m6A去甲基化酶ALKBH5阻断PI3K-Akt信号通路抑制结直肠癌增殖转移的分子机制研究

RNA m6A methylation is validated to be closely related to the genesis and development of cancer. ALKBH5 is the demethylase regulating the level of m6A. Our study found that ALKBH5 was downregulated in colorectal cancer tissue, while the level of m6A was elevated. The low expression of ALKBH5 was related with poor prognosis. Furthermore, the functional experiments found that ALKBH5 can inhibit proliferation and metastasis of colorectal cancer cell, whose specific mechanism remains unclear. We initially verified that ALKBH5 can block the PI3K-Akt signaling pathway by transcriptome sequencing and Western blotting. Therefore, we propose the hypothesis : ALKBH5 modulates the level of m6A of targeted gene, thus affecting the function of targeted gene, which blocks the PI3K-Akt pathway to inhibit proliferation and metastasis of colorectal cancer. The present study will identify the candidated genes of ALKBH5 blocking PI3K-Akt pathway, and further validate the targeted gene of ALKBH5 through MeRIP-PCR. Combining the validation of animal model and clinical sample, our study will elucidate the molecular mechnism that the blockade of Akt-PI3K signaling pathways by ALKBH5 inhibits the proliferation and metastasis of colorectal cancer. This study will provide a novel insight into target therapy for colorectal cancer.

研究证实RNA m6A甲基化与肿瘤的发生发展密切相关，ALKBH5是调控m6A水平的去甲基化酶。我们的研究发现ALKBH5在结直肠癌组织中表达下调，其调控的m6A水平升高，且ALKBH5低表达与不良预后相关，进一步功能实验表明ALKBH5可抑制结直肠癌细胞的增殖转移，但具体调控机制不明。申请人基于转录组测序及WB证实ALKBH5可阻断PI3K-Akt信号通路。由此，我们提出：ALKBH5通过改变ALKBH5靶基因的m6A水平，从而影响靶基因的功能，进而阻断PI3K-Akt信号通路抑制结直肠癌增殖转移的假说。本项目拟采用m6A-IP-seq筛选出阻断PI3K-Akt的ALKBH5靶基因；通过MeRIP-PCR鉴定ALKBH5的靶基因；结合动物模型及临床样本的验证，最终阐明m6A去甲基化酶ALKBH5阻断PI3K-Akt信号通路抑制结直肠癌增殖转移的分子机制，从而为基于此的靶向治疗提供依据。

既往研究证实RNAm6A甲基化与肿瘤的发生发展密切相关，ALKBH5是调控m6A水平的去甲基化酶，其在结直肠癌发生发展中作用及临床意义尚不清楚，对其深入探讨，有望为结直肠癌的综合治疗、个体化治疗提供新靶标和新思路，具有重临床转化价值。本项目首先我们探究了RNA m6A 去甲基化酶ALKBH5在结直肠癌中的表达及临床意义，发现ALKBH5 在结直肠癌组织 中的表达水平显著低于正常组织切与更好的预后相关。在此基础上我们构建ALKBH5过表达及敲低细胞系，并开展体内体外功能实验，证实ALKBH5 抑制结直肠癌转移和增殖，进一步通过MeRIP-seq 测序和质谱检测探索了其机制和相关信号通路，发现ALKBH5的下游主要与Wnt信号通路相关。本项目的完成为靶向ALKBH5靶向治疗提供依据。