Escherichia coli Nissle 1917 engineered to express Tum-5 can restrain murine melanoma growth.
Escherichia coli Nissle 1917 engineered to express Tum-5 can restrain murine melanoma growth.
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DOI：
10.18632/oncotarget.20486
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发表时间：
2017-10-17
期刊：
影响因子：
--
通讯作者：
Xia L
中科院分区：
文献类型：
--
作者：
He L;Yang H;Liu F;Chen Y;Tang S;Ji W;Tang J;Liu Z;Sun Y;Hu S;Zhang Y;Liu X;Huang W;Ding X;Xia L
肿瘤的生长和转移依赖于血管生成。因此，抑制肿瘤血管生成已成为近年来很有前途的肿瘤治疗策略。Tumstatin是一种比Endostatin更有效的血管生成抑制剂。本研究将编码肿瘤抑素45-132氨基酸(Tum-5)的抗血管生成活性片段亚克隆到四种不同的诱导表达载体中，并在大肠杆菌BL21(DE3)中成功地进行了可溶性表达。在此基础上，构建了透明颤菌血红蛋白基因启动子PvHb在大肠杆菌Nissle 1917(ECN)中的厌氧诱导表达载体。用免疫印迹和免疫化学方法检测Tum-5在工程菌中的体内外分泌性表达。抗肿瘤作用检测表明，ECN能特异性地定植于肿瘤内，ECN(Tum-5)对B16黑色素瘤的生长有明显的抑制作用。免疫荧光显示ECN(Tum-5)治疗组小鼠肿瘤切片中血管内皮细胞黏附分子-1(PECAM-1/CD31)表达相对减少。ECN处理组小鼠的肝、肾、脾组织形态与对照组无明显差异，说明ECN能被免疫系统清除，不会对小鼠造成全身毒性。这些结果表明，Tum-5基因转移到实体瘤中可能是一种有效的肿瘤治疗策略。
Tumor growth and metastasis depend on angiogenesis. Thus, inhibiting tumor angiogenesis has become promising cancer therapeutic strategy in recent years. Tumstatin is a more powerful angiogenesis inhibitor than endostatin. Anti-angiogenic active fragment encoding amino acids 45–132 (Tum-5) of tumstatin was subcloned into four different inducible expression vectors and successfully solubly expressed in Escherichia coli BL21 (DE3) in this study. Subsequently, an anaerobic inducible expression vector was constructed under Vitreoscilla hemoglobin gene promoter Pvhb in E. coli Nissle 1917 (EcN). The secretory expression of Tum-5 in the engineered bacterium was determined in vitro and in vivo by Western blot or immunochemistry. The anti-tumor effect detection demonstrated that EcN could specifically colonize the tumor, and B16 melanoma tumor growth was remarkably restrained by EcN (Tum-5) in mice bearing B16 melanoma tumor. Abundant infiltrating inflammatory cells were observed in tumor areas of the EcN-treated group through hematoxylin and eosin staining, with a relatively reduced expression of endothelial marker platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31) by immunofluorescence in tumor sections of EcN (Tum-5)-treated mice. No significant morphological differences were observed in the liver, kidney and spleen between EcN-treated mice and the control group, indicating that EcN was cleared by the immune system and did not cause systemic toxicity in mice. These findings demonstrated that the gene delivery of Tum-5 to solid tumors could be an effective strategy for cancer therapy.
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