Effect of bortezomib on human neuroblastoma cell growth, apoptosis, and angiogenesis
Effect of bortezomib on human neuroblastoma cell growth, apoptosis, and angiogenesis
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DOI：
10.1093/jnci/djj309
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发表时间：
2006-08-16
期刊：
影响因子：
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通讯作者：
Ponzoni, Mirco
中科院分区：
文献类型：
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作者：
Brignole, Chiara;Marimpietri, Danilo;Ponzoni, Mirco
背景资料：硼替佐米是一种选择性和可逆的26 S蛋白酶体抑制剂，在体外和体内对几种成人癌症显示出有效的抗肿瘤活性。没有关于硼替佐米对人小儿神经母细胞瘤活性的数据。方法：检测10个神经母细胞瘤细胞系和3例患者的原代神经母细胞瘤细胞悬液对硼替佐米的敏感性。集落形成，细胞增殖，细胞周期进程和凋亡进行了评价，通过克隆形成试验和测量H-3-胸苷掺入，溴脱氧尿苷摄取，DNA片段化，磷脂酰丝氨酸暴露和碘化丙啶染色，分别。通过鸡胚绒毛尿囊膜（CAM）试验评估血管生成。使用模拟人类神经母细胞瘤生长和扩散的两种小鼠异种移植模型来检查神经母细胞瘤对硼替佐米的体内敏感性。所有统计检验均为双侧检验。结果：硼替佐米对神经母细胞瘤细胞株的增殖和集落形成具有时间和剂量依赖性抑制作用。在72小时时，导致50%生长抑制的平均硼替佐米浓度为6.1 nM（95%置信区间[CI]=0.9 - 11.3 nM）。硼替佐米处理的神经母细胞瘤细胞被阻滞在G（2）/M，并发生凋亡（用20 nM硼替佐米处理24小时的4种神经母细胞瘤细胞系中凋亡细胞的平均百分比范围为20%至35%，半胱氨酸天冬氨酸蛋白酶被激活为未处理细胞的2至5倍）。暴露于硼替佐米的原代神经母细胞瘤细胞也获得了类似的结果。硼替佐米抑制神经母细胞瘤细胞系条件培养基、神经母细胞瘤异种移植物和原发性神经母细胞瘤活检标本刺激的CAM中的血管生成（微血管面积：2.9x10（-2）mm（2），95% CI=1.8x10（-2）至3.8x10（-2）mm（2）（仅用活检标本处理的CAM）和1.3x10（-2）mm（2），在用活检标本加硼替佐米治疗的CAM中，95% CI= 1x 10（-2）至1.5x10（-2）mm（2），P= 0.024）。在两种小鼠模型中，硼替佐米治疗小鼠的存活时间在统计学上显著长于对照小鼠（假转移模型中的平均存活时间：74.2天vs 50.3天，P
Background: Bortezomib is a selective and reversible inhibitor of the 26S proteasome that shows potent antitumor activity in vitro and in vivo against several human cancers of adulthood. No data are available on bortezomib activity against human pediatric neuroblastoma. Methods: Ten neuroblastoma cell lines and suspensions of primary neuroblastoma cells from three patients were tested for sensitivity to bortezomib. Colony formation, cell proliferation, cell cycle progression, and apoptosis were evaluated by a clonogenic assay and by measuring H-3-thymidine incorporation, bromodeoxyuridine uptake, DNA fragmentation, and phosphatidylserine exposure and propidium iodide staining, respectively. Angiogenesis was assessed by the chick embryo chorioallantoic membrane (CAM) assay. Two mouse xenograft models that mimic the growth and spread of neuroblastoma in humans were used to examine in vivo sensitivity of neuroblastoma to bortezomib. All statistical tests were two-sided. Results: Bortezomib inhibited proliferation and colony formation of neuroblastoma cell lines in a time- and dose-dependent manner. The mean bortezomib concentration that caused 50% inhibition of growth was 6.1 nM (95% confidence interval [CI]=0.9 to 11.3 nM) at 72 hours. Bortezomib-treated neuroblastoma cells were arrested at G(2)/M and underwent apoptosis (mean percentage of apoptotic cells in four neuroblastoma cell lines treated with 20 nM bortezomib for 24 hours ranged from 20% to 35%, and caspases were activated by two- to fivefold with respect to untreated cells). Similar results were obtained for primary neuroblastoma cells exposed to bortezomib. Bortezomib inhibited angiogenesis in CAMs stimulated by conditioned medium from neuroblastoma cell lines, by neuroblastoma xenografts, and by primary neuroblastoma biopsy specimens (microvessel area: 2.9x10(-2) mm(2), 95% CI=1.8x10(-2) to 3.8x10(-2) mm(2) in CAMs treated with biopsy specimens alone and 1.3x10(-2) mm(2), 95% CI=1x10(-2) to 1.5x10(-2) mm(2) in CAMs treated with biopsy specimens plus bortezomib, P=.024). In both mouse models, mice treated with bortezomib lived statistically significantly longer than control mice (mean survival time in the pseudometastatic model: 74.2 versus 50.3 days, P