Inhibition of 6-phosphofructo-2-kinase (PFKFB3) induces autophagy as a survival mechanism.
Inhibition of 6-phosphofructo-2-kinase (PFKFB3) induces autophagy as a survival mechanism.
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DOI：
10.1186/2049-3002-2-2
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发表时间：
2014-01-23
影响因子：
5.9
通讯作者：
Telang S
中科院分区：
文献类型：
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作者：
Klarer AC;O'Neal J;Imbert-Fernandez Y;Clem A;Ellis SR;Clark J;Clem B;Chesney J;Telang S
与直接将葡萄糖分解为丙酮酸的糖酵解酶不同，6-磷酸果糖-2-激酶/果糖-2,6-二磷酸酶（PFKFBs）家族控制果糖-6-磷酸与果糖-2,6-二磷酸的转化，是糖酵解酶磷酸果糖激酶-1 （PFK-1）的关键调节因子。其中一个家族成员PFKFB3已被证明在人类癌细胞中高度表达和激活，PFKFB3抑制剂的衍生物3-（3-吡啶基）-1-（4-吡啶基）-2-丙烯-1- 1 （3PO）目前正在临床试验中开发。然而，3PO等靶向能量途径的药物的有效性受到存活途径的限制，存活途径可以通过减少ATP和营养摄取而激活。其中一种途径是细胞自我分解代谢过程，称为自噬。我们假设抑制PFKFB3诱导肿瘤细胞的功能性葡萄糖饥饿可诱导自噬作为促生存机制，而自噬抑制剂可增强PFKFB3抑制剂的抗肿瘤作用。基于LC3-II和p62蛋白表达、酸性空泡吖吖橙荧光和自噬体电镜检测，我们发现转染siRNA或3PO选择性抑制HCT-116结肠腺癌细胞的PFKFB3可导致葡萄糖摄取显著减少，同时自噬增加。由于n -乙酰半胱氨酸阻断了HCT-116细胞暴露于3PO后，酸性囊泡中LC3-I向LC3-II的转化和吖啶橙荧光的增加，因此PFKFB3抑制诱导自噬需要活性氧的增加。我们推测自噬的诱导可能会保护细胞免受3PO的促凋亡作用，并发现破坏自噬的药物，包括氯喹，增加了3PO诱导的凋亡，这是通过Annexin V和碘化丙啶双染色在HCT-116细胞和Lewis肺癌（LLC）细胞中测量的。氯喹还增强了3PO在体内对LLCs的抗生长作用，导致肿瘤内凋亡细胞增加。我们得出结论，PFKFB3抑制剂抑制葡萄糖摄取，这反过来导致自噬增加。将选择性自噬抑制剂添加到3PO及其更有效的衍生物中，可能被证明是治疗癌症的合理组合。
Unlike glycolytic enzymes that directly catabolize glucose to pyruvate, the family of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatases (PFKFBs) control the conversion of fructose-6-phosphate to and from fructose-2,6-bisphosphate, a key regulator of the glycolytic enzyme phosphofructokinase-1 (PFK-1). One family member, PFKFB3, has been shown to be highly expressed and activated in human cancer cells, and derivatives of a PFKFB3 inhibitor, 3-(3-pyridinyl)-1-(4-pyridinyl)-2-propen-1-one (3PO), are currently being developed in clinical trials. However, the effectiveness of drugs such as 3PO that target energetic pathways is limited by survival pathways that can be activated by reduced ATP and nutrient uptake. One such pathway is the process of cellular self-catabolism termed autophagy. We hypothesized that the functional glucose starvation induced by inhibition of PFKFB3 in tumor cells would induce autophagy as a pro-survival mechanism and that inhibitors of autophagy could increase the anti-tumor effects of PFKFB3 inhibitors. We found that selective inhibition of PFKFB3 with either siRNA transfection or 3PO in HCT-116 colon adenocarcinoma cells caused a marked decrease in glucose uptake simultaneously with an increase in autophagy based on LC3-II and p62 protein expression, acridine orange fluorescence of acidic vacuoles and electron microscopic detection of autophagosomes. The induction of autophagy caused by PFKFB3 inhibition required an increase in reactive oxygen species since N-acetyl-cysteine blocked both the conversion of LC3-I to LC3-II and the increase in acridine orange fluorescence in acidic vesicles after exposure of HCT-116 cells to 3PO. We speculated that the induction of autophagy might protect cells from the pro-apoptotic effects of 3PO and found that agents that disrupt autophagy, including chloroquine, increased 3PO-induced apoptosis as measured by double staining with Annexin V and propidium iodide in both HCT-116 cells and Lewis lung carcinoma (LLC) cells. Chloroquine also increased the anti-growth effect of 3PO against LLCs in vivo and resulted in an increase in apoptotic cells within the tumors. We conclude that PFKFB3 inhibitors suppress glucose uptake, which in turn causes an increase in autophagy. The addition of selective inhibitors of autophagy to 3PO and its more potent derivatives may prove useful as rational combinations for the treatment of cancer.