光辉霉素消耗特异性蛋白1并激活p53以介导恶性胸膜间皮瘤细胞的衰老和凋亡。
Mithramycin Depletes Specificity Protein 1 and Activates p53 to Mediate Senescence and Apoptosis of Malignant Pleural Mesothelioma Cells.
作者信息
Rao Mahadev, Atay Scott M, Shukla Vivek, Hong Young, Upham Trevor, Ripley R Taylor, Hong Julie A, Zhang Mary, Reardon Emily, Fetsch Patricia, Miettinen Markku, Li Xinmin, Peer Cody J, Sissung Tristan, Figg William D, De Rienzo Assunta, Bueno Raphael, Schrump David S
机构信息
Thoracic Epigenetics Section, Thoracic and GI Oncology Branch, National Cancer Institute, Bethesda, Maryland.
Laboratory of Pathology, National Cancer Institute, Bethesda, Maryland.
出版信息
Clin Cancer Res. 2016 Mar 1;22(5):1197-210. doi: 10.1158/1078-0432.CCR-14-3379. Epub 2015 Oct 12.
PURPOSE
Specificity protein 1 (SP1) is an oncogenic transcription factor overexpressed in various human malignancies. This study sought to examine SP1 expression in malignant pleural mesotheliomas (MPM) and ascertain the potential efficacy of targeting SP1 in these neoplasms.
EXPERIMENTAL DESIGN
qRT-PCR, immunoblotting, and immunohistochemical techniques were used to evaluate SP1 expression in cultured MPM cells and MPM specimens and normal mesothelial cells/pleura. MTS, chemotaxis, soft agar, β-galactosidase, and Apo-BrdUrd techniques were used to assess proliferation, migration, clonogenicity, senescence, and apoptosis in MPM cells following SP1 knockdown, p53 overexpression, or mithramycin treatment. Murine subcutaneous and intraperitoneal xenograft models were used to examine effects of mithramycin on MPM growth in vivo. Microarray, qRT-PCR, immunoblotting, and chromatin immunoprecipitation techniques were used to examine gene expression profiles mediated by mithramycin and combined SP1 knockdown/p53 overexpression and correlate these changes with SP1 and p53 levels within target gene promoters.
RESULTS
MPM cells and tumors exhibited higher SP1 mRNA and protein levels relative to control cells/tissues. SP1 knockdown significantly inhibited proliferation, migration, and clonogenicity of MPM cells. Mithramycin depleted SP1 and activated p53, dramatically inhibiting proliferation and clonogenicity of MPM cells. Intraperitoneal mithramycin significantly inhibited growth of subcutaneous MPM xenografts and completely eradicated mesothelioma carcinomatosis in 75% of mice. Mithramycin modulated genes mediating oncogene signaling, cell-cycle regulation, senescence, and apoptosis in vitro and in vivo. The growth-inhibitory effects of mithramycin in MPM cells were recapitulated by combined SP1 knockdown/p53 overexpression.
CONCLUSIONS
These findings provide preclinical rationale for phase II evaluation of mithramycin in patients with mesothelioma.
目的
特异性蛋白1（SP1）是一种致癌转录因子，在多种人类恶性肿瘤中过表达。本研究旨在检测恶性胸膜间皮瘤（MPM）中SP1的表达，并确定靶向SP1对这些肿瘤的潜在疗效。
实验设计
采用qRT-PCR、免疫印迹和免疫组化技术评估培养的MPM细胞、MPM标本以及正常间皮细胞/胸膜中SP1的表达。采用MTS、趋化性、软琼脂、β-半乳糖苷酶和Apo-BrdUrd技术评估SP1敲低、p53过表达或光神霉素处理后MPM细胞的增殖、迁移、克隆形成、衰老和凋亡。利用小鼠皮下和腹腔异种移植模型检测光神霉素对MPM体内生长的影响。采用微阵列、qRT-PCR、免疫印迹和染色质免疫沉淀技术检测光神霉素介导的基因表达谱，以及联合SP1敲低/p53过表达，并将这些变化与靶基因启动子内的SP1和p53水平相关联。
结果
与对照细胞/组织相比，MPM细胞和肿瘤中SP1 mRNA和蛋白水平更高。SP1敲低显著抑制MPM细胞的增殖、迁移和克隆形成。光神霉素消耗SP1并激活p53，显著抑制MPM细胞的增殖和克隆形成。腹腔注射光神霉素显著抑制皮下MPM异种移植瘤的生长，并在75%的小鼠中完全根除间皮瘤癌。光神霉素在体外和体内调节介导癌基因信号传导、细胞周期调控、衰老和凋亡的基因。联合SP1敲低/p53过表达可重现光神霉素对MPM细胞的生长抑制作用。
结论
这些发现为光神霉素用于间皮瘤患者的II期评估提供了临床前依据。
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