Monitoring the functional expression of multidrug resistance gene (MDR1) and study on potentiation of antitumor agent by MDR1 inhibitors in brain tumors.

本研究旨在探讨^<99m>Tc-MIBI是否适用于阐明恶性肿瘤中MDR1抑制剂的多药耐药和预测抗肿瘤药物的增强作用。为了诱导和维持肿瘤细胞（RG2和C6胶质瘤，Walker 256癌：W256）的多药耐药，用低剂量长春新碱（VCR,1 ng/ml）连续孵育。不同浓度VCR （0.001 ~ 10 μg/ml）暴露3 d后，采用MTT法评估多药耐药情况。在1 μg/ml VCR剂量下，MTT法显示，与未给药的细胞系RG2、C6、W256相比，所有VCR耐药亚系（RG2R、C6R、W256R）的存活组分均显著增加（p<0.01）。在所有vcr耐药亚系中，RT-PCR显示MDR1 mRNA的表达高于药物初始细胞系。表达高水平MDR1 mRNA的vcr耐药亚株的Vds <99m>Tc-MIBI显著低于未接触药物的细胞（p<0.01）。Vd <99m>Tc-MIBI与MDR1 mRNA表达呈负相关（r=0.98, p<0.01）。这些发现表明，耐药的发展与^<99m>Tc-MIBI挤压增强有关。MDR - 1抑制剂（verpamil, cyclosporin, FK506）浓度为5 μM预处理3天后，与未处理的vcr -耐药亚群相比，所有vcr -耐药亚群的存活部分均显著减少。MTT试验显示MDR - 1抑制剂预处理对VCR细胞毒性有增强作用。在所有vcr耐药亚组中，MDR 1抑制剂浓度为5 μM预处理1小时后，Tc-MIBI ^<99m>的Vd均显著升高（p<0.01）。这些发现表明，MDR - 1抑制剂可以抑制mbi依赖的VCR挤压机制，这可能具有重要的临床意义，例如^<99m>Tc MIBI可以有效地监测主要作用于MDR - 1影响的系统的治疗。Tc-MIBI SPECT有望为恶性脑肿瘤患者化疗前后的多药耐药监测提供更明确的标准。少

The aim of this study is to explore whether ^<99m>Tc-MIBI is suitable to elucidate multidrug resistance and prediction of potentiation of antitumor agents by MDR1 inhibitors in malignant tumors.In order to induce and maintain multidrug resistance, tumor cells (RG2 and C6 gliomas, Walker 256 carcinoma : W256) were continuously incubated with low dose vincristine (VCR,1 ng/ml). MTT assay was performed following a 3-day exposure of various concentration of VCR (0.001-10 μg/ml) to assess the status of multidrug resistance. At a dose of 1 μg/ml of VCR, MTT assay demonstrated significant increase of surviving fractions in all VCR-resistant sublines (RG2R, C6R, W256R) as compared with those of drug-naive cell lines (RG2, C6, W256)(p<0.01). In all VCR-resistant sublines, RT-PCR revealed higher expression of MDR1 mRNA as compared with drug-naive cell lines. Vds of ^<99m>Tc-MIBI in VCR-resistant sublines expressing higher level of MDR1 mRNA was significantly lower than those of drug-naive cell l … More ines expressing lower levels of MDR1 mRNA (p<0.01). Vd of ^<99m>Tc-MIBI is negatively correlated with MDR1 mRNA expression (r=0.98, p<0.01) among drug-naive cell lines and VCR-resistant sublines. These findings indicated that the development of drug resistance was associated with enhanced ^<99m>Tc-MIBI extrusion.After 3-day pretreatment with MDR 1 inhibitors (verpamil, cyclosporin, FK506) at a concentration of 5 μM, surviving fractions of all VCR-resistant sublines significantly decreased as compared with those of non-treated VCR-resistant sublines. MTT assay revealed the enhancing effects on VCR cytotoxity following pretreatment with MDR 1 inhibitors. Vd of ^<99m>Tc-MIBI significantly increased after one-hour pretreatment with MDR 1 inhibitors at a concentration of 5 μM in all VCR-resistant sublines (p<0.01). These findings indicated that MIBI-dependent mechanisms allowing VCR extrusion were inhibited by MDR 1 inhibitors, which might have significant clinical implications such as ^<99m>Tc MIBI could be effective in monitoring therapies which act primarily on systems effected by MDR 1. ^<99m>Tc-MIBI SPECT is expected to provide more definitive criteria in monitoring multidrug resistance before and after chemotherapy in patients with malignant brain tumors. Less