G9a小分子抑制剂对ER阳性乳腺癌的抑制效果、分子机制和临床意义研究

Epigenetic small-molecule inhibitors, especially the inhibitors that target Histone Methyltransferases (HMTs), recently became very efficient and safe drug candidates for cancer therapy. Estrogen Receptor alpha (ER) is a well-known target for the endocrine therapy of breast cancer. Histone H3K9 HMT G9a is a coactivator of ER. G9a targeting gene SOX2 is also reported to be a downstream molecule of ER/miR-140 signaling pathway. Our previous study found G9a methylates ER protein at both K235 and K303, while K303R mutation is associated with high risk of breast cancer. Depletion of G9a greatly abolishes ER target genes transcription and cell proliferation of ER positive breast cancer cells. Here we propose that small-molecule inhibitor of G9a could inhibit tumor growth and metastasis of breast cancer through its inhibition of ER/miR-140/SOX2 signaling pathway. In this study, we will investigate the anti-tumor effect and molecular mechanism of the G9a small-molecule inhibitor in ER positive breast cancer, both in vivo and in vitro, through cellular biology assays, ChIP, RNA sequencing and mutation analysis of patients samples. Our study will provide novel strategy for the precision medicine and personalized therapy of breast cancer.

以表观遗传修饰酶特别是组蛋白甲基化酶为靶点的小分子抑制剂，近年来成为令人瞩目的靶向抗肿瘤药。雌激素受体（ER）是乳腺癌内分泌治疗的关键靶点。组蛋白甲基化酶G9a是ER的转录共促活因子，其靶基因包括ER/miR-140通路的下游分子SOX2。我们最近的研究发现G9a甲基化ER的K235和K303位点，而K303的点突变是乳腺癌高风险的预测指标。G9a敲除后严重抑制ER靶基因的表达和ER阳性乳腺癌细胞的增殖。因此我们预测G9a的小分子抑制剂能够通过下调信号通路ER/miR-140/SOX2而控制乳腺癌的生物活性。为了验证这一假说，我们将通过乳腺癌细胞模型、裸鼠模型及病例样本三个水平，使用细胞功能实验、ChIP、RNAseq、突变分析等技术，筛选对抑癌作用最优的G9a小分子抑制剂并研究其信号通路和临床意义。我们的研究将为乳腺癌的精准医疗和个性化用药提供新的理论依据。

乳腺癌现有的临床诊疗标志物都是基于几个经典的临床特征和细胞肿瘤生物学的标记，比如雌激素受体 ER和人类表皮生长因子受体HER-2等。这些标志物是目前乳腺癌治疗分组的黄金标准，但是也有几点局限性，比如准确性和特异性有限，大量病理数据人工观察容易产生主观偏见等。本课题通过乳腺癌中筛选出9个具有鉴别诊断意义的表观遗传酶，包括5个蛋白甲基化酶，2个蛋白去甲基化酶，1个DNA甲基化酶； 在细胞模型和动物模型中进一步鉴定EHMT1/2等表观遗传酶的表达与乳腺癌的分化程度有关；使用近300例乳腺癌组织芯片样本库鉴定了一批以EHMT1/2介导的H3K9me为代表的组蛋白表观遗传修饰在乳腺癌组织中的表达水平在恶性程度不同的乳腺癌中存在差异表达；另外本研究还测试了电子图像特征作为机器学习数据进行建模和诊断预测的意义。 综上，本研究初步建立肿瘤智能免疫监测和诊断系统，为乳腺癌和肺癌的个性化智能医疗提供基础。

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