HOXB-AS3/HOXB7/PAK4信号轴调控结直肠癌侵袭转移的分子机制研究

lncRNA HOXB-AS3 and HOXB7 gene were upregulated in colorectal tissue samples and cell lines with high metastatic potential, both of which related with poor prognosis in patients with colorectal cancer. There was a positive correlation between the expression of HOXB-AS3 and HOXB7. We demonstrated that both of HOXB-AS3 and HOXB7 could promote the invasion and migration of the colorectal cancer cells. HOXB-AS3 could regulate the expression and biological function of HOXB7. We also found that HOXB7 can active the PI3K/AKT and ERK/MAPK pathways. Besides, HOXB7 could regulate the expression of PAK4. The purpose of this study is: a) to confirm the function of HOXB-AS3 and HOXB7 in colorectal cancer using 3-D culture, whole-body visualized animal model system etc, b) explore the mechanism of HOXB-AS3 regulating HOXB7 by ChIP, dual luciferase reporter assay etc., c) reveal the mechanism of HOXB7 regulating PI3K/AKT and ERK/MAPK pathways via PAK4. Finally, we may reveal the molecular mechanism of HOXB-AS3/HOXB7/PAK4 axis in metastasis of colorectal cancer , which will provide new drug target for treatment of colorectal cancer metastasis.

我们前期研究发现lncRNA HOXB-AS3和HOXB7基因在结直肠癌高转移细胞株及结直肠癌组织中高表达，两者表达呈正相关，且均与患者不良预后相关。HOXB-AS和HOXB7均可促进结直肠癌细胞的侵袭迁移能力，HOXB-AS3可调控HOXB7的表达及生物学功能。HOXB7可调控PAK4激酶的表达，可激活PI3K/AKT、ERK/MAPK信号通路。本研究拟构建过表达及干扰细胞株模型，用三维培养、可视化结直肠癌原位及转移动物模型等实验验证HOXB-AS3及HOXB7对结直肠癌细胞侵袭迁移能力的影响；用染色质免疫共沉淀、双荧光素酶基因报告系统等方法分析HOXB-AS3调控HOXB7的分子机制，HOXB7通过PAK4调控PI3K/AKT、ERK/MAPK信号通路的分子机制。揭示HOXB-AS3/HOXB7/PAK4信号轴在结直肠癌侵袭转移中的分子机制，为结直肠癌转移的治疗和药物筛选提供新靶点。

结直肠癌是常见的恶性肿瘤。原位杂交检测结果显示lncRNA HOXB-AS3在结直肠癌中表达定位于细胞核。实时荧光定量PCR检测发现lncRNA HOXB-AS3在肿瘤组织中较癌旁组织中表达更低；其在肿瘤组织中低表达与5-氟尿嘧啶治疗反应不佳相关；且lncRNA HOXB-AS3的低表达与高T分期，高N分期相关。构建过表达lncRNA HOXB-AS3的SW620和HCT116细胞株。药敏实验结果示在含5-氟尿嘧啶生长环境中，过表达组较对照空载组，药物抑制肿瘤细胞增殖的抑制率增加。流式细胞术检测结果示过表达组较对照空载组，细胞凋亡率增加；且western blot检测示促凋亡蛋白Bax和促凋亡活性酶Caspase 3表达均上调。构建干扰lncRNA HOXB-AS3的SW480和LS174T细胞株。药敏实验发现干扰组较对照组，药物抑制肿瘤细胞增殖的抑制率降低。流式细胞术检测结果示干扰组细胞凋亡率降低；且western blot检测结果示促凋亡蛋白Bax和促凋亡活性酶Caspase 3表达均下调。Western blot检测结果示过表达lncRNA HOXB-AS3细胞组NFkB信号通路活性降低， JNK信号通路活性上调；而干扰组则NFkB信号通路活性上调，JNK信号通路活性下调。本研究揭示了lncRNA HOXB-AS3通过调节NFkB和JNK 信号通路活性，诱导凋亡，从而增强结直肠癌细胞对5-FU的敏感性。本课题研究结果为lncRNA HOXB-AS3是结直肠癌治疗的重要预测因子及靶点提供了理论基础。