開發抗中東呼吸症候群冠狀病毒棘蛋白之單株抗體

Abstract

2012年，在沙烏地阿拉伯通報了首例新型中東呼吸症候群冠狀病毒 (MERS-CoV) 感染人的病例，並開始在全球散播。MERS-CoV的致死率高達36%，高於曾經在2002年造成全球性傳染的SARS-CoV的9%致死率。因此，為了預防疫情發生，開發出有效的診斷方法和治療藥物為當務之急。MERS-CoV棘蛋白 (Spike protein) 是病毒表面的重要抗原，棘蛋白根據其不同的功能被分S1和S2兩段次單元，其S1含有一段受體結合域 (receptor binding domain, RBD)，可介導病毒與宿主細胞的DPP4 (dipeptidyl peptidase receptor 4) 表面受體結合。本論文成功開發出可結合於MERS-CoV Spike RBD之C端的兩株單株抗體A8和G2，經比較之後發現與實驗室已開發的可結合在RBD 之N端的單株抗體 6-9具有不同之抗原決定基 (epitope)。單株抗體型別鑑定結果顯示A8、G2和6-9 皆屬於IgG1並帶有kappa 輕鏈。Sandwich ELISA 的實驗結果也揭示了G2和6-9能同時結合至RBD上。這些單株抗體將可開發成能準確診斷MERS-CoV的酵素連結免疫吸附分析法及快速免疫色譜分析試片。而這些單株抗體是否能阻斷棘蛋白與DPP4結合的能力仍需進一步測試。

In 2012, a novel coronavirus was reported in Saudi Arabia and spread out worldwide. The novel coronavirus was then officially named as Middle East respiratory syndrome coronavirus (MERS-CoV) by WHO. The mortality of MERS-CoV reaches 36%, much higher than 9.6% for SARS-CoV which caused worldwide pandemic in 2002. Thus, an effective diagnostic tool and medical treatment would be urgently needed to prevent worse pandemic. The MERS-CoV spike protein is an important surface antigen known to mediate host-receptor binding interaction and virus entry. This spike protein can be divided to two functional subunits. The S1 subunit, including receptor binding domain (RBD), can bind with dipeptidyl peptidase 4 (DPP4) on host cells. In this research, two clones of monoclonal antibodies (mAbs) A8 and G2 against MERS-CoV RBD were successfully produced. The epitope mapping experiments reveal that A8, G2, and another 6-9 mAbs previously developed in our laboratory have different binding epitopes. The results of antibody isotyping exhibit that A8, G2 and 6-9 mAbs are all IgG1 with kappa light chains. Results of sandwich ELISA (enzyme-linked immunosorbent assay) illustrated that G2 and 6-9 mAbs can simultaneously bind on RBD. G2 and 6-9 mAbs would be worth to develop into ELISA and rapid immunochromatographic strip test that accurately diagnose MERS-CoV. The capabilities of mAbs for blocking spike protein binding to DPP4 shall be further characterized.