基于小分子化合物Arenobufagin的c-Myc靶向抑制剂的发现及抗肿瘤作用机理研究

The transcription factor c-Myc is over-expressed in various cancers, and as a key gene can drive the initiation and development of cancer, making it a promising therapeutic target of cancer treatment. Recently, we found that compound Arenobufagin (Are) could rapidly down-regulate c-Myc in lung cancer and breast cancer cells at nM level, thereby inhibiting cell proliferation and inducing apoptosis, and showed lower activity against normal cells at the same doses, reflecting a broad spectrum of anticancer activity and selectivity of Are. Although a small number of compounds have been reported to target c-Myc, they generally exhibit activity requiring higher concentrations and their applications are limited. Therefore, the discovery of novel highly effective c-Myc inhibitors is of great significance for cancer treatment. This project is intended to solve the above problems. Using c-Myc highly-expressed cancers, such as breast cancer, as the object of our study, on the one hand, the molecular mechanism of Are-induced down-regulation of c-Myc, identification of c-Myc as a direct target of Are and the in vitro and in vivo anti-tumor activity will be fully investigated; on the other hand, structural modification based on Are, the structure effect relationship studies and targeted validation will be performed to obtain novel c-Myc targeted compounds with highly activity and low toxicity. The successful implementation of this project not only enriches the theoretical research of c-Myc targeted compounds, but also provides new options for cancer therapy.

转录因子c-Myc在多种癌症中高表达，且作为关键基因能驱动癌症的发生发展，是一个很有前途的癌症治疗靶标。最近我们发现化合物Arenobufagin（Are）能在nM水平迅速下调肺癌和乳腺癌等多种癌细胞中c-Myc蛋白，进而抑制细胞增殖并诱导凋亡，且同等剂量对正常细胞活性较低，体现出广谱的抗癌活性以及选择性。虽然目前已报道一些化合物可以抑制c-Myc，但通常发挥活性需较高浓度而极大限制他们的应用，因此，发现新的高效靶向c-Myc抑制剂对于癌症治疗具有重要意义。本项目拟解决上述问题，以乳腺癌等高表达c-Myc的癌症为研究对象，一方面，研究Are下调c-Myc的分子机制、是否直接靶向c-Myc以及体内外抗肿瘤活性；另一方面，对Are进行结构修饰和改造，开展构效关系研究，以期获得高效低毒的靶向c-Myc的化合物。本项目的顺利实施，不仅能丰富化合物靶向c-Myc的理论研究，还可为癌症治疗提供新选择。

转录因子c-Myc在多种肿瘤中高表达且在肿瘤的发生发展过程中起到重要作用，是一个潜在的肿瘤治疗靶标。但目前针对c-Myc的小分子抑制剂仍然面临特异性不高及活性低相关问题，因而，开发新一类的c-Myc小分子抑制剂具有重要意义。本项目从前期发现的降解c-Myc蛋白小分子化合物Are出发，系统研究Are调控c-Myc蛋白的分子机制及其体内外抗乳腺癌活性。结果发现，Are能在翻译后水平促进c-Myc蛋白通过泛素–蛋白酶体通路降解，E3泛素连接酶Fbw7介导c-Myc的泛素化降解过程。Are还可下调PP2A内源性抑制蛋白CIP2A及抑制PP2A下游靶蛋白AKT的活性，且PP2A激活剂DT-061也能下调c-Myc、CIP2A及pAKT，提示Are通过激活PP2A进而诱导c-Myc通过E3泛素连接酶Fbw7介导的蛋白酶体通路降解。体内外活性研究显示，Are不仅能在体外细胞水平显著抑制乳腺癌细胞的增殖并诱导凋亡，同样能够抑制裸鼠乳腺癌细胞移植瘤模型中肿瘤细胞的生长，并显著增强小鼠肿瘤组织中Fbw7蛋白表达以及下调c-Myc蛋白的表达。并且，siRNA干扰实验显示Fbw7-c-Myc通路在Are抗乳腺癌活性中起到重要作用。同时，Are和临床常用化疗药物多西紫杉醇docetaxel (Doc) 联合具有显著的抑制乳腺癌细胞作用。上述结果阐明了Are抑制c-Myc蛋白表达的分子机制，为基于c-Myc蛋白的乳腺癌以及其它c-Myc高表达癌症的治疗提供理论基础。此外，我们通过高压均质法对Are进行脂质体包裹，Are水溶性增强且毒性降低，为基于Are的进一步开发指明了方向。