應用類病毒顆粒技術發展廣效性流感疫苗的機制研究

流感病毒藉由其基因體的多變性持續影響人類公共衛生。疫苗是有效控制流感的重要利器，而以新技術產製新式疫苗已成為未來研發趨勢的主流。本研究利用哺乳類細胞，針對H6N1禽流感與H3N2人類季節性流感病毒，開發可提供廣效性保護力的類病毒顆粒(virus like particle, VLP)疫苗，突破病毒抗原飄移(antigenic drift)對於當前疫苗策略所造成的瓶頸；並以小鼠作為動物模式，比較此種新式疫苗與由去活化病毒(whole inactivated virus, WIV)組成的傳統疫苗，在病毒控制功效上的差異，進而探討造成保護力差異的可能機制。研究結果顯示，利用人類H6N1或H3N2流感病毒所衍生的兩種VLP疫苗，經以肌肉注射方式對小鼠進行初次與加強免疫(prime-boost immunization)之後，在不需佐劑的情形下，均可在其體內產生高力價且持久的抗體反應。這些抗體針對同源(homologous)的流感病毒具優異的中和效力。值得注意的是，與傳統去活化疫苗相比，H6N1-VLP與H3N2-VLP疫苗皆可進一步抑制異源性(heterologous)流感病毒在哺乳類細胞複製的能力，並誘發小鼠產生較強且針對病毒紅血球凝集素(hemagglutinin, HA)蛋白根部(stalk)區域的抗體反應(anti-HA stalk response)。此種抗體經後續證實與疫苗的廣效保護力有關，顯示由VLP疫苗所誘發的功能性抗體對於流感病毒的抗原飄移較不敏感，可延長保護效果。此外，進一步分析發現，誘導產生HA柄部抗體反應的機制與VLP所特有未經切割的HA0原態蛋白(uncleaved HA0 precursor)有關。因蛋白酶將HA0原態蛋白切割成HA1與HA2結構域(protein domain)為流感病毒複製所必需，這也是以去活化病毒組成的傳統疫苗無法有效誘發此類廣效性中和抗體的原因之一。這些發現除了首度找出VLP疫苗較易誘發可抵抗流感病毒變異之廣效性中和抗體的相關機制外，亦顯示使用類病毒顆粒疫苗可有效免除世衛組織每年皆須更新季節性流感疫苗抗原的不便。綜合以上結果，本研究所開發的新疫苗策略，因其結合以哺乳類細胞產製、採用類病毒顆粒抗原型式以及特有未經蛋白酶切割的HA0原態蛋白等特性，可做為未來研發廣效性通用疫苗(universal vaccine)的立論基礎，有效提高疫苗對於各類流感病毒的保護力。

關鍵字

H6N1流感病毒； H3N2流感病毒；抗原飄移；類病毒顆粒疫苗；廣效性保護抗體；蛋白酶切割

並列摘要

Influenza viruses impact public health due to their continual evolution. Vaccination is an effective countermeasure for influenza intervention. To prepare for possible threats caused by avian and human influenza viruses, we investigated the drift-tolerance of virus-like particles (VLPs) as an improved vaccine candidate. Prime-boost intramuscular immunization in mice with unadjuvanted H6N1-VLPs or H3N2-VLPs engineered using mammalian cells induced long-lasting neutralizing antibody responses against the homologous viruses. Both vaccines were able to confer superior cross-clade humoral immunity against the heterologous viruses belonging to distinct representative antigenic clusters in comparison with the cognate conventional whole-inactivated virus (WIV) vaccines, indicating the antibody repertoire induced by VLP vaccine formulations was insensitive to viral antigenic drift. Furthermore, the H6N1-VLPs and H3N2-VLPs also elicited significantly higher levels of anti-stalk antibodies contributing to a broadened neutralization effect. This advantageous effect was attributed to the uncleaved precursor of their HA proteins. These results uncover a mechanism for induction of wide-range immunity that better tolerates the evolutionary dynamics of influenza viruses and point to the possible use of a VLP as a next-generation vaccine candidate. Collectively, our strategy that combines favorable features of mammalian cell culture-based manufacture, VLP structure and the full-recombinant precursor HA trimer thus builds proof of concept of a new vaccine and may fit into the larger picture of universal/broadly cross-reactive vaccination approach to enhance cross-protection against the dynamically evolving influenza viruses.

並列關鍵字

influenza A (H6N1) virus ； influenza A(H3N2) virus ； antigenic drift ； virus-like particle vaccine ； cross-protective antibody ； proteolytic cleavage

參考文獻

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應用類病毒顆粒技術發展廣效性流感疫苗的機制研究

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