Kajiwara Yoshinori, Panchabhai Sonali, Liu Diane D, Kong Maiying, Lee J Jack, Levin Victor A
Department of Neuro-Oncology, The University of Texas M. D. Anderson Cancer Center, 1515 Holcombe Boulevard, Houston, TX 77230-1402, USA.
Technol Cancer Res Treat. 2009 Apr;8(2):163-76. doi: 10.1177/153303460900800210.
The goal of our study was two-fold: (i) develop a robust 3D colony assay methodology to interrogate drug combinations using GelCount and (ii) to develop 2-drug combinations that might be useful in the clinic for the treatment of high-grade gliomas. We used three glioma cell lines (U251MG, SNB19, and LNZ308) and two adenocarcinoma cell lines (MiaPaCa and SW480) grown as colonies in a two-tiered agarose cultures. We evaluated two-drug combinations of difluoromethylornithine (DFMO), carboplatin, vorinostat (SAHA), and docetaxel. To analyze for antitumor efficacy we used GelCount to measure the area under the curve for tumor colony volumes (microm(2) x OD) in each plate. The non-linear dose-response E(max) model and the interaction index based on the Loewe additivity are applied to calculate two-drug synergy, additive, and antagonistic interactions. For glioblastoma cell lines, (i) carboplatin followed by DFMO was synergistic or additive in 2/3 cell lines, (ii) carboplatin before SAHA was synergistic in 1 cell line, (iii) carboplatin before docetaxel was synergistic in 2/3 cell lines and partially additive in the third, (iv) SAHA before docetaxel was synergistic in 1/3 cell lines, (v) docetaxel before DFMO was additive or partially active in 3/3 cell lines, and (vi) DFMO plus SAHA was inactive regardless of order. In the MiaPaCA cell line, synergy occurred when DFMO followed carboplatin and, at short exposure times, when SAHA was combined with carboplatin (regardless of order). In the SW480 cell line synergy occurred only in short exposures for carboplatin followed by docetaxel; additive and mixed partial effects were also seen with DFMO plus carboplatin or docetaxel (regardless of order), carboplatin before DFMO, carboplatin before SAHA, and docetaxel before carboplatin. In conclusion, by applying the Gelcount automated counting and sizing of colonies and the use of E(max) and Loewe models to define drug interactions, we can reliably define drug combination efficacy as a function of log dose and duration of drug exposure.
（i）开发一种强大的三维集落测定方法，以使用GelCount研究药物组合；（ii）开发可能在临床上用于治疗高级别胶质瘤的双药组合。我们使用了三种胶质瘤细胞系（U251MG、SNB19和LNZ308）和两种腺癌细胞系（MiaPaCa和SW480），它们在双层琼脂糖培养物中形成集落生长。我们评估了二氟甲基鸟氨酸（DFMO）、卡铂、伏立诺他（SAHA）和多西他赛的双药组合。为了分析抗肿瘤疗效，我们使用GelCount测量每个平板中肿瘤集落体积的曲线下面积（平方微米×光密度）。应用非线性剂量反应E(max)模型和基于Loewe加和性的相互作用指数来计算双药协同、相加和拮抗相互作用。对于胶质母细胞瘤细胞系，（i）卡铂后接DFMO在2/3的细胞系中具有协同或相加作用，（ii）卡铂在SAHA之前在1个细胞系中具有协同作用，（iii）卡铂在多西他赛之前在2/3的细胞系中具有协同作用，在第三个细胞系中部分相加，（iv）SAHA在多西他赛之前在1/3的细胞系中具有协同作用，（v）多西他赛在DFMO之前在3/3的细胞系中具有相加或部分活性，（vi）DFMO加SAHA无论顺序如何均无活性。在MiaPaCA细胞系中，当DFMO在卡铂之后以及在短暴露时间下SAHA与卡铂联合使用时（无论顺序如何）会出现协同作用。在SW480细胞系中，仅在卡铂后接多西他赛的短暴露中出现协同作用；DFMO加卡铂或多西他赛（无论顺序如何）、卡铂在DFMO之前、卡铂在SAHA之前以及多西他赛在卡铂之前也出现相加和混合的部分效应。总之，通过应用Gelcount对集落进行自动计数和测量大小，并使用E(max)和Loewe模型来定义药物相互作用，我们可以可靠地将药物组合疗效定义为对数剂量和药物暴露持续时间的函数。
