基于调控p21和CDK4的天然产物griffipavixanthone抑制卵巢癌耐药性的机制研究

The main obstacle to a successful treatment for ovarian cancer (OC) is the development of drug resistance to platinum-taxanes centered chemotherapy, which represents the gold standard treatment. Therefore, it is urgent to develop novel and effective chemotherapeutic drugs to reduce or even reverse drug resistance in OC. Griffipavixanthone, a natural dimeric xanthone, was poorly understood on its pharmacological roles, and rarely known about the functions on drug resistance in cancer or anti-cancer effects in OC. We found that griffipavixanthone significantly inhibited the cell proliferation in OC cells and mouse models. Also, griffipavixanthone reduced the colony-formation ability, induced cell cycle arrest, and induced cell senescence in OC cells. Furthermore, the inhibitory effects of this compound on the cell viability and colony-formation ability of drug-resistant cells were more significant compared with the sensitive cells. Most importantly, griffipavixanthone could synergize with carboplatin/paclitaxel to reduce drug resistance in OC. In drug-sensitive/resistant cells and subcutaneous tumors, griffipavixanthone treatment upregulated the protein expression level of the tumor suppressor gene p21 and downregulated the expression level of oncogene CDK4. And a more pronounced effect on the expression level of these two proteins was observed when the cells were treated with this compound combined with paclitaxel. Thus, in this application, a scientific hypothesis is proposed that griffipavixanthone inhibits drug resistance of ovarian cancer by modulating the expression of p21, CDK4 and other key proteins. By means of in vitro and in vivo assays, we aim to illustrate the role and molecular mechanism of griffipavixanthone in the inhibition of drug resistance in ovarian cancer. The impact of this application is to provide a natural lead compound and lay a solid foundation for developing new anti-cancer agents to treat drug-resistant ovarian cancer. It could also provide candidate molecular targets for the reversal of drug resistance and the clinical application in ovarian cancer.

对铂类/紫杉醇为主的耐药是卵巢癌化疗失败的主要原因，急需寻求新型有效的化疗药物以降低乃至逆转耐药。Griffipavixanthone是氧杂蒽酮类单体化合物，其药理研究较少，与肿瘤耐药或抗卵巢癌研究未见报道。我们发现其在细胞和动物模型中显著抑制卵巢癌生长，还能抑制克隆形成、阻滞细胞周期、诱导细胞衰老。且其对耐药细胞的抑制作用更强，并能与紫杉醇/卡铂协同增效而降低耐药性。该化合物在亲本和耐药细胞及移植瘤中均能上调抑癌基因p21的蛋白表达，在细胞中下调癌基因CDK4的蛋白表达，且与紫杉醇联用对这两个蛋白的表达影响更显著。由此我们提出科学假设：天然产物griffipavixanthone通过调控p21、CDK4及其它关键因子而抑制卵巢癌的耐药性。本项目将阐明该化合物对卵巢癌耐药的抑制作用及机制，为卵巢癌特别是耐药性卵巢癌治疗药物研发提供候选先导化合物，为卵巢癌临床治疗和耐药逆转提供候选分子靶点。

卵巢癌是致死率最高的女性生殖系统恶性肿瘤。耐药是卵巢癌化疗失败的主要原因，急需寻求新型有效的化疗药物以降低乃至逆转耐药。Griffipavixanthone是一种氧杂蒽酮类单体化合物，其药理活性研究较少，与肿瘤耐药或抗卵巢癌相关研究未见报道。本项目研究发现griffipavixanthone（文中缩写为SZ-NCI-0819）可显著抑制人卵巢癌细胞HeyA8裸鼠皮下异种移植瘤的生长，抑制卵巢癌细胞的增殖活力和平板克隆形成能力，增加卵巢癌细胞活性氧生成，降低细胞线粒体膜电位。SZ-NCI-0819显著增加卵巢癌细胞自噬小体数量，阻滞细胞自噬流，抑制自噬小体与溶酶体的结合，减弱溶酶体水解酶ACP活性，影响卵巢癌细胞中自噬相关基因/蛋白LC3B-II、SQSTM1/p62、ULK1、Beclin-1和SLC7A11等的表达水平。SZ-NCI-0819有效抑制C57鼠同种移植瘤的转移及腹水的形成，抑制卵巢癌细胞的迁移能力和侵袭能力，影响肿瘤转移相关EMT蛋白的表达，例如SZ-NCI-0819处理导致E-Cadherin蛋白表达水平上调，N-Cadherin、Vimentin及Slug蛋白表达水平下调。CCK-8和平板克隆形成实验发现SZ-NCI-0819可降低卵巢癌耐药细胞H-R对紫杉醇的耐药性。通过表达谱芯片分析、自噬数据库等大样本数据研究、分子生物实验和细胞生物学实验，筛选和鉴定了SLC7A11等潜在分子靶点，验证了SZ-NCI-0819可通过调控SLC7A11而降低卵巢癌耐药细胞对紫杉醇的耐药性。在裸鼠皮下移植瘤模型中证明了该单体化合物和紫杉醇联用有协同增效作用。所取得的研究成果为今后进一步开展氧杂蒽酮类单体化合物的抗肿瘤研究提供了实验依据，为卵巢癌治疗药物特别是耐药性卵巢癌治疗药物的研发提供了候选先导化合物，为临床治疗卵巢癌提供了新的候选分子靶点，具有重要的理论和应用价值。