Sitravatinib联合Niraparib对黏膜黑色素瘤细胞系增殖、凋亡和自噬的影响及其机制  

Effect and mechanism of Sitravatinib combined with Niraparib on proliferation,apoptosis and autophagy in mucosal melanoma cell lines

作　　者：胡子衿 孔燕[1] 吴晓雯 郭倩 郭军[1] HU Zijin;KONG Yan;WU Xiaowen;GUO Qian;GUO Jun(Department of Melanoma and Sarcoma,Key Laboratory of Carcinogenesis and Translational Research(Ministry of Education),Peking University Cancer Hospital&Institute,Beijing 100142,China)

机构地区：[1]北京大学肿瘤医院暨北京市肿瘤防治研究所、黑色素瘤与肉瘤内科恶性肿瘤发病机制及转化研究教育部重点实验室,北京100142

出　　处：《基础医学与临床》2024年第3期295-302,共8页Basic and Clinical Medicine

基　　金：国家自然科学基金(82272848,81972557)。

摘　　要：目的研究抗血管生成药物Sitravatinib联合多聚(腺苷二磷酸[ADP]-核糖)聚合酶抑制剂(PARPi)Niraparib对黏膜黑色素瘤细胞的作用及其可能的机制。方法CCK8法检测Sitravatinib和Niraparib在黏膜黑色素瘤(MM)细胞系的半数抑制浓度(IC 50);CompuSyn模型计算不同浓度联合条件下的联合指数(CI)。细胞集落形成实验测定细胞增殖;流式细胞测量术测定细胞凋亡;Western blot检测蛋白质表达;RT-qPCR检测mRNA表达。结果在人阴道黏膜来源黑色素瘤细胞系(HMVⅡ)和人外阴黏膜黑色素瘤腹股沟淋巴结转移病灶来源细胞系(GAK)中Sitravatinib(2μmol/L)联合Niraparib(20μmol/L)条件下CI值分别为0.19和0.15;与对照组及单药组相比,联合组细胞增殖能力显著下降(P<0.05或P<0.01或P<0.001);细胞凋亡率显著升高(P<0.01或P<0.001),凋亡标志物蛋白质和mRNA表达均显著升高(P<0.001);细胞自噬标志物蛋白质和mRNA表达显著升高(P<0.01或P<0.001);DNA损伤相关蛋白质表达显著升高。而与对照组相比,Sitravatinib单药组和联合组重组酶辐射敏感蛋白51(RAD51)表达显著下降。随着Sitravatinib剂量逐渐升高至2μmol/L,RAD51蛋白和mRNA表达显著下降(P<0.05或P<0.01),BRCA1与BRCA2的mRNA表达显著下降(P<0.05或P<0.01或P<0.001)。结论Sitravatinib联合Niraparib可抑制黏膜黑色素瘤细胞增殖,诱导细胞凋亡并促进细胞自噬,其机制可能与Sitravatinib抑制同源重组修复(HRR)水平相关。Objective To investigate the effect of anti-angiogenic drug Sitravatinib combined with poly(adenosine diphosphate[ADP]-ribose)polymerase inhibitor(PARPi)Niraparib on mucosal melanoma cell lines and its possible mechanism.Methods The CCK8 assay was used to detect the maximal half inhibitory concentration(IC 50)of Sitravatinib and Niraparib targeting at mucosal melanoma(MM)cell lines.CompuSyn was used to detect the Combination Index(CI)in different concentrations of the two drugs.Flow cytometry was used to detect the effect of drugs on cell apoptosis.Colony formation assay was used to detect the effect of drugs on cell proliferation.Western blot was used to detect the protein expressions and RT-qPCR was used to detect mRNA expression.Results CI values was respectively 0.19 and 0.15 for Sitravatinib(2μmol/L)in combination with Niraparib(20μmol/L)in a human vaginal maligant melanoma cell line(HMVII)and a metastasis inguinal lymph node of vulvar malignant melanoma cell line(GAK).Compared with the control group and single-drug groups,the cell proliferation of the combination group was significantly reduced(P<0.05 or P<0.01 or P<0.001).The cell apoptosis rate was significantly increased(P<0.01 or P<0.001).The protein and mRNA expression of apoptosis-related biomarkers significantly increased(P<0.001);In addition,the protein and mRNA expression of cell autophagy biomarkers significantly increased(P<0.01 or P<0.001).The protein expression of DNA damage marker significantly increased.Moreover,compared with the control group,The expression of radiation sensitive protein 51(RAD51)recombinase in the Sitravatinib single-drug group and combination group significantly reduced.As the dose of Sitravatinib gradually increased up to 2μmol/L,the protein and mRNA expression of RAD51 both significantly reduced(P<0.05 or P<0.01),the mRNA expression of BRCA1 and BRCA2 also significantly reduced(P<0.05 or P<0.01 or P<0.001).Conclusions Sitravatinib combined with Niraparib inhibits the proliferation of mucosal melanoma cells,induces cel

关 键 词：黏膜型黑色素瘤 抗血管生成药物 多聚(腺苷二磷酸核糖)聚合酶抑制剂 同源重组修复 

分 类 号：R739.5[医药卫生—肿瘤]