JMJD2B调控氨基酸代谢途径影响大肠癌发生发展的机制研究

Altered tumor metabolism including amino acids metabolic reprogramming is now a generally regarded hallmark of cancer. Abnormal amino acid uptake, alterations in the amino acids derived from the autophagy-lysosome pathway and so on lead to the amino acid metabolic reprogramming. Post-translational modifications of histones are critical for the maintenance of rapid tumor cell proliferation adapting to nutrient-deprived tumor microenvironment through regulation of metabolic reprogramming. Our previous study found that the expression of histone demethylase JMJD2B was significantly elevated in colorectal carcinoma tissues. We also found that JMJD2B was up-regulated during glucose deprivation, and JMJD2B silencing inhibited CRC cells proliferation, along with a significant change in intermediates of amino acid metabolism. Moreover, JMJD2B may also regulate autophagy-lysosome pathway during glucose deprivation. We therefore assume that JMJD2B may be emerging as a potential regulator of amino acid metabolic alterations in the occurrence and development of CRC. Based on our former results, in this study we aim to ① clarify the effect of amino acid metabolic alteration by JMJD2B on the CRC malignant phenotype; ②explore the underlying mechanism of JMJD2B regulating amino acid metabolism during glucose deprivation; ③analyze the correlation between JMJD2B and related autophagy-lysosome protein or amino acid transporter, and their relationship with clinical pathological features and prognosis. Our research may provide theoretical basis for potential therapeutic target in the treatment of CRC.

肿瘤发生发展中存在氨基酸代谢异常等细胞代谢改变，其具体机制包括氨基酸摄入异常以及自噬-溶酶体途径影响的氨基酸生成变化等。而组蛋白翻译后修饰可影响代谢重编程，在肿瘤细胞适应自身生长微环境改变中发挥重要作用。近年我们发现，组蛋白去甲基化酶JMJD2B在大肠癌组织中呈高表达。在葡萄糖缺乏等代谢应激下，JMJD2B表达上调；静默JMJD2B抑制细胞增殖，伴随氨基酸代谢中间产物明显变化。亦发现JMJD2B参与调控自噬-溶酶体途径。据此我们推测，JMJD2B可能作用于氨基酸代谢过程而影响大肠癌发生发展。因此，本研究拟在前期工作基础上：①明确JMJD2B是否通过调控大肠癌细胞氨基酸代谢而影响其恶性表型；②深入研究JMJD2B影响大肠癌氨基酸代谢的具体机制；③探讨大肠癌组织JMJD2B和自噬-溶酶体蛋白或氨基酸转运蛋白表达的相关性及与临床病理特征及预后的关系。本课题的实施将为大肠癌临床治疗提供新思路。

背景：近年来，肿瘤中细胞代谢的改变越来越受到关注；且组蛋白甲基化修饰作为一种重要的翻译后修饰， 参与肿瘤代谢重编程。而JMJD2B作为组蛋白去甲基化酶，其在大肠癌细胞代谢改变中的作用尚未明确。.主要研究内容：采用非靶向代谢质谱评价JMJD2B在葡萄糖剥夺条件下对大肠癌细胞代谢的作用。通过细胞活力、免疫印迹、克隆形成实验和流式细胞术分析JMJD2B诱导的氨基酸改变对大肠癌细胞存活的影响。在体外（大肠癌细胞系）和体内（异种移植模型）模型中通过免疫荧光染色、染色质免疫沉淀分析、电子显微镜等评估JMJD2B影响大肠癌细胞代谢的潜在机制。采用免疫组织化学方法在大肠癌组织标本中评估JMJD2B与LC3B表达的相关性。从而明确在葡萄糖剥夺条件下，JMJD2B通过影响肿瘤细胞氨基酸代谢而促进大肠癌细胞的存活。.重要结果：在葡萄糖缺乏的情况下，JMJD2B敲除的大肠癌细胞中发生了明显的代谢重编程，尤以氨基酸代谢产物变化更为显著。静默JMJD2B可降低葡萄糖缺乏引起的某些氨基酸的水平。其中，天冬酰胺（Asn）、苯丙氨酸（Phe）和组氨酸（His）在葡萄糖剥夺下当JMJD2B被敲除时，可促进大肠癌细胞存活。机制上，JMJD2B的下调通过LC3B的表观遗传调节抑制大肠癌细胞中的自噬，继而在葡萄糖剥夺下降低细胞内氨基酸（Asn、Phe、His）水平，从而抑制大肠癌细胞的存活。利用裸鼠异种移植模型，我们证实抑制JMJD2B可降低氨基酸（Asn、Phe、His）水平。而过表达LC3B，则可恢复JMJD2B敲除所导致的对肿瘤生长和氨基酸水平的抑制作用。此外在60例人大肠癌组织中，JMJD2B高表达的组织中更有可能出现LC3B的高表达（P<0.001）。.结论和科学意义：以上结果表明，JMJD2B在葡萄糖缺乏时部分通过LC3B的表观遗传调节而以自噬方式维持了大肠癌细胞内氨基酸水平，从而在大肠癌恶性进程中发挥作用。这一发现为未来大肠癌患者的治疗提供了新的思路。