素，而骨肉瘤干细胞活化的机制还不明确。本项目通过RNA-seq及组织芯片筛选到肿瘤-睾丸抗原GAGE2B在转移/耐药骨肉瘤中高表达，已知GAGE家族对精原干细胞干性维持有重要作用，我们构建GAGE2B基因沉默/过表达骨肉瘤细胞系，其转移耐药及干性也相应改变，且其与多种干细胞标志物存在共表达，进一步筛选干细胞转录因子发现STAT3及其下游通路与GAGE2B密切相关，上游分析发现GAGE2B调控STAT3通路细胞膜表面受体OSMR的表达。证实了GAGE2B调控骨肉瘤转移耐药及干性的作用，明确GAGE2B对STAT3信号通路的调控作用机制，为靶向骨肉瘤干细胞治疗策略提供新思路。在本项目的资助下，我们获得授权发明专利3项，拟投稿两篇SCI论文。