Department of Pediatrics and Human Development, College of Human Medicine, Michigan State University, Grand Rapids, MI 49503, U.S.A.
Department of Medical Biology, Academic Medical Center, University of Amsterdam, 1105 AZ Amsterdam, The Netherlands.
Biochem J. 2018 Jan 31;475(2):531-545. doi: 10.1042/BCJ20170597.
The eukaryotic initiation factor 5A (eIF5A), which contributes to several crucial processes during protein translation, is the only protein that requires activation by a unique post-translational hypusine modification. eIF5A hypusination controls cell proliferation and has been linked to cancer. eIF5A hypusination requires the enzymes deoxyhypusine synthase (DHPS) and deoxyhypusine hydroxylase and uniquely depends on the polyamine (PA) spermidine as the sole substrate. Ornithine decarboxylase (ODC) is the rate-limiting enzyme in PA biosynthesis. Both ODC and PAs control cell proliferation and are frequently dysregulated in cancer. Since only spermidine can activate eIF5A, we chose the hypusine-PA nexus as a rational target to identify new drug combinations with synergistic antiproliferative effects. We show that elevated mRNA levels of the two target enzymes DHPS and ODC correlate with poor prognosis in a large cohort of neuroblastoma (NB) tumors. The DHPS inhibitor GC7 (-guanyl-1,7-diaminoheptane) and the ODC inhibitor α-difluoromethylornithine (DFMO) are target-specific and in combination induced synergistic effects in NB at concentrations that were not individually cytotoxic. Strikingly, while each drug alone at higher concentrations is known to induce p21/Rb- or p27/Rb-mediated G cell cycle arrest, we found that the drug combination induced caspase 3/7/9, but not caspase 8-mediated apoptosis, in NB cells. Hypusinated eIF5A levels and intracellular spermidine levels correlated directly with drug treatments, signifying specific drug targeting effects. This two-pronged GC7/DFMO combination approach specifically inhibits both spermidine biosynthesis and post-translational, spermidine-dependent hypusine-eIF5A activation, offering an exciting clue for improved NB drug therapy.
真核起始因子 5A（eIF5A）是唯一一种需要通过独特的翻译后氢化修饰来激活的蛋白质，它参与了蛋白质翻译过程中的几个关键步骤。eIF5A 的氢化修饰控制着细胞的增殖，并与癌症有关。eIF5A 的氢化修饰需要脱氧鸟氨酸合酶（DHPS）和脱氧鸟氨酸羟化酶的酶促作用，并且独特地依赖多胺（PA）精胺作为唯一的底物。鸟氨酸脱羧酶（ODC）是 PA 生物合成的限速酶。ODC 和 PA 都控制着细胞的增殖，并且在癌症中经常失调。由于只有精胺才能激活 eIF5A，我们选择氢化-PA 连接作为一个合理的靶点，以确定具有协同抗增殖作用的新药物组合。我们发现，在一个大型神经母细胞瘤（NB）肿瘤队列中，两个靶酶 DHPS 和 ODC 的高 mRNA 水平与预后不良相关。DHPS 抑制剂 GC7（-鸟嘌呤-1,7-二氨基庚烷）和 ODC 抑制剂 α-二氟甲基鸟氨酸（DFMO）是针对特定目标的，并且在 NB 中以不会单独产生细胞毒性的浓度联合使用时会产生协同作用。引人注目的是，虽然每种药物在较高浓度下单独使用已知会诱导 p21/Rb 或 p27/Rb 介导的 G 细胞周期停滞，但我们发现药物组合会在 NB 细胞中诱导 caspase 3/7/9，但不会诱导 caspase 8 介导的凋亡。氢化的 eIF5A 水平和细胞内精胺水平与药物处理直接相关，表明了特定的药物靶向作用。这种 GC7/DFMO 联合应用方法特异性地抑制了精胺生物合成和翻译后依赖精胺的氢化修饰，为改善 NB 药物治疗提供了一个令人兴奋的线索。
