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

Research Project

Project/Area Number	11671351
Research Category	Grant-in-Aid for Scientific Research (C)
Allocation Type	Single-year Grants
Section	一般
Research Field	Cerebral neurosurgery
Research Institution	Akita University
Principal Investigator	SASAJIMA Toshio Akita Univ., Sch.of Med., LECTURER, 医学部, 講師 (40235289)
Project Period (FY)	1999 – 2000
Project Status	Completed (Fiscal Year 2000)
Budget Amount *help	¥3,700,000 (Direct Cost: ¥3,700,000) Fiscal Year 2000: ¥700,000 (Direct Cost: ¥700,000) Fiscal Year 1999: ¥3,000,000 (Direct Cost: ¥3,000,000)
Keywords	malignant tumor / drug resistance / ^<99m>Tc-MIBI / MDR1mRNA expression / MTT assay / Tc^<99m>-MIBI / MDR1mRNA発見 / 増殖能
Research Abstract	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