Zoli Wainer, Ricotti Luca, Tesei Anna, Ulivi Paola, Gasperi Campani Anna, Fabbri Francesco, Gunelli Roberta, Frassineti Giovanni Luca, Amadori Dino
Department of Medical Oncology, Pierantoni Hospital, Forlì, Italy.
Clin Cancer Res. 2004 Feb 15;10(4):1500-7. doi: 10.1158/1078-0432.ccr-1107-03.
The aim of the study was to evaluate the activity of epidoxorubicin (EPI) and gemcitabine (GEM) and to define the most effective schedule in human bladder cancer cells.
The study was performed on HT1376 and MCR cell lines. Cells were exposed for 1 and 24 h to drugs used in different schemes. Cytotoxic activity was evaluated by the sulforhodamine B assay, potential clinical activity was estimated by relative antitumor activity, and the type of drug interaction was assessed using the method of Chou and Talalay. Cell cycle perturbations and apoptosis were assessed by flow cytometry; BAX, BCL-2, and P53 expression was evaluated by Western blot; and DNA damage was assessed using the alkaline Comet assay.
EPI and GEM produced a cytotoxic effect in both cell lines, with 50% inhibitory concentration and relative antitumor activity values suggestive of a high clinical activity. Simultaneous treatment with EPI and GEM and the sequence GEM-->EPI caused an antagonistic interaction (combination index > 1) after both 1- and 24-h treatments. Conversely, the inverse sequence, EPI-->GEM, produced a synergistic interaction that was more pronounced in MCR cells than in HT1376 cells. The increase in DNA-damaged cells from 10% to 20% after single-drug exposure to 40-60% at the end of EPI-->GEM treatment may explain the synergistic interaction produced by the anthracycline-antimetabolite sequence.
Our findings show that the efficacy of the EPI and GEM combination is highly schedule dependent and indicate that the most active scheme is EPI followed by GEM, which is currently being validated in an ongoing intravesical Phase I-II clinical protocol.
本研究旨在评估表柔比星（EPI）和吉西他滨（GEM）的活性，并确定在人膀胱癌细胞中最有效的给药方案。
本研究在HT1376和MCR细胞系上进行。细胞分别在1小时和24小时内暴露于不同方案使用的药物。通过磺酰罗丹明B法评估细胞毒性活性，通过相对抗肿瘤活性估计潜在临床活性，并使用Chou和Talalay方法评估药物相互作用类型。通过流式细胞术评估细胞周期扰动和凋亡；通过蛋白质免疫印迹法评估BAX、BCL-2和P53表达；并使用碱性彗星试验评估DNA损伤。
EPI和GEM在两种细胞系中均产生细胞毒性作用，其50%抑制浓度和相对抗肿瘤活性值表明具有较高的临床活性。在1小时和24小时处理后，EPI和GEM同时给药以及GEM→EPI的给药顺序均导致拮抗相互作用（联合指数>1）。相反，EPI→GEM的相反给药顺序产生协同相互作用，在MCR细胞中比在HT1376细胞中更明显。单药暴露后DNA损伤细胞从10%增加到20%，而在EPI→GEM治疗结束时增加到40%-60%，这可能解释了蒽环类药物-抗代谢物给药顺序产生的协同相互作用。
我们的研究结果表明，EPI和GEM联合使用的疗效高度依赖给药方案，并表明最有效的方案是先给予EPI后给予GEM，目前正在一项正在进行的膀胱内I-II期临床方案中进行验证。
