Balzarinia J, Degrève B, Zhu C, Durini E, Porcu L, De Clercq E, Karlsson A, Manfredini S
Rega Institute for Medical Research, K.U. Leuven, Minderbroedersstraat 10, B-3000, Leuven, Belgium.
Biochem Pharmacol. 2001 Mar 15;61(6):727-32. doi: 10.1016/s0006-2952(01)00529-9.
Introduction of a bulky lipophilic acyl entity at the 2'-OH position of both 1-beta-D-arabinofuranosylthymine (araT) and (E)-5-(2-bromovinyl)-1-beta-D-arabinofuranosyluracil (BVaraU), consistently resulted in a marked ( approximately 10-fold) increase in the inhibitory activity of these new arabinosyl nucleoside analogues for the mitochondrial thymidine kinase (TK-2)-catalysed conversion of 2 microM [methyl-(3)H]dThd to [methyl-(3)H]dTMP. The most potent derivatives were inhibitory to [methyl-(3)H]dThd phosphorylation by TK-2 within the lower micromolar concentration range. Substitution of the arabinosyl nucleoside derivatives with the acyl groups also dramatically increased the selectivity of these compounds. The inhibitory activity of araT and BVaraU to dThd phosphorylation by other related nucleoside kinases, including herpes simplex virus type 1 TK, varicella-zoster virus TK, and cytosolic TK-1, was completely annihilated upon 2'-O-acyl substitution (IC(50) > or = 1000 microM). Kinetic analysis revealed purely competitive inhibition of 2'-O-acyl-BVaraU against TK-2-catalysed thymidine phosphorylation (K(i)/K(m): 2.3). However, 2'-O-acyl-BVaraU was extremely poorly converted to the corresponding arabinosyl nucleoside 5'-monophosphate by TK-2 as revealed by [gamma-(32)P]phosphate transfer studies from [gamma-(32)P]ATP. Thus, the 2'-O-acyl derivatives of BVaraU did not behave as substrates, but rather as potent and highly selective inhibitors of TK-2. This is the first report on such a highly selective arabinosyl nucleoside inhibitor of mitochondrial TK-2, and opens perspectives for the rational design of selective mitochondrial TK-2 inhibitors.
在1-β-D-阿拉伯呋喃糖基胸腺嘧啶（araT）和（E）-5-（2-溴乙烯基）-1-β-D-阿拉伯呋喃糖基尿嘧啶（BVaraU）的2'-OH位置引入一个庞大的亲脂性酰基实体，始终导致这些新的阿拉伯糖基核苷类似物对线粒体胸苷激酶（TK-2）催化的2μM [甲基-（3）H] dThd转化为[甲基-（3）H] dTMP的抑制活性显著（约10倍）增加。最有效的衍生物在低微摩尔浓度范围内对TK-2催化的[甲基-（3）H] dThd磷酸化具有抑制作用。用酰基取代阿拉伯糖基核苷衍生物也显著提高了这些化合物的选择性。araT和BVaraU对其他相关核苷激酶（包括单纯疱疹病毒1型TK、水痘带状疱疹病毒TK和胞质TK-1）催化的dThd磷酸化的抑制活性在2'-O-酰基取代后完全消失（IC50≥1000μM）。动力学分析表明，2'-O-酰基-BVaraU对TK-2催化的胸苷磷酸化具有纯粹的竞争性抑制作用（K i /K m：2.3）。然而，如[γ-（32）P]磷酸从[γ-（32）P]ATP的转移研究所揭示的，2'-O-酰基-BVaraU被TK-2转化为相应的阿拉伯糖基核苷5'-单磷酸的效率极低。因此，BVaraU的2'-O-酰基衍生物不是作为底物，而是作为TK-2的有效且高度选择性的抑制剂。这是关于线粒体TK-2的这种高度选择性阿拉伯糖基核苷抑制剂的首次报道，并为选择性线粒体TK-2抑制剂的合理设计开辟了前景。
