黑色素瘤是一种病死率高的恶性肿瘤，进展快、易耐药，针对黑色素瘤细胞干性的分子靶向治疗是近年研究热点。黑色素瘤中医属热盛血瘀肉腐之证，寒凉中药鸦胆子有清热去腐、清火攻实之效，可治疗多种肿瘤，但对黑色素瘤的作用机制尚无深入研究。前期研究显示，鸦胆子主要活性成分鸦胆子苦醇可上调USP10及BASP1，后者进一步抑制其靶基因HOXB9表达。课题组已发表成果表明，HOXB9参与黑色素瘤细胞干性的调控。据此我们推测，鸦胆子苦醇通过USP10/BASP1/HOXB9通路抑制黑色素瘤细胞干性。本课题拟通过泛素化实验和免疫共沉淀等检测USP10与BASP1的相互作用，利用荧光素酶报告基因检测等明确BASP1参与HOXB9转录调控，通过动物模型研究鸦胆子苦醇对黑色素瘤在体内的抑制作用，并分析上述分子的临床相关性。从临床、动物、细胞、分子水平，研究鸦胆子苦醇对黑色素瘤的作用机制，充实“清火攻实”的中医理论内涵。

Melanoma is a malignant skin tumor with a high mortality rate and rapid progress. It is prone to drug resistance. Molecular targeted therapy focusing stemness of melanoma stem cells has been hot topic of research in recent years. According to traditional Chinese medical theory, melanoma belongs to the heat predominance-blood stasis-tissue decay syndrome. The herb of cold and cool nature, traditional Chinese medicine brucea javanica has the effects of clearing heat-fire, inhibiting decay and targeting sthenia pathogenic factors. It has been used to treat several kinds of tumors. However, its effects and mechanism in melanoma hasn’t been deeply studied. Our previous data revealed that the main active constituent of brucea javanica, brusatol, could upregulate USP10 and BASP1, and BASP1 could further inhibit the expression of its downstream target gene HOXB9. Our published data suggested that HOXB9 could regulate the stemness of melanoma cells. Thus, we speculate that brusatol might inhibit melanoma cell stemness through the USP10/BASP1/HOXB9 pathway. This project tries to explore the interaction between USP10 and BASP1 through co-immunoprecipitation and ubiquitination assay, confirm the transcriptional regulation of HOXB9 by BASP1 through Luciferase and realtime PCR, elucidate the inhibitory effects of brusatol on melanoma through tumor xenograft mouse model, and analyze the clinical correlation of the above molecules through statistical analysis. We endeavor to study the mechanism of brusatol’s effects on melanoma on the clinical, animal, celluar and molecular levels, and enrich the connotation of “Qing huo gong shi” traditional theory.