Department of Pharmacology, Research Center for Resistant Cells, Chosun University Medical School, 375 Seosuk-dong Dong-gu, Gwangju, 501-759, Korea.
Cancer Chemother Pharmacol. 2010 May;65(6):1131-6. doi: 10.1007/s00280-009-1121-7. Epub 2009 Sep 18.
Multidrug resistance (MDR) is one of the major causes of clinical cancer chemotherapy failure. PSC833 is well known as a non-immunosuppressant cyclosporine analogue that functionally inhibits P-glycoprotein (Pgp), a product of the MDR1 gene. We investigated whether PSC833 could also alter MDR1 expression and, if so, which mitogen-activated protein kinases (MAPKs) and nuclear factor-kappaB (NF-kappaB) pathways were involved in this event.
MTT assay and flow cytometry were used for the analysis of cytotoxicity and intracellular drug accumulation, respectively. RT-PCR and Western blot assays for analysis of gene expression and electrophoretic mobility shift assays for determination of DNA-binding activity of transcription factors were used.
The doxorubicin-resistant lung cancer cell subline (SK-MES-1/DX1000), selected from SK-MES-1/WT cells, upregulated MDR1 expression, thereby showing MDR phenotypes. PSC833 sensitized SK-MES-1/DX1000 cells to doxorubicin. PSC833 (5 microM) also decreased the intracellular accumulation of fluorescent Pgp substrates such as rhodamine 123 and daunorubicin in SK-MES-1/DX1000 cells. PSC833 downregulated MDR1 mRNA and Pgp expression in a time- and concentration-dependent manner. PSC833 activated c-Jun NH2-terminal kinase (JNK)/c-Jun and enhanced AP-1 DNA-binding activity, but suppressed nuclear translocation of NF-kappaB, all of which were prevented by pretreatment with a JNK inhibitor SP600125.
These results indicate that PSC833 not only sensitizes SK-MES-1/DX1000 cells to doxorubicin by enhancing drug accumulation, but also downregulates MDR1 expression by activating JNK/c-Jun/AP-1 and suppressing NF-kappaB.
多药耐药（MDR）是临床癌症化疗失败的主要原因之一。PSC833 是一种众所周知的非免疫抑制剂环孢菌素类似物，它能有效地抑制多药耐药基因 1（MDR1）产物 P-糖蛋白（Pgp）。我们研究了 PSC833 是否也能改变 MDR1 的表达，如果能，哪些有丝分裂原激活的蛋白激酶（MAPKs）和核因子-κB（NF-κB）途径参与了这一事件。
MTT 法和流式细胞术分别用于细胞毒性和细胞内药物积累的分析。RT-PCR 和 Western blot 用于基因表达分析，电泳迁移率变动分析用于测定转录因子的 DNA 结合活性。
从 SK-MES-1/WT 细胞中选择的多柔比星耐药肺癌细胞亚系（SK-MES-1/DX1000）上调了 MDR1 的表达，从而表现出 MDR 表型。PSC833 使 SK-MES-1/DX1000 细胞对多柔比星敏感。PSC833（5 μM）也降低了 SK-MES-1/DX1000 细胞内荧光 Pgp 底物如罗丹明 123 和柔红霉素的积累。PSC833 以时间和浓度依赖的方式下调 MDR1 mRNA 和 Pgp 的表达。PSC833 激活 c-Jun NH2-末端激酶（JNK）/c-Jun 并增强 AP-1 DNA 结合活性，但抑制 NF-κB 的核转位，所有这些都被 JNK 抑制剂 SP600125 预处理所阻止。
这些结果表明，PSC833 不仅通过增强药物积累使 SK-MES-1/DX1000 细胞对多柔比星敏感，而且通过激活 JNK/c-Jun/AP-1 和抑制 NF-κB 来下调 MDR1 的表达。
