Structure-guided affinity maturation of a single-chain variable fragment antibody against the Fu-bc epitope of the dengue virus envelope protein.
Structure-guided affinity maturation of a single-chain variable fragment antibody against the Fu-bc epitope of the dengue virus envelope protein.
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DOI：
10.1016/j.jbc.2022.101772
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发表时间：
2022-04
期刊：
影响因子：
--
通讯作者：
Gupta RD
中科院分区：
文献类型：
--
作者：
Sarker A;Rathore AS;Khalid MF;Gupta RD
登革热是最主要的节肢动物传播的病毒性疾病之一，全世界每年至少感染3.9亿人。尽管如此，没有有效的治疗登革热的方法，唯一可用的疫苗由于严重的不利影响已经撤回。因此，使用单克隆抗体的被动免疫疗法现在正在寻求作为治疗选择。迄今为止，已经鉴定出许多登革单克隆抗体，其中大多数是登革类型特异性的，并且其中只有少数是交叉反应性的。此外，在血清型内交叉反应的抗体是弱中和的，并且经常诱导抗体依赖性增强，这促进病毒进入和复制。因此，治疗登革热需要没有抗体依赖性增强风险的广泛中和抗体。在此，我们从抗融合环E53抗体（PDB：2 IGF）开发了单链可变片段（scFv）抗体。我们将先前预测的有利的互补决定区（CDR）突变引入编码scFv抗体的基因中用于亲和力成熟，并在体外测试所得变体对登革病毒包膜蛋白的高度保守的融合和bc表位。我们展示了在三个不同CDR中具有两个至三个取代突变的这些scFv变体中的一些具有20至200 nM范围内的亲和常数（KD）。含有D31 L、Y105 W和S227 W取代的scFv-突变体15显示出最低的亲和力常数（KD = 24 ± 7 nM），比其亲本构建体低约100倍。我们建议，scFv衍生抗体可能是一个很好的候选人，为发展一种有效和安全的免疫治疗。
Dengue is one of the most dominant arthropod-borne viral diseases, infecting at least 390 million people every year throughout the world. Despite this, there is no effective treatment against dengue, and the only available vaccine has already been withdrawn owing to the significant adverse effects. Therefore, passive immunotherapy using monoclonal antibodies is now being sought as a therapeutic option. To date, many dengue monoclonal antibodies have been identified, most of which are serotype-specific, and only a few of which are cross-reactive. Furthermore, antibodies that cross-react within serotypes are weakly neutralizing and frequently induce antibody-dependent enhancement, which promotes viral entry and replication. Therefore, broadly neutralizing antibodies with no risk of antibody-dependent enhancement are required for the treatment of dengue. Here, we developed a single-chain variable fragment (scFv) antibody from an anti-fusion loop E53 antibody (PDB: 2IGF). We introduced previously predicted favorable complementarity-determining region (CDR) mutations into the gene encoding the scFv antibody for affinity maturation, and the resultant variants were tested in vitro against the highly conserved fusion and bc epitope of the dengue virus envelope protein. We show some of these scFv variants with two to three substitution mutations in three different CDRs possess affinity constants (KD) ranging from 20 to 200 nM. The scFv-mutant15, containing D31L, Y105W, and S227W substitutions, showed the lowest affinity constant, (KD = 24 ± 7 nM), approximately 100-fold lower than its parental construct. We propose that the scFv-derivative antibody may be a good candidate for the development of an effective and safe immunotherapy.
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DOI：
10.1073/pnas.0703498104
发表时间：
2007-05-29
影响因子：
11.1
作者：
Goncalvez, Ana P.;Engle, Ronald E.;Lai, Ching-Juh
通讯作者：
Lai, Ching-Juh
影响因子：
64.8
作者：
Modis, Y;Ogata, S;Harrison, SC
通讯作者：
Harrison, SC
影响因子：
7.6
作者：
Ramadhany, Ririn;Hirai, Itaru;Kurosu, Takeshi
通讯作者：
Kurosu, Takeshi
影响因子：
64.8
作者：
Nybakken GE;Oliphant T;Johnson S;Burke S;Diamond MS;Fremont DH
通讯作者：
Fremont DH
影响因子：
3.7
作者：
Crill WD;Hughes HR;Delorey MJ;Chang GJ
通讯作者：
Chang GJ