m6A修饰介导CDCP1+亚群参与Kras突变型大肠癌复发转移的机制研究

Colorectal cancer (CRC) patients harboring Kras mutations are more likely to develop recurrence of lung metastases.CDCP1+ subpopulation is the unit of selection responsible for this process.However,the underlying mechanism(s) still remain elusive.In our preliminary studies, we observed that CDCP1+ subpopulation derived from mutant Kras CRCs exhibited decreased m6A levels, which was mediated by fat mass and obesity-associated protein (FTO), and that Foxd3 served as a major downstream target of FTO. Here, we aim to uncover the critical role of m6A demethylation in pulmonary recurrence both in vitro and in vivo. Utilizing RIP, ChIP-PCR and promoter reporter system, we attempt to clarify the potential metabolic involvement in regulating the demethylase activity of FTO. Moreover, with a lentiviral-based knockdown approach, we try to determine whether targeting FTO in vivo represents a novel therapeutic approach to prevent pulmonary recurrence in patients with mutant Kras CRCs. Thus, further support of this program would provide a new insight into the process during post-therapeutic recurrence of mutant Kras CRCs and a promising perspective for therapy against this subtype of cancer.

肺脏是Kras突变型大肠癌患者术后复发转移的最常见部位，作为复发根源的CDCP1+亚群在此过程中扮演了关键角色，然而相关作用机制目前尚未阐明。我们在前期研究中利用体内复发转移模型和高通量筛选等方法，初步证实Kras突变型大肠癌中CDCP1+亚群发生了FTO介导的m6A去修饰，Foxd3是FTO下游主要的效应靶基因。本项目拟利用不同遗传背景的同源性大肠癌细胞系、大肠癌临床标本和小鼠体内模型，从体内外两方面进一步探讨m6A去修饰主导CDCP1+亚群参与术后肺脏复发转移的机制；采用RIP、ChIP-PCR和启动子基因报告等技术，阐明调控FTO活性的代谢偶联机制；并通过动物实验观察靶向干扰FTO对抑制术后复发转移的作用，从而为有效改善其临床预后提供新思路和新手段。

肺脏是Kras突变型大肠癌患者术后复发转移的最常见部位，作为复发根源的CDCP1+亚群在此过程中扮演了关键角色，然而相关作用机制目前尚未阐明。在本项研究中，我们发现Kras突变型大肠癌CDCP1+亚群中Kras和CDCP1协同作用下谷氨酸代谢发生了重编程，α-酮戊二酸/琥珀酸比值显著增高，从而促进了FTO介导的m6A 去修饰。随后的高通量筛选结果显示，Foxd3是受FTO调控的主要效应靶点之一。一方面，Foxd3通过激活OCT1使CDCP1+亚群处于静息状态，从而产生化疗耐受性；另一方面，Foxd3通过促进CDCP1+亚群自我更新潜能帮助其在肺脏发生定植。因此，靶向CDCP1+亚群中的FTO-Foxd3能够成为潜在的有效预防Kras突变型大肠癌术后复发转移的治疗靶点。