海绵来源倍半萜醌类NF-κB抑制剂的发现及其抗肿瘤作用机制研究

Abnormal activation of NF-κB pathway is closely related with the generation, metastasis, and drug resistance of tumor, which makes NF-κB pathway become an important target for the development of anticancer drugs. The applicant obtained four marine sponges with potent cytotoxic activity from 23 sponges collected from the Xisha Islands in South China Sea by antitumor activity screening. 16 new sesquiterpene quinones have been isolated from the four marine sponges, some of which showed potent antitumor activity against human myeloma cell line NCI-H929 with IC50 values less than 0.1 μM. The results of preliminary experiments showed that bioactive sesquiterpene quinones prevented proliferation of tumor cells by inhibiting NF-κB pathway. Some bioactive fractions from the four sponges showed more potent cytotoxic and NF-κB inhibitory activities..Based on the above results, the applicant is going to efficiently discover new bioactive sesquiterpene quinones from the four sponges, Dysidea avara, Dysidea fragilis, Spongia sp., and Strepsichordaia sp., by HPLC-DAD-MS/MS combined with bioactive screening, and elucidate their planar structures and absolute configurations. In addition, we will evaluate their cytotoxic and NF-κB inhibitory activities, discuss their structure-activity relationship, investigate their antitumor activity by Hollow Fiber Assay, and reveal the regulation mechanism of bioactive sesquiterpene quinone on NF-κB pathway to find one or two antitumor compounds with definite mechanisms, which have important implications for the development of new marine drugs.

NF-κB通路的异常活化与肿瘤的发生、转移以及耐药性的产生密切相关，是抗肿瘤药物研发的重要靶点。申请人从23种西沙海绵中筛选出4种具有显著细胞毒活性的海绵，并从中发现了16个新结构倍半萜醌，部分化合物对人骨髓瘤NCI-H929等瘤株显示出强细胞毒活性，IC50值小于0.1μM，初步的机制研究发现其活性是通过抑制NF-κB通路产生的。此外还发现多个馏分的细胞毒和NF-κB抑制活性更强。.本项目拟在此基础上，采用HPLC-DAD-MS/MS和活性筛选相结合的方法，从贪婪倔海绵、酥脆倔海绵、沐浴海绵和Dactylospongia sp.等4种西沙海绵中高效发现微量新结构活性倍半萜醌，确定其平面结构和绝对构型，进行细胞毒和NF-κB抑制活性评价，探讨其构效关系，采用中空纤维法评价强活性倍半萜醌的体内抗肿瘤活性，并阐明其对NF-κB通路的调控机制，发现1-2个作用机制明确的抗肿瘤先导化合物。

海绵是海洋天然产物和海洋药物发现的重要源泉之一。申请人采用LC-DAD/MS和活性筛选相结合的方法，从采自西沙群岛的10种海绵样本中分离鉴定了101个化合物，采用HRESIMS、NMR、X-ray单晶衍射、计算ECD法确定了这些化合物的结构和绝对构型，主要为倍半萜醌类化合物，其中新化合物61个，包括5个新颖碳骨架。对这些萜醌类化合物进行了系统的细胞毒和NF-κB抑制活性筛选，发现了51个活性化合物，并对其进行了初步的构效关系探讨，确定了它们的活性官能团，从中优选出1个作用于NF-B信号通路的抗多发性骨髓瘤先导化合物dysihebol A，初阐明了它的抗肿瘤作用机制。在完成预期研究目标的基础上，申请人还对所获得的萜醌类化合物进行了广泛、深入的药理活性筛选，发现dysiarenone能够选择性的抑制脂多糖诱导的小鼠巨噬细胞RAW264.7中COX-2酶的表达，并呈现出良好的剂量依赖关系；frondoplysins A-B对PTP1B酶具有很强的抑制活性，IC50值分别为0.39和0.65 μM，为目前从海绵中发现的活性最强的PTP1B酶抑制剂之一；septosone A对CuSO4诱导的转基因斑马鱼模型中炎症的产生和扩散具有非常强的抑制活性，与强效非甾体抗炎药吲哚美辛活性相当；hippolide J对3株医院来源的致病性真菌具有很强的抑制活性，MIC值为0.125-0.25 μg/mL，与阳性药酮康唑相当；spiroetherones A-B对转基因斑马鱼模型中新生血管的产生具有很强的抑制活性，IC50值分别为2.4和8.7μM。还完成了抗过敏分子dysivillosin A的作用机制研究。Dysiarenone、frondoplysin A septosone A、hippolide J等4个新骨架分子被著名期刊Nat. Prod. Rep.选为“热点报道”。该研究对高效利用我国南海海绵生物资源、阐明海绵萜醌类化合物的结构多样性和生物活性多样性、推动我国海洋抗肿瘤药物的研发都具有重要意义。.以第一作者或通讯作者发表基金标注SCI论文17篇，第一基金标注7篇。申请国家发明专利3项。协助培养了3名博士生和3名硕士生。在本项目的资助下，申请人于2017年获得国家自然科学应急管理项目1项，2019年入选“上海市人才发展资金”、上海交通大学医学院“双百人”计划。