Antiangiogenic potential of camptothecin and topotecan
Antiangiogenic potential of camptothecin and topotecan
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DOI：
10.1007/s002800050997
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发表时间：
1999-11-01
影响因子：
3
通讯作者：
Daoud, SS
中科院分区：
文献类型：
--
作者：
Clements, MK;Jones, CB;Daoud, SS
目的：研究亚致死剂量喜树碱(CPT)和拓扑替康(TPT)对体外培养的人内皮细胞的抑制作用，以及与另一种抗血管生成化合物TNP-470和非特异性细胞毒药物顺铂的体内抗血管生成活性。方法：采用结晶紫法测定细胞生长抑制率，测定相对细胞数。用H-3-胸腺嘧啶核苷摄取法测定CPT和TPT对DNA合成的抑制作用。台盼蓝拒染法检测细胞存活率。流式细胞仪分析细胞对CPT的细胞周期反应。通过盘状血管生成系统(DAS)检测体内血管生成的抑制情况，将手术海绵盘置入大鼠背部皮下，评估脂质体CPT(LCPT)、TPT、TNP-470或顺铂全身治疗对盘内血管生长的抑制能力。用切片的甲苯胺蓝染色和数字图像分析来确定血管生长的数量。结果：结晶紫染色显示50 nM CPT或TPT可抑制人脐静脉内皮细胞(HUVEC)的生长，但对细胞无细胞毒性。事实证明，细胞数量并没有随着治疗的增加或减少而增加或减少，但在治疗后细胞存活超过96h时保持不变。HUVEC对~3H-胸腺嘧啶核苷的摄取早在5min就受到抑制，24 h达到最大值，并持续96h以上。细胞周期分析显示，CPT作用人脐静脉内皮细胞12h后，细胞周期停滞于S期，同时GI细胞数量减少。24 h时可见G(2)/M期细胞积聚，并伴有S期抑制。给DF大鼠隔日1 mg/kg LCPT或TPT治疗14天，可使进入椎间盘的血管生长受到约30%的抑制。这种抑制作用类似于TNP-470的抑制作用，TNP-470是一种公认的有效的血管生成抑制剂。相比之下，顺铂在抑制进入椎间盘的血管生长方面效果不佳。结论：在这项工作中，CPT和TPT以非细胞毒性的方式抑制体外培养的人内皮细胞的生长，并且这种抑制作用在药物去除后持续96h以上。我们还表明，LCPT和TPT与非特异性细胞毒剂顺铂不同，在体内椎间盘血管生成模型中，在抑制血管生成方面与TNP-470一样有效。根据这一观察结果，我们认为喜树碱除了其已证实的杀瘤活性外，还可能通过抑制血管生成而在体内具有间接的抗肿瘤作用。
Purpose: To determine the inhibitory nature of sublethal doses of camptothecin (CPT) and topotecan (TPT) treatments on normal human endothelial cells in vitro, as well as the in vivo antiangiogenic activity as compared to another antiangiogenic compound, TNP-470 and to a nonspecific cytotoxic agent, cisplatin. Methods: Growth inhibition was determined by the crystal Violet assay to measure relative cell numbers. H-3-thymidine uptake was used to determine the inhibitory effect of CPT and TPT on DNA synthesis in vitro. Cell viability was determined using trypan blue exclusion assays. Cell cycle response to CPT was determined by flow cytometric analysis of propidium iodide-stained nuclei. In vivo inhibition of angiogenesis was determined by the disc angiogenesis system (DAS), where surgical sponge discs were placed subcutaneously in the rat dorsum and the ability of systemic treatment with liposomal CPT (LCPT), TPT, TNP-470 or cisplatin to inhibit Vascular growth into the discs was evaluated. Quantitation of vascular growth was determined using toluidine blue staining of sectioned discs followed by digital image analysis. Results: Treatment with 50 nM CPT or TPT inhibited human umbilical venular endothelial cell (HUVEC) growth as shown by crystal violet staining, but was not cytotoxic to the cells. This was evidenced by the fact that cell numbers did not increase or decrease with treatment, but remained static while cells were Viable for over 96 h posttreatment. 3H-thymidine uptake in HUVEC was inhibited as early as 5 min, reached a maximum inhibition at 24 h and lasted over 96 h posttreatment. Cell cycle analysis of CPT-treated HUVEC showed arrest in S-phase at 12 h with a concurrent decrease in population of cells in GI. Accumulation of cells at the G(2)/M-phase was discernible at 24 h along with the S-phase inhibition. Treatment df rats with 1 mg/kg LCPT or TPT every other day for 14 days resulted in approximately 30% inhibition of vascular growth into the discs. This inhibition was similar to the inhibition seen with TNP-470, an established and potent angiogenic inhibitor. In contrast, cisplatin was not as effective in inhibiting vascular growth into the discs. Conclusions: In this work we showed that CPT and TPT inhibit human endothelial cell growth in vitro in a non-cytotoxic manner and that this inhibition lasts more than 96 h after drug removal. We also showed that LCPT and TPT, unlike a nonspecific cytotoxic agent, cisplatin, are as effective as TNP-470 in inhibiting angiogenic growth in the in vivo disc angiogenesis model. From this observation we propose that in addition to their proven tumoricidal activities, camptothecins may have an indirect in vivo antitumor effect mediated through the inhibition of angiogenesis.