乳腺癌中蛋白质精氨酸甲基转移酶CARM1调控的甲基化修饰的网络、功能、分子机制及其应用研究

Breast cancer is among the most common cancer types observed in women worldwide, with its notoriously high mortality being a serious health issue. The number of reported cases of breast cancer has increased dramatically in China in recent years. Protein methylation has been shown to be as prevalent as phosphorylation and ubiquitination, which is dynamically and precisely regulated by protein methyltransferases and demethylases. Aberrant regulation of protein methylation is tightly linked to the initiation and progression of various types of cancers. Our preliminary data suggested that PRMT4 (protein arginine methyltransferase 4), also called CARM1 (coactivator associated arginine methyltransferase 1), is critical for estrogen receptor (ER)-positive breast cancer cell proliferation and tumor growth, which is highly dependent on its enzymatic activity. Therefore, in the current proposed study, we aim to systematically identify the substrates of CARM1 by quantitative mass spectrometry analysis of enriched peptides from anti-asymmetrical dimethylarginine antibody in both wild type (wt) and CARM1 knockout (KO) cells. Furthermore, we aim to investigate the function of the identified arginine methylated-substrates and their associated arginine methylation in breast carcinogenesis and the underlying molecular mechanisms, establishing a regulatory axis composed of CARM1, arginine methylation and breast carcinogenesis, with arginine methylation at its center. Finally, screening and optimization of small molecules targeting CARM1, and further validation of their efficacy and specificity will be done by using cell- and animal-based breast cancer models. To summarize, our ultimate goal is to reveal new components in breast cancer initiation and progression, which could be potentially serving as drug targets for the prevention and treatment of breast cancer.

本项目围绕指南重点研究方向中的生物大分子修饰在病理变化中的功能和调控机制展开研究。乳腺癌是全球女性人群中最常见且致死率最高的癌症，而中国又是乳腺癌发病率增长最快的国家之一。蛋白质甲基化修饰是细胞内普遍存在的翻译后修饰之一，其受到蛋白质甲基转移酶和去甲基化酶的精密调控。蛋白质甲基化修饰调控的紊乱被证实和癌症发生发展密切相关。申请团队的初步研究表明精氨酸甲基转移酶CARM1对乳腺癌的发生发展至关重要，并且其酶活性起重要作用。基于此，本项目拟通过甲基化富集和定量质谱等系统鉴定CARM1在乳腺癌细胞中的甲基化底物，并通过表观遗传相关技术深入地阐述精氨酸甲基化修饰的功能和潜在分子机制，确立一条以CARM1---精氨酸甲基化修饰---乳腺癌发生发展的分子通道，以期多角度地揭示新的乳腺癌诊疗标志物和药物靶点。项目后期将针对CARM1进行活性小分子筛选，发现先导化合物，为乳腺癌的防治提供新的途径。

本项目以蛋白甲基化修饰领域的引领性研究为基础，集中探索蛋白甲基转移酶CARM1在乳腺癌细胞中调控的甲基化修饰的网络、功能和分子机制。研究表明CARM1对雌激素诱导的基因转录激活和乳腺癌发生发展至关重要，并且CARM1的酶活性在其调控功能中起重要作用，因此靶向CARM1极有可能成为防治乳腺癌的有效方式。在本基金项目的支持下，我们系统鉴定了CARM1在乳腺癌中的甲基化底物，这些底物蛋白参与了细胞内多种重要的生理学进程，并且具有甲基化程度高和甲基化位点序列特殊性等特征。基于此，深入阐述了CARM1介导的甲基化修饰在乳腺癌中的分子作用机制，发现CARM1通过甲基化一些特定的底物蛋白调控雌激素诱导的基因转录激活，进而影响乳腺癌的发生发展。针对CARM1及其介导的甲基化修饰在乳腺癌发生发展中的关键作用，我们通过大规模虚拟筛选得到了特异性靶向CARM1的活性小分子抑制剂，并且证明了其在体内外的抗肿瘤活性，小分子抑制剂的开发和应用将为乳腺癌的防治提供新的策略。

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