Laboratory of Experimental Medicine, Department of Pediatrics, Faculty of Medicine, Palacký University & Faculty Hospital in Olomouc, Olomouc, Czech Republic.
Neoplasma. 2010;57(2):161-9. doi: 10.4149/neo_2010_02_161.
Bohemine and roscovitine are the most important representatives of the group of compounds structurally derived from olomoucine. Biologically they function as inhibitors of cyclin-dependent kinases (CDKs), the key regulators of cell cycle, which is often disrupted in cancer cells resulting in uncontrollable proliferation. Bohemine and roscovitine have demonstrated their cytostatic and cytotoxic in vitro and also in vivo effects. Currently the phase II clinical trials for roscovitine are underway. The aim of the study was to evaluate the potential in vitro radiosensitising effect of bohemine (BOH) and roscovitine (ROS). Clonogenic survival assay and human lung adenocarcinoma cell line A549 were used. Tested schedules were: A-pretreatment, B-concomitant application and C-posttreatment. Concentrations corresponded to IC10, IC25 and IC50 for BOH/ROS (0.1-30 microM). The radiation doses were 1, 2 and 3 Gy. Flow cytometry and western blot analysis were used to characterize cell cycle distribution, BrdU incorporation and DNA repair processes. The highest in vitro radiosensitising effect of BOH/ROS was observed for Schedule A in all tested concentrations (SER(37%) 1.46-3.20). Cell cycle analysis showed an inclination towards G0/G1 delay 48 hours posttreatment and unaltered level of apoptosis. Changes in the DNA repair processes were observed - inhibition of DNA-PK kinase, inhibition of BrdU incorporation, strong and enduring induction of p21 protein and long-lasting phosphorylation of gammaH2AX(Ser139). Certain low concentration activities of BOH/ROS in monotherapy were detected, mainly the activation of DNA-PK kinase. The results demonstrated strong in vitro radiosensitising effect of BOH/ROS that is concentration and especially schedule dependent. The strong cytostatic effect of the pretreatment schedule is mediated through the inhibition/rearrangements of DNA repair processes.
苯并咪嗪和罗司渥丁是结构上源自洛莫司汀的一组化合物的最重要代表。它们在生物学上作为细胞周期依赖性激酶 (CDK) 的抑制剂起作用，CDK 是细胞周期的关键调节剂，而在癌细胞中经常会发生细胞周期失控，导致无法控制的增殖。苯并咪嗪和罗司渥丁已在体外和体内显示出其细胞生长抑制和细胞毒性作用。目前，罗司渥丁的 II 期临床试验正在进行中。本研究旨在评估苯并咪嗪 (BOH) 和罗司渥丁 (ROS) 的体外放射增敏作用。使用克隆存活测定和人肺腺癌细胞系 A549。测试方案为：A-预处理、B-同时应用和 C-后处理。测试浓度对应于 BOH/ROS 的 IC10、IC25 和 IC50(0.1-30μM)。辐射剂量为 1、2 和 3Gy。流式细胞术和 Western blot 分析用于表征细胞周期分布、BrdU 掺入和 DNA 修复过程。在所有测试浓度下，方案 A 观察到 BOH/ROS 的体外放射增敏作用最高（SER(37%)1.46-3.20）。细胞周期分析显示，48 小时后处理时向 G0/G1 延迟的倾向和未改变的凋亡水平。观察到 DNA 修复过程的变化-抑制 DNA-PK 激酶、抑制 BrdU 掺入、强烈和持久诱导 p21 蛋白以及γH2AX(Ser139)的持久磷酸化。还检测到 BOH/ROS 在单独治疗中的某些低浓度活性，主要是 DNA-PK 激酶的激活。结果表明，BOH/ROS 具有强烈的体外放射增敏作用，这种作用取决于浓度，特别是方案。预处理方案的强烈细胞生长抑制作用是通过抑制/重组 DNA 修复过程来介导的。
