Department of Chemistry , The University of Hong Kong , Pokfulam Road , Hong Kong , P. R. China.
Department of Medcine, Queen Mary Hospital , The University of Hong Kong , Hong Kong , P.R. China.
Anal Chem. 2018 Sep 4;90(17):10465-10471. doi: 10.1021/acs.analchem.8b02444. Epub 2018 Aug 21.
Arsenic has long been used as therapeutic agents. Understanding the mechanism of action of arsenic-based drugs enables more effective arsenic drugs to be developed. Cell cycle has been known to play a critical role for cell division and growth. Herein, we establish a methodology to evaluate the uptake of two arsenic-based drugs (ATO and ZIO-101) across the cell cycle in single leukemia cells, i.e., NB4 and HL60, using a double thymidine block combined with time-resolved inductively coupled plasma mass spectrometry (ICPMS). We show that cells absorb maximal amounts of both ATO and ZIO-101 in G2/M phase and minimum in S phase, and such variation is less apparent for ZIO-101 than ATO (NB4-G2/M:S = 2.5:1 for ATO and 1.6:1 for ZIO-101). We subsequently demonstrate that AQP9, an ATO transporter, is highly expressed in the G1 phase (50.2-46.9%) and minimum value was observed in the S phase (27.6-24.6%); whereas xCT, a ZIO-101 transporter, is maximally expressed in the G2/M phase (74.8-76.1%) and minimally expressed in the G1 phases (55.4-59.8%). Different expression levels of AQP9 and xCT are only partially accountable for the observed differences in arsenic uptake across cell cycle, indicative of the presence of other importers for both ATO and ZIO-101. Furthermore, we show that the cytotoxicity of ATO and ZIO-101 on NB4 cells is also cell cycle dependent, with the highest cytotoxicity at S + G2/M phase and the lowest at G1 + S phase. Our studies provide the first evidence on cell cycle dependent uptake and cytotoxicity of arsenic-based drugs at single cell levels, may have general implications for precise evaluation of other anticancer drugs by considering cell cycle phase.
砷一直被用作治疗剂。了解基于砷的药物的作用机制，可以开发出更有效的砷药物。细胞周期一直被认为对细胞分裂和生长起着至关重要的作用。在此，我们建立了一种方法，用于评估两种基于砷的药物（ATO 和 ZIO-101）在单个白血病细胞（即 NB4 和 HL60）中的细胞周期内的摄取情况，方法是使用双胸腺嘧啶阻断法结合时间分辨电感耦合等离子体质谱法（ICPMS）。我们发现，细胞在 G2/M 期吸收 ATO 和 ZIO-101 的量最大，在 S 期吸收的量最小，而 ZIO-101 比 ATO 的变化不太明显（NB4-G2/M：ATO 为 2.5:1，ZIO-101 为 1.6:1）。随后，我们证明 ATO 转运体 AQP9 在 G1 期高度表达（50.2-46.9%），在 S 期表达最低（27.6-24.6%）；而 ZIO-101 的转运体 xCT 在 G2/M 期表达最高（74.8-76.1%），在 G1 期表达最低（55.4-59.8%）。AQP9 和 xCT 的不同表达水平仅部分解释了砷在细胞周期内摄取的差异，表明两种 ATO 和 ZIO-101 都存在其他的摄取体。此外，我们还表明，ATO 和 ZIO-101 对 NB4 细胞的细胞毒性也与细胞周期有关，在 S+G2/M 期的细胞毒性最高，在 G1+S 期的细胞毒性最低。我们的研究首次在单细胞水平上提供了基于砷的药物摄取和细胞毒性的细胞周期依赖性证据，这可能对通过考虑细胞周期相来精确评估其他抗癌药物具有普遍意义。
