Tumor-suppressive effects of neutral endopeptidase in androgen-independent prostate cancer cells.
Tumor-suppressive effects of neutral endopeptidase in androgen-independent prostate cancer cells.
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发表时间：
2001-05
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通讯作者：
Jie Dai;R. Shen;Makoto Sumitomo;Jonathan S. Goldberg;Yiping Geng;Daniel Navarro;Su Xu;J. Koutcher;Mark Garzotto;C. T. Powell;D. Nanus
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作者：
Jie Dai;R. Shen;Makoto Sumitomo;Jonathan S. Goldberg;Yiping Geng;Daniel Navarro;Su Xu;J. Koutcher;Mark Garzotto;C. T. Powell;D. Nanus
中性内肽酶(NEP)24.11在转移性、雄激素非依赖性前列腺癌(PC；C.N.Papandreou等人，NAT：MED：，4：50--57,1998)中的表达降低。为了确定细胞表面NEP过表达对雄激素非依赖性PC细胞的影响，利用可诱导的四环素调控基因表达系统将NEP基因稳定地导入并表达于产生WT-5细胞的雄激素非依赖性TSU-PR1细胞中，WT-5细胞在没有四环素的情况下培养时表达高水平的酶活性NEP蛋白。以含有相同载体但不含NEP基因且不表达NEP的TN12细胞作为对照。在去除四环素后，WT-5细胞中NEP的表达导致细胞增殖在一周内与对照细胞相比抑制了80%(P<；0.005)。注射2×10(6)WT-5细胞并饲喂多西环素(NEP抑制)的5只裸鼠中有4只于30天后在原位注射裸鼠的前列腺中形成肿瘤，所有注射TN12细胞并饲喂或不饲喂多西环素的小鼠均有肿瘤形成。相比之下，在接受WT-5细胞注射和没有接受多西环素治疗的小鼠中，只有1/5的小鼠前列腺发生了肿瘤。对NEP诱导的生长抑制机制的分析表明，NEP在WT-5细胞中的表达诱导PC细胞发生凋亡的数量增加4倍，Western印迹结果显示p21抑癌基因蛋白的表达和视网膜母细胞瘤蛋白的非磷酸化水平增加。流式细胞仪分析显示，诱导WT-5细胞NEP表达导致细胞周期G(1)期停滞。这些数据表明，NEP可以抑制PC细胞的生长和致瘤性，提示NEP具有治疗雄激素非依赖性PC的潜力。
Expression of neutral endopeptidase (NEP) 24.11 is diminished in metastatic, androgen-independent prostate cancers (PCs; C. N. Papandreou et al., NAT: MED:, 4: 50--57, 1998). To determine the effects on androgen-independent PC cells of overexpressing cell-surface NEP, an inducible tetracycline-regulatory gene expression system was used to stably introduce and express the NEP gene in androgen-independent TSU-Pr1 cells generating WT-5 cells, which expressed high levels of enzymatically active NEP protein when cultured in the absence of tetracycline. TN12 cells, which contain the identical vectors without the NEP gene and do not express NEP, were used as control. Expression of NEP in WT-5 cells after removal of tetracycline from the media resulted in a >80% inhibition in cell proliferation over a 1-week period (P < 0.005) compared with control cells. Tumor formation occurred in the prostate glands of orthotopically injected athymic mice killed at 30 days in 4 of 5 mice that were given injections of 2 x 10(6) WT-5 cells and were fed doxycycline (NEP suppressed), and in all mice that were given injections of TN12 cells and were fed with or without doxycycline. In contrast, only 1 of 5 mouse prostates developed a tumor in mice that were given injections of WT-5 cells and that did not receive doxycycline. Analysis of the mechanisms of NEP-induced growth suppression revealed that NEP expression in WT-5 cells induced a 4-fold increase in the number of PC cells undergoing apoptosis, and increased the expression of p21 tumor suppressor gene protein and the level of unphosphorylated retinoblastoma protein as determined by Western blot. Flow cytometric analysis show that induced NEP expression in WT-5 cells resulted in a G(1) cell cycle arrest. These data show that NEP can inhibit PC cell growth and tumorigenicity and suggest that NEP has potential as therapy for androgen-independent PC.