肿瘤干细胞及CXCR4/CXCL12信号通路在喉癌转移中作用的研究

Many tumors,including laryngeal cancer, are maintained by a subpopulation of cells that display stem cell properties, mediate metastasis, and contribute to treatment resistance. These cancer stem cells are regulated by complex interactions with the components of the tumor microenvironment.Cancer stem cell is very important for the occurrence and progression of cancer.The chemokine receptor CXCR4 and its cognate ligand CXCL12 recently have been proposed to regulate the directional trafficking and invasion of malignant tumor. We have identified the CD133 positive cell in laryngeal carcinoma cell. CD133 is one of the marker of cancer stem cell. We also found that CXCL12 was higher in laryngeal cancer, which had positive relationship with that of lymph node metastasis. Our project is to fulfill the following aim based on our previous researches: (1) To block the CXCR4/CXCL12 and PI3K/AKT pathway with RNAi method and LY294002, and then to detect the invasion ability and CXCR4/CXCL12 and PI3K/AKT genes expression of the CD 133 positive Hep-2 cell. (2) To make a mouse model with cancer, and then block the CXCR4/CXCL12 pathway. We will observe the development and metastasis ability of the cancer in the animal model, and then detect the expression of CXCR4/CXCL12 and PI3K/AKT. (3) To collect the laryngeal cancer specimens, and select the CD133 positive cells. We will meature the expression defferences of CXCR4/CXCL12 and PI3K/AKT between the laryngeal carcinoma with lymph node metastasis and the laryngeal cancer without lymph node metastasis. Reframed in the"seed and soil"hypothesis of tumor metastasis, the seeds are cancer stem cells，and the soils is the rich microenviroment. The CXCR4/CXCL12 axis is pivotal for modulating the trafficking of both cancer and normal stem cells. It has great value to elucidate the relationship between cancer stem cell, CXCR4/CXCL12 pathway and the metastasis of laryngeal carcinoma. The project will give rise to clarify the metastasis mechnism of laryngeal cancer, to provide new therapeutic targetic drugs and to direct appropriate clinical treatments.

肿瘤干细胞在肿瘤发生、发展中起重要作用，CXCR4/CXCL12信号通路可以调节干细胞的迁移和定向运动，我们已证实喉癌中存在CD133阳性细胞，并且CXCL12有异常升高，均与喉癌淋巴结转移有关。本项目将在此基础上，（1）分别阻断CXCR4/CXCL12及PI3K/AKT信号通路，观察喉癌Hep-2中CD133阳性的细胞侵袭能力以及各信号通路基因表达的变化；（2）制造裸鼠移植瘤模型，阻断CXCR4/CXCL12信号通路后，检测CD133阳性Hep-2细胞在裸鼠体内肿瘤的生长、转移能力，观察肿瘤内CXCR4/CXCL12以及下游PI3K/AKT基因表达的改变；（3）收集临床喉癌标本，应用流式细胞仪分离CD133阳性细胞，检测其CXCR4/CXCL12以及下游基因PI3K/AKT表达及活性。本研究对于明确肿瘤干细胞及CXCR4/CXCL12在喉癌转移中作用具有重要的理论和科学价值。

背景： 肿瘤干细胞在肿瘤发生、发展中起重要作用，CXCR4/CXCL12可以调节干细胞的迁移和定向运动，我们已发现喉癌组织中存在CD133阳性的细胞亚群，并且CXCL12有异常升高，可见CXCR4/CXCL12及干细胞可能参与喉癌淋巴结转移。方向：本项目以CXCR4/CXCL12信号通路及肿瘤干细胞为切入点，以研究喉癌淋巴结转移机制为目标进行研究。主要内容：培养HEP-2细胞，筛选出CD133阳性的细胞，分别阻断CXCR4/CXCL12信号通路及PI3K/AKT信号通路，观察HEP-2细胞中CD133阳性的细胞侵袭能力以及各信号通路基因表达情况。结果：成功筛选出CD133阳性细胞，敲出CXCR4基因，加入PI-103，经细胞功能试验发现，MTT发现与对照组相比，活细胞数减低（P<0.01）；Transwell试验发现，敲出CXCR4基因，加入PI-103，与对照组比，CD133阳性细胞侵袭力下降（P<0.01）。敲出CXCR4基因，加入PI-103，对于CXCR4、AKT\PI3K、mTOR等表达有重要影响。CXCR4/CXCL12信号通路与PI3K/AKT/mTOR有相互影响。