Shin Yong Won, Chang Ki-Hwan, Hong Gwang-Won, Yeo Sang-Gu, Jee Youngmee, Kim Jong-Hyun, Oh Myoung-Don, Cho Dong-Hyung, Kim Se-Ho
R&D Center, GC Pharma, Yongin 16924, Republic of Korea.
Division of VPD Control and NIP, Korea Centers for Disease Control and Prevention, Ministry of Health and Welfare, Osong Health Technology Administration Complex, Cheongju 28159, Republic of Korea.
J Microbiol Biotechnol. 2019 Apr 28;29(4):651-657. doi: 10.4014/jmb.1812.12024.
Although smallpox was eradicated in 1980, it is still considered a potential agent of biowarfare and bioterrorism. Smallpox has the potential for high mortality rates along with a major public health impact, eventually causing public panic and social disruption. Passive administration of neutralizing monoclonal antibodies (mAbs) is an effective intervention for various adverse reactions caused by vaccination and the unpredictable nature of emerging and bioterrorist-related infections. Currently, vaccinia immune globulin (VIG) is manufactured from vaccinia vaccine-boosted plasma; however, this production method is not ideal because of its limited availability, low specific activity, and risk of contamination with blood-borne infectious agents. To overcome the limitations of VIG production from human plasma, we isolated two human single chain variable fragments (scFvs) (SC34 and SC212) bound to vaccinia virus (VACV) from a scFv phage library constructed from the B cells of VACV vaccine-boosted volunteers. The scFvs were converted to human IgG1 (VC34 and VC212). These two anti-VACV mAbs were produced in Chinese Hamster Ovary (CHO) DG44 cells. The binding affinities of VC34 and VC212 were estimated by competition ELISA to IC values of 2 μg/mL (13.33 nM) and 22 μg/mL (146.67 nM), respectively. Only the VC212 mAb was proven to neutralize the VACV, as evidenced by the plaque reduction neutralization test (PRNT) result with a PRNT of 0.16 mg/mL (1.07 μM). This VC212 could serve as a valuable starting material for further development of VACV-neutralizing human immunoglobulin for a prophylactic measure against post-vaccination complications and for post-exposure treatment against smallpox.
虽然天花于1980年被根除，但它仍被视为生物战和生物恐怖主义的潜在媒介。天花有可能导致高死亡率，并对公众健康产生重大影响，最终引发公众恐慌和社会混乱。被动给予中和性单克隆抗体（mAb）是针对疫苗接种引起的各种不良反应以及新出现的和与生物恐怖主义相关感染的不可预测性的有效干预措施。目前，牛痘免疫球蛋白（VIG）是由接种牛痘疫苗后的血浆制成的；然而，这种生产方法并不理想，因为其供应有限、比活性低以及有被血源感染因子污染的风险。为了克服从人血浆生产VIG的局限性，我们从由接种牛痘疫苗的志愿者的B细胞构建的单链抗体噬菌体文库中分离出两个与牛痘病毒（VACV）结合的人单链可变片段（scFv）（SC34和SC212）。这些scFv被转化为人IgG1（VC34和VC212）。这两种抗VACV单克隆抗体在中国仓鼠卵巢（CHO）DG44细胞中产生。通过竞争ELISA估计，VC34和VC212的结合亲和力的IC值分别为2 μg/mL（13.33 nM）和22 μg/mL（146.67 nM）。噬斑减少中和试验（PRNT）结果显示，只有VC212单克隆抗体能中和VACV，其PRNT为0.16 mg/mL（1.07 μM）。这种VC212可作为进一步开发VACV中和性人免疫球蛋白的有价值的起始材料，用于预防疫苗接种后并发症以及天花暴露后治疗。
