藜芦胺类化合物调控AP-1活性抑制皮肤细胞恶性转化的分子机制研究

Transcription factor activator protein-1(AP-1) can be activated by many carcinogens and is an important target for cancer chemoprevention, . Up to now, the compounds regulating AP-1 activity is mainly targeting the upstream kinases of AP-1.The compounds, which can directly targeting AP-1, probably has bettter chemoprevention effect than compounds targeting AP-1 upstream kinase are eagerly wanted.In our previous research, we found veratramine and its analogues inhibit tansactivation activity of AP-1 better than compound PD98059, an ERK inhibitor. Furthermore, veratramine and its analogues supress TPA induced skin cell transformation.However, the kinases which regulating AP-1 activity were not affected by veratramine treatment. These data indicated veratramine and its analogues can directly targeting AP-1 and inhibited its activity. However, the molecular target of veratramine and its analogues, the mechamism of veratramine regulating AP-1 activity are still elusive. In the present study,we will explore the binding target of veratramine and its analogues,investigate the the mechanisms of veratramine and its analogues regulating AP-1 activity.The chemoprevention effect of veratramine also been investigated through 7,12- dimethylbenzanthracene (DMBA)/ 12-O-Tetradecanoyl phorbol 13-acetate (TPA) induced two-stage cancinogenesis model. The present research will set the base for further chemoprevention research targeting AP-1.

转录因子激活蛋白1(AP-1)是皮肤等肿瘤化学预防的重要靶点。以往抑制AP-1活性的小分子化合物均为通过调节AP-1上游激酶的间接抑制剂，寻找AP-1分子直接抑制剂，提高肿瘤化学预防的效果，是目前亟待解决的问题。我们前期研究发现藜芦胺与已有的间接抑制剂比较，能够更有效地抑制AP-1活性并抑制TPA诱导的皮肤细胞转化，激酶芯片研究结果发现藜芦胺类化合物对调控AP-1活性的激酶没有明显抑制。由此推测藜芦胺为AP-1的直接抑制剂。本课题拟进一步研究藜芦胺类化合物作用于AP-1的分子靶点和具体的结合位点；藜芦胺类化合物通过结合位点调控AP-1转录激活活性的分子机制。并采用DMBA/TPA诱导的皮肤乳头状瘤小鼠模型，与AP-1间接抑制剂比较，验证藜芦胺对皮肤乳头状瘤的化学预防作用。本课题旨在阐明藜芦胺类化合物直接抑制AP-1活性的具体分子机制，为以抑制AP-1活性为目标的肿瘤化学预防提供依据。

转录因子AP-1通过调节多种蛋白质编码基因来调控细胞的功能，如细胞增殖、转化、EMT和凋亡。如果能够靶向抑制AP-1的活性，这对肿瘤和炎症的治疗有重要意义。我们研究发现，天然化合物藜芦胺类化合物能够特异性结合位点(TRE 5`-TGACTCA-3`)，这个特异性位点是AP-1的靶向DNA序列，AP-1可以上调含有沸波酯 DNA 应答元件 (TRE 5'-TGACTCA-3') 的基因的转录。基因表达谱芯片结果进一步确证藜芦胺类化合物对AP-1活性的直接调控和抑制AP-1活性对下游蛋白分子表达的差异。但是藜芦胺类化合物对Hedgehog信号通路并没有任何影响，这说明藜芦胺类化合物可能不会有致畸性和毒性。研究还表明，藜芦胺类化合物可以抑制EGF诱导的AP-1的转录活性和JB6细胞的转化。此外，小鼠体内实验的数据表明，藜芦胺类化合物还能够抑制紫外线诱导的AP-1活性。