汉黄芩素调节乳腺癌组蛋白乙酰化诱导细胞衰老的作用及机制研究

Pro-senescence drugs are promising for cancer treatment, but such kind of effective drugs have not yet been developed. As such, researches for understanding the functions and regulation of senescence in tumors are being actively carried out around the world currently. Cellular epigenome changes dramatically in the progress of senescence, but its roles in this process are elusive. Whether epigenetic drugs can be used to promote cancer senescence requires more investigations. Wogonin, a type of flavonoid, has strong anti-tumor effect in a plethora of cancers and is in Phase I clinical trial in China now. Nevertheless, understanding its working mechanisms facilitates its application in cancer therapy. In this respect, we hypothesize that Wogonin regulates the metabolism of Acetyl-CoA in breast cancer cells, altering their epigenome, such as histone acetylation and chromatin structure. These epigenetic changes induce cellular senescence and the expression of senescence-associated-secretory-proteins (SASP), activating the immune surveillance mechanism to clear the senescent cancer cells. In this study, we will investigate the functions of Wogonin in inducing cellular senescence and immune clearance in cultured breast cancer cells and with animal models. A series of gene-modified breast cancer cell lines will be constructed by CRISPR-Cas9 technology or lentivirus. These cells are then used to reveal the molecular mechanisms underlying the pro-senescence activity of Wogonin by employing a variety of genomic, proteomic and epigenetic methods. Our results will highlight the importance of the epigenetic axis ASF1/CBP/P300-H3K56Ac-CAF1 in controlling cellular senescence, flourishing the regulatory mechanisms of senescence and uncovering potential targets for developing pro-senescence drugs and epigenetic drugs. In addition, it diversifies the working mechanisms of Wogonin in cancer treatment, which will be beneficial for its clinical application.

促细胞衰老药物在肿瘤治疗中正引起广泛关注，但尚缺乏有效药物。细胞衰老伴随表观遗传学图谱的剧烈改变，但这些变化对细胞衰老的影响仍不清楚，表观遗传学药物可否用于促细胞衰老疗法也需探索。汉黄芩素在肿瘤治疗中很有前景，对其作用机理的研究将促进其临床应用。本课题将研究汉黄芩素可否影响肿瘤细胞的乙酰辅酶A代谢，改变其表观遗传学图谱如组蛋白乙酰化和染色质结构，诱导细胞衰老及相关分泌蛋白的表达；这些蛋白是否进而激活机体的免疫监测机制清除肿瘤细胞。我们将在细胞和动物水平上考察汉黄芩素诱导乳腺癌细胞衰老和免疫清除的功能；运用基因编辑技术，结合基因组、蛋白质组和表观遗传学探讨汉黄芩素的作用机制。从而揭示表观遗传相关蛋白ASF1/CBP/P300-H3K56Ac-CAF1轴调控细胞衰老的功能，丰富细胞衰老的分子机制，为药物开发提供靶点；阐明汉黄芩素作用于该通路诱导细胞衰老而抑瘤的新机理，促进其在肿瘤治疗中的应用。

衰老细胞处于周期阻滞状态而不能分裂增殖，因此促细胞衰老药物可以有效的抑制肿瘤生长，在肿瘤治疗中正引起广泛关注。本课题在体外和体内证明了汉黄芩素具有引起乳腺癌细胞衰老的作用。汉黄芩素降低组蛋白乙酰转移酶KAT5和KAT6B的表达，同时可能抑制细胞核中PDH复合物的活性，降低细胞中的组蛋白乙酰化和染色质的可接近性，进而抑制细胞中抗氧化蛋白TXNRD2的表达，增加乳腺癌细胞中活性氧水平，引起细胞DNA损伤，诱导乳腺癌细胞衰老。在衰老的乳腺癌细胞中，转录因子NFκB被激活，引起细胞衰老相关蛋白的表达和分泌，募集巨噬细胞和NK细胞并促进巨噬细胞向M1性分化，表明汉黄芩素可以激活机体的免疫监测机制。此外还初步研究了汉黄芩素的衍生物GL-V9对衰老乳腺癌细胞的影响。发现汉黄芩素衍生物GL-V9使衰老细胞的溶酶体碱化，抑制自噬，诱导细胞凋亡，从而能够在体内和体外清除化疗药物如多柔比星等诱导的衰老的乳腺癌细胞。因此本课题丰富了细胞衰老的分子机制，阐明了汉黄芩素抑制乳腺癌的新机理，有助于促进汉黄芩素及其衍生物在肿瘤治疗中的应用。