Gene-expression profiling reveals down-regulation of equilibrative nucleoside transporter 1 (ENT1) in Ara-C-resistant CCRF-CEM-derived cells
Gene-expression profiling reveals down-regulation of equilibrative nucleoside transporter 1 (ENT1) in Ara-C-resistant CCRF-CEM-derived cells
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DOI：
10.1093/jb/mvh180
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发表时间：
2004-11-01
影响因子：
2.7
通讯作者：
Yano, J
中科院分区：
文献类型：
--
作者：
Takagaki, K;Katsuma, S;Yano, J
我们使用专为分析白血病细胞而设计的内部 cDNA 微阵列研究了白血病细胞对 Ara-C 的耐药机制。我们从 CCRF-CEM（急性淋巴细胞白血病）细胞系中产生了 Ara-C 抗性细胞，并将其基因表达谱与野生型细胞进行了比较。腺苷脱氨酶 (ADA) 基因在 Ara-C 耐药细胞中高度上调，而平衡核苷转运蛋白 1 (ENT1) 和几个细胞周期相关基因则下调。在所有这些基因中，ENT1 似乎最有可能与 Ara-C 抗性相关。为了研究 ENT1 在 Ara-C 抗性细胞中的作用，我们用该基因转染了细胞。 ENT1转染的Ara-C抗性细胞在生长速率、Ara-C摄取特征和ADA表达水平方面比未转染的Ara-C抗性细胞更类似于野生型CCRF-CEM细胞。除了 Ara-C 流入受阻之外，ENT1 基因的下调预计还会导致核苷酸缺乏。因此，Ara-C 抗性细胞显示出较低的生长速率，通过 ENT1 转染可恢复生长速率。这些低生长速率也与细胞周期检查点激酶 2 的磷酸化水平相关。在这项研究中，我们确定 ENT1 的下调是造成 Ara-C 耐药性的因素，这一知识可用于设计克服耐药性的临床方案。
We have investigated the mechanism of resistance of leukemia cells to Ara-C using an in-house cDNA microarray designed for the analysis of leukemia cells. We produced Ara-C-resistant cells from the CCRF-CEM (acute lymphoblastic leukemia) cell line and compared their gene-expression profile with that of wild-type cells. The adenosine deaminase (ADA) gene was highly up-regulated in Ara-C-resistant cells, while equilibrative nucleoside transporter 1 (ENT1) and several cell-cycle-related genes were down-regulated. Of all these genes, ENT1 seemed the most likely to be relevant to Ara-C resistance. To investigate the role of ENT1 in Ara-C-resistant cells, we transfected the cells with the gene. ENT1-transfected Ara-C-resistant cells resembled wildtype CCRF-CEM cells more closely than untransfected Ara-C-resistant cells in terms of growth rate, Ara-C-uptake characteristics, and ADA expression levels. The down-regulation of the ENT1 gene is expected to result in nucleotide deficiency in addition to blockage of Ara-C influx. Accordingly, Ara-C-resistant cells showed low growth rates, which were restored by transfection with ENT1. These low growth rates were also correlated with the phosphorylation level of cell-cycle checkpoint kinase 2. In this study we identified down-regulation of ENT1 as the factor responsible for Ara-C resistance, and this knowledge may be used to devise a clinical regimen that will overcome the resistance.