Human Cytomegalovirus UL97 Kinase Is Involved in the Mechanism of Action of Methylenecyclopropane Analogs with 6-Ether and -Thioether Substitutions
Human Cytomegalovirus UL97 Kinase Is Involved in the Mechanism of Action of Methylenecyclopropane Analogs with 6-Ether and -Thioether Substitutions
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DOI：
10.1128/aac.01726-13
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发表时间：
2014-01-01
影响因子：
4.9
通讯作者：
Bowlin, Terry L.
中科院分区：
文献类型：
--
作者：
Komazin-Meredith, Gloria;Chou, Sunwen;Bowlin, Terry L.
由于良好的临床前数据和有限的更昔洛韦交叉耐药性，亚甲基环丙烷核苷 (MCPN) 类似物正在研究用于治疗人巨细胞病毒 (HCMV) 感染。在嘌呤 6 位上带有醚和硫醚官能团的单羟甲基 MCPN 除 HCMV 外，还具有针对单纯疱疹病毒 (HSV) 和水痘带状疱疹病毒 (VZV) 的抗病毒活性。检查了 HCMV UL97 激酶在这些衍生物的作用机制中的作用。当针对激酶失活的 UL97 K355M 病毒进行测试时，观察到 50% 有效浓度 (EC50) 适度增加 5 至 7 倍，而环丙韦或更昔洛韦则增加 13 至 25 倍。 HCMV在这些化合物中的两种下的连续繁殖选择了编码氨基酸取代D456N、C480R和Y617del的三个新的UL97突变。当转移到基线实验室 HCMV 毒株时，这些突变分别对所有测试的 MCPN、更昔洛韦和马里巴韦产生耐药性。然而，工程菌株也表现出与激酶失活突变体类似的严重生长缺陷和异常细胞病变效应。表达和纯化的 UL97 激酶显示新测试的 MCPN 的体外磷酸化。因此，HCMV UL97激酶参与了这些MCPN的抗病毒作用，但UL97缺陷型病毒的体外选择表明，它们针对更典型的更昔洛韦抗性、具有生长能力的UL97突变体的活性可能相对保留。
Methylenecyclopropane nucleoside (MCPN) analogs are being investigated for treatment of human cytomegalovirus (HCMV) infection because of favorable preclinical data and limited ganciclovir cross-resistance. Monohydroxymethyl MCPNs bearing ether and thioether functionalities at the purine 6 position have antiviral activity against herpes simplex virus (HSV) and varicella-zoster virus (VZV) in addition to HCMV. The role of the HCMV UL97 kinase in the mechanism of action of these derivatives was examined. When tested against a kinase-inactive UL97 K355M virus, a moderate 5- to 7-fold increase in 50% effective concentration (EC50) was observed, in comparison to a 13- to 25-fold increase for either cyclopropavir or ganciclovir. Serial propagation of HCMV under two of these compounds selected for three novel UL97 mutations encoding amino acid substitutions D456N, C480R, and Y617del. When transferred to baseline laboratory HCMV strains, these mutations individually conferred resistance to all of the tested MCPNs, ganciclovir, and maribavir. However, the engineered strains also demonstrated severe growth defects and abnormal cytopathic effects similar to the kinase-inactive mutant. Expressed and purified UL97 kinase showed in vitro phosphorylation of the newly tested MCPNs. Thus, HCMV UL97 kinase is involved in the antiviral action of these MCPNs, but the in vitro selection of UL97-defective viruses suggests that their activity against more typical ganciclovir-resistant growth-competent UL97 mutants may be relatively preserved.