廈門大學夏寧邵團隊JVI發表呼吸道合胞病毒新的高效抗體研究成果

近日，Journal of Virology雜誌在線發表了廈門大學國家傳染病診斷試劑與疫苗工程技術研究中心夏寧邵教授團隊關於呼吸道合胞病毒預防性抗體研究成果。夏寧邵教授團隊與美國國立衛生研究院疫苗研究中心Barney S. Graham教授團隊合作發現了特異性針對RSV融合蛋白F的融合前構象（pre-F構象）的鼠源高效中和單抗5C4，並對5C4與1129（唯一商品化抗RSV預防性抗體藥物Palivizumab的鼠源前導單抗）進行了體內外的平行比較，發現5C4中和效價顯著優於1129。原文題為「Discovery of a Prefusion Respiratory Syncytial Virus F-Speci?c Monoclonal Antibody That Provides Greater In Vivo Protection than the Murine Precursor of Palivizumab」。

呼吸道合胞病毒（Respiratory Syncytial Virus，RSV）是導致兒童因下呼吸道感染致死的第三大病原體，每年報告病例在3000萬以上，其中超過10%的兒童需住院治療。高危新生兒的（早產兒、支氣管/氣管發育不良者、先天性心臟病及免疫缺陷兒童）RSV發病率高達50–70%。RSV至今缺乏安全有效的疫苗，上世紀70年代的備選疫苗FI-RSV在III期臨床研究中不僅未顯示出保護性效果，還增強了RSV感染所引發的相關肺部炎症。單抗藥物Palivizumab（由鼠源前導單抗1129進行人源化改造獲得）是唯一的預防性藥物。其自身效價偏低，導致了Palivizumab高昂的價格，並限制了其預防效果和應用範圍，僅在歐美等發達地區用於高危兒童產後預防。

RSV的融合蛋白F介導了病毒感染的膜融合過程，是抑制RSV病毒感染的重要靶標，F蛋白存在融合前（pre-F）及融合後（post-F）兩種不同的構象形式。本文研究者篩選獲得了一株具有高中和效價的鼠源單抗5C4。並通過重組表達的方式獲得了具有完全相同恆定區（IgG2a及IgG1）的5C4及1129，對二者在體外反應性、中和活性及體內預防效果進行了完全平行比較。結果顯示抗體1129同時與pre-F與post-F兩種構象反應，識別site II抗原表位；而5C4抗體特異性識別pre-F構象頂端的site ?抗原表位，而不與post-F構象反應。二者對於pre-F構象蛋白具有相近的反應活性。

圖 1. 5C4與1129抗體對於小鼠肺部炎症因子的抑制效果。

圖 2. 5C4抗體與1129抗體對小鼠上下呼吸道的預防效果比較。

在此條件下，細胞中和模型顯示5C4的中和效價為1129的近50倍。而小鼠感染模型評估結果顯示二者在小鼠體內的半衰期相近，但是5C4僅需要1129標準使用劑量（15mg/kg）的1/10就可以完全控制RSV感染引發的小鼠體重下降，並有效抑制肺部的IFN-γ、TNF-α、IL-5、IL-10、IP-10以及MIP-1α等炎症因子水平，在對肺部TNF-α、IL-10及IP-10等炎症因子的控制上1.5mg/kg劑量的5C4抗體甚至略優於15mg/kg劑量的1129（圖 1）。對小鼠上呼吸道及下呼吸道的病毒滴定結果均顯示出在同等劑量下5C4能夠更有效地抑制病毒的複製與擴增，與1129相比降低了病毒峰值滴度近1000倍（圖 2）。

由於兩株抗體的體外反應活性及體內半衰期相當，該研究成果表明特異性針對pre-F上site ?抗原表位的抗體能夠更為高效的預防RSV的感染，提示如何有效刺激針對該抗原表位的抗體應答對於RSV疫苗研究具有重要意義。同時該研究也顯示出5C4具有明顯的效價優勢及產業轉化潛力，為更為高效及適用範圍更廣泛的RSV特異性預防藥物的研發提供了備選分子。

ABSTRACT

Palivizumab, a humanized murine monoclonal antibody that recognizes antigenic site II on both the prefusion (pre-F) and postfusion (post-F) conformations of the RSV F glycoprotein, is the only prophylactic agent approved for RSV. However, its relatively low neutralizing potency and high cost has limited its use to a restricted population of infants at high risk of severe disease. Previously, we isolated a high potency neutralizing antibody, 5C4, that specifically recognizes antigenic site ? at the apex of the pre-F trimer. We compared–in vitro and in vivo–the potency and protective efficacy of 5C4 and the murine precursor of palivizumab, antibody 1129. Both antibodies were synthesized on identical murine backbones as either an IgG1 or IgG2a subclass and evaluated for binding to multiple F protein conformations, in vitro inhibition of RSV infection and propagation, and protective efficacy in mice. Although 1129 and 5C4 have similar pre-F binding affinities, 5C4 neutralizing activity is nearly 50-fold greater than that of 1129 in vitro. In BALB/c mice, 5C4 reduced peak RSV titers 1000-fold more than 1129 in both upper and lower respiratory tracts. These data indicate that antibodies specific for antigenic site ? are more efficacious at preventing RSV infection than antibodies specific for antigenic site II. Our data also suggest that site ?-specific antibodies may be useful for RSV prevention or therapy and support the use of pre-F as a vaccine antigen.

IMPORTANCE

There is no vaccine yet available to prevent RSV. The licensed antibody, palivizumab, which recognizes site II on both pre-F and post-F, is restricted to prophylaxis in neonates at high-risk of severe RSV disease. Recommendations for using passive immunization in the general population or for therapy in immunocompromised persons with persistent infection is limited because of cost, determined inhigh doses needed to compensate for relatively low neutralizing potency. Prior efforts to improve in vitro potency of site II antibodies did not translate to significant in vivo dose-sparing. We isolated a pre-F-specific, high-potency neutralizing antibody (5C4) that recognizes antigenic site ?, and compared its efficacy to the murine precursor of palivizumab (1129) matched for isotype and pre-F binding affinities. Our findings demonstrate that epitope specificity is an important determinant of antibody neutralizing potency, and defining mechanisms of neutralization has the potential to identify improved products for RSV prophylaxis and therapy.

來源：中國病毒學英文版、Journal of Virology

本期編輯：Tony

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