Transcriptional regulation of inducible nitric oxide synthase gene therapy: targeting early stage and advanced prostate cancer
Transcriptional regulation of inducible nitric oxide synthase gene therapy: targeting early stage and advanced prostate cancer
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DOI：
10.1002/jgm.1495
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发表时间：
2010-09-01
影响因子：
3.5
通讯作者：
McCarthy, Helen O.
中科院分区：
文献类型：
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作者：
Coulter, Jonathan A.;Page, Natalie L.;McCarthy, Helen O.
使用肿瘤类型特异性人骨钙素（hOC）启动子，我们以前报道过在体外激素不依赖型前列腺癌细胞中强启动子激活。在本研究中，我们在体外和体内模型中对组织特异性启动子前列腺特异性膜抗原（PSMA）和肿瘤型特异性hOC启动子驱动诱导型一氧化氮合酶（iNOS）转基因进行了比较研究。方法采用克隆法测定体外细胞毒性。通过Griess试验定量测定一氧化氮的表达。体内抗肿瘤效果是通过将构建体直接注射到PC3异种移植物的肿瘤生长延迟来确定的。此外，死后解剖肿瘤，检查形态学差异以及凋亡蛋白表达的变化。结果PSMA/iNOS对雄激素依赖型和非雄激素依赖型细胞系均产生细胞毒性。一氧化氮定量证实细胞毒性的增加与一氧化氮的产生直接相关。在用表达iNOS的构建物治疗的所有组中，肿瘤生长延迟从hOC/iNOS单剂量治疗组的10.7天到hOC/iNOS多剂量组的最多52.2天不等。对肿瘤内iNOS和cleaved poly （adp -核糖）聚合酶蛋白表达的评估显示，这两种蛋白的表达均显著上调，表明细胞毒性通过内在凋亡途径介导。结论采用PSMA/iNOS和hOC/iNOS结构治疗后，肿瘤生长明显延迟，无不良副作用。我们认为这些发现为开发针对早期和晚期前列腺癌的系统递送PSMA/iNOS或hOC/iNOS提供了基础。版权。(C) 2010 John Wiley & Sons, Ltd。
Background Using the tumour type specific human osteocalcin (hOC) promoter, we have previously reported strong promoter activation in hormone independent prostate cancer cells in vitro. In the present study, we present a comparative study of the tissue specific promoter prostate specific membrane antigen (PSMA), and the tumour-type specific hOC promoter driving the inducible nitric oxide synthase (iNOS) transgene using both in vitro and in vivo models.Methods In vitro cytotoxicity was assessed by clonogenic assay. Quantification of nitric oxide expression was determined by the Griess test. In vivo anti-tumour efficacy was determined by tumour growth delay following direct intra-tumoural injection of the constructs into PC3 xenografts. In addition, tumours were dissected post mortem and examined for morphological differences as well as changes in apoptotic protein expression.Results PSMA/iNOS produced cytotoxicity in both androgen dependant and independent cell lines. Nitric oxide quantification confirmed that increased cytotoxicity was directly associated with nitric oxide production. Tumour growth delays were observed in all groups treated with the iNOS-expressing constructs ranging from 10.7 days for the hOC/iNOS single dose treatment group to a maximum of 52.2 days for the hOC/iNOS multiple dose group. Intra-tumoural assessment of iNOS and cleaved poly (ADP-ribose) polymerase protein expression demonstrated a significant up-regulation of both proteins, indicating cytotoxicity mediated through the intrinsic apoptotic pathway.Conclusions Highly significant tumour growth delay coupled with no detrimental side-effects were observed following treatment with the PSMA/iNOS and hOC/iNOS constructs. We consider that these findings provide a basis for the development of systemically delivered PSMA/iNOS or hOC/iNOS targeting early stage and advanced prostate cancer. Copyright. (C) 2010 John Wiley & Sons, Ltd.