Turner Joel G, Gump Jana L, Zhang Chunchun, Cook James M, Marchion Douglas, Hazlehurst Lori, Munster Pamela, Schell Michael J, Dalton William S, Sullivan Daniel M
H. Lee Moffitt Cancer Center and Research Institute, 12902 Magnolia Dr, Tampa, FL 33612, USA.
Blood. 2006 Dec 1;108(12):3881-9. doi: 10.1182/blood-2005-10-009084. Epub 2006 Aug 17.
We investigated the role of the breast cancer resistance protein (BCRP/ABCG2) in drug resistance in multiple myeloma (MM). Human MM cell lines, and MM patient plasma cells isolated from bone marrow, were evaluated for ABCG2 mRNA expression by quantitative polymerase chain reaction (PCR) and ABCG2 protein, by Western blot analysis, immunofluorescence microscopy, and flow cytometry. ABCG2 function was determined by measuring topotecan and doxorubicin efflux using flow cytometry, in the presence and absence of the specific ABCG2 inhibitor, tryprostatin A. The methylation of the ABCG2 promoter was determined using bisulfite sequencing. We found that ABCG2 expression in myeloma cell lines increased after exposure to topotecan and doxorubicin, and was greater in logphase cells when compared with quiescent cells. Myeloma patients treated with topotecan had an increase in ABCG2 mRNA and protein expression after treatment with topotecan, and at relapse. Expression of ABCG2 is regulated, at least in part, by promoter methylation both in cell lines and in patient plasma cells. Demethylation of the promoter increased ABCG2 mRNA and protein expression. These findings suggest that ABCG2 is expressed and functional in human myeloma cells, regulated by promoter methylation, affected by cell density, up-regulated in response to chemotherapy, and may contribute to intrinsic drug resistance.
我们研究了乳腺癌耐药蛋白（BCRP/ABCG2）在多发性骨髓瘤（MM）耐药中的作用。通过定量聚合酶链反应（PCR）评估人MM细胞系以及从骨髓中分离出的MM患者浆细胞的ABCG2 mRNA表达，并通过蛋白质印迹分析、免疫荧光显微镜检查和流式细胞术评估ABCG2蛋白。通过在存在和不存在特异性ABCG2抑制剂曲古抑菌素A的情况下使用流式细胞术测量拓扑替康和阿霉素的外排来确定ABCG2的功能。使用亚硫酸氢盐测序确定ABCG2启动子的甲基化。我们发现骨髓瘤细胞系在暴露于拓扑替康和阿霉素后ABCG2表达增加，并且与静止期细胞相比，对数期细胞中的表达更高。接受拓扑替康治疗的骨髓瘤患者在接受拓扑替康治疗后以及复发时ABCG2 mRNA和蛋白表达增加。ABCG2的表达至少部分受细胞系和患者浆细胞中启动子甲基化的调节。启动子去甲基化增加了ABCG2 mRNA和蛋白表达。这些发现表明ABCG2在人骨髓瘤细胞中表达且具有功能，受启动子甲基化调节，受细胞密度影响，对化疗有上调反应，并且可能导致内在耐药性。
