Induction of cell-mediated immune responses in mice by DNA vaccines that express hepatitis C virus NS3 mutants lacking serine protease and NTPase/RNA helicase activities.
Induction of cell-mediated immune responses in mice by DNA vaccines that express hepatitis C virus NS3 mutants lacking serine protease and NTPase/RNA helicase activities.
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DOI：
10.1371/journal.pone.0098877
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Hotta H
中科院分区：
文献类型：
--
作者：
Ratnoglik SL;Jiang DP;Aoki C;Sudarmono P;Shoji I;Deng L;Hotta H
针对病毒感染的有效治疗性疫苗必须诱导足够水平的针对靶病毒表位的细胞介导的免疫应答，并且还必须避免伴随的风险因素，例如潜在的致癌特性。丙型肝炎病毒（HCV）的非结构蛋白3（NS 3）携带多种CD 4+和CD 8 + T细胞表位，诱导强烈的HCV特异性T细胞应答，与病毒清除和急性HCV感染的消退相关。另一方面，NS 3具有丝氨酸蛋白酶和核苷三磷酸酶（NTH）/RNA解旋酶活性，其不仅在病毒生活周期中起重要作用，而且通过解除正常细胞功能而伴随干扰宿主防御机制。本研究构建了一系列表达HCV NS 3基因的DNA疫苗。为了避免NS 3的潜在危害，我们对丝氨酸蛋白酶的催化三联体（H57 A、D81 A和S139 A）和NTR/RNA解旋酶结构域（K210 N、F444 A、R461 Q和W501 A）引入突变以消除酶活性。用每种DNA疫苗候选物免疫BALB/c小鼠表达同时缺乏丝氨酸蛋白酶和NTR/解旋酶活性的NS 3突变体的pNS 3 [S139 A/K210 N]、pNS 3 [S139 A/F444 A]、pNS 3 [S139 A/R461 Q]和pNS 3 [S139 A/W501 A]诱导的T细胞免疫应答的程度与野生型NS 3和NS 3/4A复合体诱导的T细胞免疫应答的程度相当，如干扰素-γ产生和针对NS 3的细胞毒性T淋巴细胞活性所证明的。目前的研究表明，表达NS 3突变体，NS 3（S139 A/K210 N），NS 3（S139 A/F444 A），NS 3（S139 A/R461 Q）和NS 3（S139 A/W501 A）的质粒，缺乏丝氨酸蛋白酶和NTR/RNA解旋酶活性，将是安全和有效的治疗性DNA疫苗的良好候选者。
Effective therapeutic vaccines against virus infection must induce sufficient levels of cell-mediated immune responses against the target viral epitopes and also must avoid concomitant risk factors, such as potential carcinogenic properties. The nonstructural protein 3 (NS3) of hepatitis C virus (HCV) carries a variety of CD4+ and CD8+ T cell epitopes, and induces strong HCV-specific T cell responses, which are correlated with viral clearance and resolution of acute HCV infection. On the other hand, NS3 possesses serine protease and nucleoside triphosphatase (NTPase)/RNA helicase activities, which not only play important roles in viral life cycle but also concomitantly interfere with host defense mechanisms by deregulating normal cellular functions. In this study, we constructed a series of DNA vaccines that express NS3 of HCV. To avoid the potential harm of NS3, we introduced mutations to the catalytic triad of the serine protease (H57A, D81A and S139A) and the NTPase/RNA helicase domain (K210N, F444A, R461Q and W501A) to eliminate the enzymatic activities. Immunization of BALB/c mice with each of the DNA vaccine candidates (pNS3[S139A/K210N], pNS3[S139A/F444A], pNS3[S139A/R461Q] and pNS3[S139A/W501A]) that expresses an NS3 mutant lacking both serine protease and NTPase/helicase activities induced T cell immune responses to the degree comparable to that induced by the wild type NS3 and the NS3/4A complex, as demonstrated by interferon-γ production and cytotoxic T lymphocytes activities against NS3. The present study has demonstrated that plasmids expressing NS3 mutants, NS3(S139A/K210N), NS3(S139A/F444A), NS3(S139A/R461Q) and NS3(S139A/W501A), which lack both serine protease and NTPase/RNA helicase activities, would be good candidates for safe and efficient therapeutic DNA vaccines against HCV infection.
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