Characterization of contemporary influenza B recombinant viruses harboring mutations of reduced susceptibility to baloxavir marboxil, in vitro and in mice
Characterization of contemporary influenza B recombinant viruses harboring mutations of reduced susceptibility to baloxavir marboxil, in vitro and in mice
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DOI：
10.1016/j.antiviral.2020.104807
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发表时间：
2020-07-01
影响因子：
7.6
通讯作者：
Boivin, Guy
中科院分区：
文献类型：
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作者：
Abed, Yacine;Fage, Clement;Boivin, Guy
Baloxavir marboxil（BXM）是流感病毒聚合酶酸性（PA）蛋白的强效抑制剂。然而，主要涉及甲型流感（H1N1）和甲型流感（H3 N2）感染的临床试验表明，BXM表现出低耐药性屏障。与甲型流感病毒相比，对BXM耐药的B型流感病毒变异株的记录仍然很少。我们评估了先前在甲型流感病毒中报道的138 T/M和E23 K PA替换对当代B型流感重组病毒的体外特性和毒力的影响。通过在ST 6 GalI-MDCK细胞中进行空斑减少试验，评估了乙型流感病毒B/Phuket/3073/2013重组野生型（WT）病毒和I38 T、I38 M和E23 K PA突变体对巴洛韦酸（BXA）（BXM的活性代谢产物）的敏感性。进行基于荧光素酶的微型基因组测试以确定聚合酶活性。在ST 6 GalI-MDCK细胞中评价复制动力学和遗传稳定性。在BALB/c小鼠中评价毒力。与WT相比，I38 T、I38 M和E23 K取代分别使BXA IC 50值增加12.6倍、5.5倍和2.6倍。微型基因组测定显示，与WT相比，E23 K取代的聚合酶活性损失46%，而I38 T和I38 M PA变体保留80%的活性。WT、I38 T和I38 M重组体的峰值病毒滴度相当（分别为7.95 +/- 0.5、7.45 +/- 0.25和8.11 +/- 0.28 logTCID（50）/mL），而E23 K突变体的峰值病毒滴度显著较低（6.28 +/- 0.28 logTCID（50）/mL）。P < 0.05相对于WT）。在小鼠中，WT、I38 T和I38 M重组体分别诱导60%、40%和100%的死亡率，并且在感染后第3天和第6天，三组获得相似的肺病毒滴度。总之，BXA耐药I38 T和I38 M PA突变体的适应性在当代流感B病毒中似乎未发生改变，从而对其出现进行了监测。
Baloxavir marboxil (BXM) is a potent inhibitor of the polymerase acidic (PA) protein of influenza viruses. However, clinical trials predominantly involving influenza A(H1N1) and A(H3N2) infections showed that BXM exhibited a low barrier of resistance. Contrasting with influenza A viruses, BXM-resistant influenza B variants remain poorly documented. We evaluated the impact of 138 T/M and E23K PA substitutions, previously reported in influenza A viruses, on in vitro properties and virulence of contemporary influenza B recombinant viruses. Influenza B/Phuket/3073/2013 recombinant wild-type (WT) virus and the I38T, I38M and E23K PA mutants were assessed for their susceptibility to baloxavir acid (BXA), the active metabolite of BXM, by plaque reduction assays in ST6GalI-MDCK cells. Luciferase-based minigenome tests were performed to determine polymerase activity. Replication kinetics and genetic stability were evaluated in ST6GalI-MDCK cells. Virulence was evaluated in BALB/c mice. The I38T, I38M and E23K substitutions increased BXA IC50s values by 12.6-, 5.5-, and 2.6-fold, respectively, compared to the WT. Minigenome assays revealed a 46% loss of polymerase activity for the E23K substitution vs the WT while the I38T and I38M PA variants retained 80% of activity. Peak viral titers were comparable for the WT, I38T and I38M recombinants (7.95 +/- 0.5, 7.45 +/- 0.25 and 8.11 +/- 0.28 logTCID(50)/mL), respectively, whereas it was significantly lower for the E23K mutant (6.28 +/- 0.28 logTCID(50)/mL. P < 0.05 vs the WT). In mice, the WT, I38T and I38M recombinants induced mortality rates of 60%, 40% and 100%, respectively and similar lung viral titers were obtained for the three groups at days 3 and 6 p.i. In conclusion, the fitness of BXA-resistant I38T and I38M PA mutants appears unaltered in contemporary influenza B viruses warranting surveillance for their emergence.