氯喹增强氟尿嘧啶对结肠癌细胞的抗癌作用。

BACKGROUND: Chloroquine (CQ), the worldwide used anti-malarial drug, has recently being focused as a potential anti-cancer agent as well as a chemosensitizer when used in combination with anti-cancer drugs. It has been shown to inhibit cell growth and/or to induce cell death in various types of cancer. 5-Fluorouracil (5-FU) is the chemotherapeutic agent of first choice in colorectal cancer, but in most cases, resistance to 5-FU develops through various mechanisms. Here, we focused on the combination of CQ as a mechanism to potentiate the inhibitory effect of 5-FU on human colon cancer cells. METHODS: HT-29 cells were treated with CQ and/or 5-FU, and their proliferative ability, apoptosis and autophagy induction effects, and the affection of the cell cycle were evaluated. The proliferative ability of HT-29 was analyzed by the MTS assay. Apoptosis was quantified by flow-cytometry after double-staining of the cells with AnnexinV/PI. The cell cycle was evaluated by flow-cytometry after staining of cells with PI. Autophagy was quantified by flow-cytometry and Western blot analysis. Finally, to evaluate the fate of the cells treated with CQ and/or 5-FU, the colony formation assay was performed. RESULTS: 5-FU inhibited the proliferative activity of HT-29 cells, which was mostly dependent on the arrest of the cells to the G0/G1-phase but also partially on apoptosis induction, and the effect was potentiated by CQ pre-treatment. The potentiation of the inhibitory effect of 5-FU by CQ was dependent on the increase of p21Cip1 and p27Kip1 and the decrease of CDK2. Since CQ is reported to inhibit autophagy, the catabolic process necessary for cell survival under conditions of cell starvation or stress, which is induced by cancer cells as a protective mechanism against chemotherapeutic agents, we also analyzed the induction of autophagy in HT-29. HT-29 induced autophagy in response to 5-FU, and CQ inhibited this induction, a possible mechanism of the potentiation of the anti-cancer effect of 5-FU. CONCLUSION: Our findings suggest that the combination therapy with CQ should be a novel therapeutic modality to improve efficacy of 5-FU-based chemotherapy, possibly by inhibiting autophagy-dependent resistance to chemotherapy.

背景：氯喹（CQ）是一种广泛用于抗疟疾的药物，最近也被关注为一种潜在的抗癌药物，以及与抗癌药物联合使用时的化疗增敏剂。它已被证明可抑制各种类型癌症的细胞生长和/或诱导细胞死亡。5-氟尿嘧啶（5-FU）是结直肠癌的首选化疗药物，但在大多数情况下，通过多种机制产生对 5-FU 的耐药性。在这里，我们专注于 CQ 作为一种机制，以增强 5-FU 对人结肠癌细胞的抑制作用。

方法：用 CQ 和/或 5-FU 处理 HT-29 细胞，评估其增殖能力、凋亡和自噬诱导作用以及细胞周期的影响。通过 MTS 分析评估 HT-29 的增殖能力。通过用 AnnexinV/PI 双重染色后通过流式细胞术定量细胞凋亡。通过 PI 染色后通过流式细胞术评估细胞周期。通过流式细胞术和 Western blot 分析定量自噬。最后，通过集落形成实验评估用 CQ 和/或 5-FU 处理的细胞的命运。

结果：5-FU 抑制 HT-29 细胞的增殖活性，这主要依赖于细胞停滞在 G0/G1 期，但部分也依赖于凋亡诱导，并且 CQ 预处理可增强该作用。CQ 增强 5-FU 的抑制作用依赖于 p21Cip1 和 p27Kip1 的增加和 CDK2 的减少。由于 CQ 被报道抑制自噬，自噬是细胞在饥饿或应激条件下作为对化疗药物的保护机制所必需的分解代谢过程，我们还分析了 HT-29 中的自噬诱导。HT-29 对 5-FU 诱导自噬，CQ 抑制这种诱导，这可能是增强 5-FU 抗癌作用的机制之一。

结论：我们的研究结果表明，CQ 联合治疗可能是一种新的治疗方式，可通过抑制化疗耐药性的自噬依赖性机制来提高基于 5-FU 的化疗的疗效。