Bastin-Coyette Laurent, Smal Caroline, Cardoen Sabine, Saussoy Pascale, Van den Neste Eric, Bontemps Françoise
de Duve Institute, Université catholique de Louvain, B-1200 Brussels, Belgium.
Biochem Pharmacol. 2008 Apr 1;75(7):1451-60. doi: 10.1016/j.bcp.2007.12.007. Epub 2007 Dec 27.
2-chloroadenosine (2-CAdo) is an adenosine deaminase-resistant analogue of adenosine, widely used as an adenosine receptor agonist. This compound has been shown to induce apoptosis in several cell types either via activation of adenosine receptors or via intracellular metabolism. However, the molecular mechanisms of 2-CAdo-induced apoptosis are unclear. Here, we analyzed the effects of 2-CAdo in the leukemia cell line EHEB. 2-CAdo was found to induce apoptosis in EHEB cells, as shown by caspase-3 activation, DNA fragmentation, poly(ADP-ribose) polymerase (PARP) cleavage and phosphatidylserine exposure. Cytotoxicity of 2-CAdo was completely suppressed by 5-iodotubercidin, an adenosine kinase inhibitor, indicating that apoptosis induced by 2-CAdo was the result of its intracellular metabolism. Accordingly, we found that 2-CAdo was efficiently converted into 2-chloroATP. In parallel, a decrease of intracellular ATP concentration as well as a general inhibition of macromolecular synthesis, involving DNA, RNA and protein synthesis, was observed. Moreover, 2-CAdo induced cytochrome c release into the cytosol, indicating activation of the intrinsic pathway of apoptosis. This was found associated with a decline in Mcl-1 protein level and p53-independent. Inhibition of AMP deaminase by coformycin markedly prevented ATP depletion, and also significantly reduced 2-CAdo cytotoxicity and caspase-3 activation. In conclusion, our data show that intracellular metabolism of 2-CAdo can lead to activation of the intrinsic pathway of apoptosis and that ATP depletion, in addition to the accumulation of the triphosphate analogue, contributes to 2-CAdo-induced apoptosis.
2-氯腺苷（2-CAdo）是腺苷的一种抗腺苷脱氨酶类似物，被广泛用作腺苷受体激动剂。该化合物已被证明可通过激活腺苷受体或通过细胞内代谢在多种细胞类型中诱导凋亡。然而，2-CAdo诱导凋亡的分子机制尚不清楚。在此，我们分析了2-CAdo对白血病细胞系EHEB的影响。结果发现，2-CAdo可诱导EHEB细胞凋亡，表现为半胱天冬酶-3激活、DNA片段化、聚（ADP-核糖）聚合酶（PARP）裂解和磷脂酰丝氨酸暴露。腺苷激酶抑制剂5-碘杀结核菌素可完全抑制2-CAdo的细胞毒性，表明2-CAdo诱导的凋亡是其细胞内代谢的结果。因此，我们发现2-CAdo可有效转化为2-氯ATP。与此同时，观察到细胞内ATP浓度降低以及包括DNA、RNA和蛋白质合成在内的大分子合成普遍受到抑制。此外，2-CAdo诱导细胞色素c释放到细胞质中，表明凋亡的内在途径被激活。这与Mcl-1蛋白水平下降且与p53无关有关。助间霉素对AMP脱氨酶的抑制作用显著阻止了ATP的消耗，也显著降低了2-CAdo的细胞毒性和半胱天冬酶-3的激活。总之，我们的数据表明，2-CAdo的细胞内代谢可导致凋亡内在途径的激活，并且除了三磷酸类似物的积累外，ATP耗竭也有助于2-CAdo诱导的凋亡。
