Investigation of expression regulatory mechanisms of drug transporters with its possible application for circumventing anticancer drug resistance

ABCG 2/BCRP是ATP结合盒转运蛋白G家族的成员，作为外排泵发挥作用，将米托蒽醌、SN-38（伊立替康的活性代谢产物）和托泊替康等抗癌药物排除在细胞膜之外。因此，当在癌细胞中过表达时，ABCG 2引起多药耐药性。我们已经阐明，雌激素通过转录后方式显著下调雌激素受体阳性乳腺癌细胞中ABCG 2的表达，但由于其原始生理功能，雌激素在实际临床中的使用似乎受到限制。我们随后将ABCG 2 cDNA转染到表达内源性ABCG 2的乳腺癌MCF-7细胞和胃癌MKN 1和NCI-N87细胞中，在MCF-7/ABCG 2、MKN 1/ABCG 2和NCI-N87/ABCG 2细胞中，外源性ABCG 2蛋白表达被p44/p42丝裂原激活剂显著抑制，而在MCF-7/ABCG 2、MKN 1/ABCG 2和NCI-N87/ABCG 2细胞中，外源性ABCG 2蛋白表达被p44/p42丝裂原激活剂显著抑制。 ...更多信息 艾德蛋白激酶（MAPK）、PD 98059和U 0126呈剂量依赖性。这些化合物几乎完全克服了MCF-7/ABCG 2和NCI-N87/ABCG 2细胞的米托蒽醌或SN-38抗性。流式细胞仪分析显示，逆转作用是由于细胞内抗癌药物摄入增加所致。定量RT-PCR分析表明，ABCG 2 mRNA水平不受PD 98059或U 0126处理的影响。PD 98059介导的ABCG 2蛋白降解不受MG 132（一种泛素/内体抑制剂）的影响，但被巴弗洛霉素A1（一种内体抑制剂）完全阻断。这些数据表明，抑制p44/p42 MAPK通路可能会加速ABCG 2蛋白的内体降解，从而使克服ABCG 2介导的多药耐药成为可能。这些数据也提示p44/p42 MAPK抑制剂可用于建立安全有效的化疗方案。少

ABCG2/BCRP, a member of the ATP-binding cassette transporter G family, functions as an efflux pump that excludes such anticancer agents as mitoxantrone, SN-38 (an active metabolite of irinotecan), and topotecan, across cell membrane. Accordingly ABCG2 causes multidrug resistance when overexpressed in cancer cells. We have thus far clarified that estrogen markedly down-regulates ABCG2 expression in estrogen receptor-positive breast cancer cells in the post-transcriptional manner, but use of estrogen in the practical clinic seemed to be limited due to its original physiological function.We then transfected ABCG2 cDNA to breast cancer MCF-7 cells and gastric cancer MKN1 and NCI-N87 cells, which express endogenous ABCG2, in order to exploring small molecules that affects ABCG2 expression levels, and termed them MCF-7/ABCG2, MKN1/ABCG2, and NCI-N87/ABCG2 cells.Exogenous ABCG2 protein expression in MCF-7/ABCG2, MKN1/ABCG2, NCI-N87/ABCG2 cells was markedly repressed by p44/p42 mitogen-activat … More ed protein kinase (MAPK), PD98059 and U0126, in the dose-dependent manner. These compounds almost completely overcame mitoxantrone- or SN-38- resistance of MCF-7/ABCG2 and NCI-N87/ABCG2 cells. FACS analyses revealed that the reversal effects were due to increased intracellular uptake of anticancer agents. Quantitative RT-PCR analyses demonstrated that ABCG2 mRNA levels were not affected by the treatment with PD98059 or U0126. In addition, a half life of the ABCG2 protein was significantly short in the presence of PD98059 as compared with that in the control experiment.PD98059-mediated degradation of ABCG2 protein was not affected by MG132, an ubiquitin/endosome inhibitor, but was completely blocked by bafilomycin A1, an endosomal inhibitor. These data suggest that inhibition of p44/p42 MAPK pathway may accelerate endosomal degradation of ABCG2 protein and makes it possible to overcome ABCG2-mediated multidrug resistance. These data may also suggest that p44/p42 MAPK inhibitors may serve for establishment of safe and effective chemotherapeutic regimen. Less