基于环状RNA circSMARCA5/miR-127/MAF1轴探讨华蟾素抑制肺鳞癌细胞增殖的机制研究

Compared with lung adenocarcinoma, lung squamous cell carcinoma (LUSC) lacks effective targeted drugs. Our previous studies found that huanchansu(HCS), the traditional Chinese medicine toad venom preparations, has the exact clinic antitumor effects for LUSC. However, HCS mechanism of action in LUSC has not been unveiled completely. HCS treatment could effectively inhibit tumor proliferation of subcutaneous tumor-bearing nude mice. Microarray data showed that the expression levels of a large number of circular RNAs and mRNAs significantly changed in the tumor tissue in response to HCS treatment, of which, circSMARCA5 was the most obvious one. Co-expression network analysis showed that the important tumor-suppressor factor MAF1 had a strong positive correlation with it. This project will take these as the breakthrough point to put forward the scientific hypothesis: HCS could upregulate circSMARCA5, promoting MAF1 expression, and thus inhibiting the proliferation of LUSC. Therefore, we intend to investigate the function of circSMARCA5 in proliferation using clinical samples, animal models and molecular cell biological means; after that, bioinformatics analysis prediction and related experiments will be taken to verify that circSMARCA5 may elevate MAF1 expression by working as a miR-127 absorption;then, clarify the relationship between the in vivo efficacy of HCS treatment and this the pathway. Above all, we characterize a new mechanism mediated by circSMARCA5/miR-127/MAF1 axis in HCS function against LUSC, providing a novel insight into LUSC therapy.

与肺腺癌相比，肺鳞癌缺乏有效的靶向药物。课题组临床研究发现中药蟾酥制剂华蟾素治疗肺鳞癌具有确切疗效但其主导的分子机制尚不清楚。利用华蟾素处理皮下成瘤裸鼠可有效抑制肿瘤增殖。全转录组芯片数据显示药物处理组瘤体内大量环形RNA及mRNA发生显著改变，其中circSMARCA5上调最明显。共表达网络分析显示重要的抑癌因子MAF1与其相关性最高。 本项目以此为切入点提出科学假说：华蟾素通过上调 促进MAF1表达从而抑制肺鳞癌增殖。故拟结合临床样本、动物模型及分子细胞学手段深入研究circSMARCA5调控肺鳞癌细胞增殖方面的功能；其后采用生信手段及相关实验证实circSMARCA5可能通过吸附miR-127上调下游靶点MAF1；进一步验证华蟾素治疗肺鳞癌的体内疗效及与该通路的关系，最终阐明华蟾素治疗肺鳞癌的作用机制，为华蟾素临床治疗肺磷癌提供基础实验依据。

中药蟾酥提取物华蟾素（cinobufacini）是临床应用较广的抗肿瘤中药制剂，属于新一代单味多组份的广谱抗癌中药。其主要成分为蟾毒灵也广泛应用于抗肿瘤研究。课题组临床研究发现中药蟾酥制剂华蟾素治疗肺鳞癌具有确切疗效但其主导的分子机制尚不清楚。实验发现华蟾素在体内外有效抑制肿瘤增殖。全转录组芯片数据显示药物处理组大量环形RNA及mRNA发生显著改变，其中circSMARCA5显著上调。环状 RNA（Circular RNA，circRNA） 是最新发现的一种非编码 RNA， 它是一类不具有 5’末端和 3’末端、 以共价键形成环形结构的 RNA。环状 RNA在生理过程和疾病中具有特殊的生物学功能。其高度稳定性和特异性使之有望成为肿瘤潜在的预测和治疗靶点。近几年药物与circRNA 的关系也引起了科研人员的关注。本项目后期以circSMARCA5主要研究内容，结合临床样本、动物模型及分子细胞学手段深入分析circSMARCA5及对肺癌细胞增殖的影响及调控的相关分析机制。我们通过体内、外实验验证华蟾素具有诱导肺鳞癌细胞凋亡的能力。circSMARCA5确实可以受到华蟾素的调控，并介导部分华蟾素的功能，是其重要的调控因子。根据生物信息学预测的circSMARCA5-miR-127-MAF1调控轴可能并不是华蟾素发挥作用的关键，但是华蟾素对circSMARCA5-免疫调控轴的影响，可能是其发挥作用的关键。结果我们发现华蟾素处理和circSMARCA5过表达都可以影响细胞因子和免疫系统通路，表明华蟾素影响肺鳞癌细胞增殖的分子机制可能是通过circSMARCA5对免疫系统的调控通路介导的。该实验结果最终阐明华蟾素治疗肺鳞癌的作用机制，为华蟾素临床治疗肺磷癌提供基础实验依据。本项目实验结果在写高等级SCI论文一篇，在投中文核心一篇，发表科技核心一篇，培养研究生多名。