豬流感之脂質奈米微粒信使核糖核酸疫苗研發

Abstract

豬流感 A 型病毒（IAV-S）會引起豬隻高度傳染性的呼吸道疾病與發燒。IAV-S 在豬群中呈地方性流行，部分病毒株可在人類與豬隻之間進行雙向傳播，其中又以人傳豬最為常見。防治IAV-S（特別是H1亞型）對於防止跨物種傳播及減輕養豬業的經濟損失至關重要。病毒表面的血球凝集素（HA）醣蛋白能促進病毒黏附至宿主細胞並使病毒基因組進入細胞，使其成為中和抗體與疫苗開發的主要靶點。然而，抗原漂移與抗原轉變所導致的快速突變，限制了現有疫苗對抗演變中HA抗原的功效。封裝於脂質奈米顆粒（LNP）中且編碼特定病毒抗原的mRNA疫苗，因為它們能夠快速開發並激發強大的體液與細胞免疫反應，提供了一具前景的解決方案。本研究旨在製備並評估三種不同HA序列mRNA-LNP候選疫苗，包含帶有台灣豬分離株之H1 HA、2025年美國豬傳人分離株之H1 HA以及經由分析1992 到2013年十二株台灣豬分離株之共識（consensus）H1 HA，以評估其誘導廣泛保護性免疫的潛力。小鼠模型的體內（in vivo）疫苗免疫原性分析顯示，肌肉注射接種HA-mRNA-LNP在單劑免疫後即可激發強大的同源抗原特異性IgG反應。接種兩劑後，豬傳人HA-mRNA-LNP疫苗對所有三種HA抗原誘導出最廣泛的交叉反應性體液IgG反應，而台灣豬H1 HA-mRNA-LNP疫苗則顯示出最有限的交叉反應性。相反地，鼻腔接種台灣豬HA-mRNA-LNP無法誘導出體液免疫與極低的黏膜免疫。經肌肉注射所有候選疫苗免疫後雖皆未能激發黏膜反應，重要的是，肌肉注射接種這三種候選疫苗的小鼠血清，均能成功產生針對台灣豬H1N2病毒的中和抗體。這些研究結果為利用HA-mRNA-LNP平台對抗台灣豬流感提供了初步基礎。這三種IAV-S-HA-mRNA-LNP具有作為豬流感多價疫苗候選藥物的潛力，然而未來仍需進一步研究，以納入細胞免疫評估、擴大樣本量並於豬隻模型中進行驗證。

Swine influenza A viruses (IAV-S)¬ cause highly contagious respiratory disease and fever in pigs. Endemic in swine populations, some IAV-S strains can transmit bidirectionally between humans and pigs, with human-to-swine transmission most frequently observed. Managing IAV-S, particularly the H1 subtype, is vital both to prevent interspecies transmission and to mitigate economic losses in the swine industry. The viral hemagglutinin (HA) surface glycoprotein facilitates viral attachment to host cells and eventual transfer of the viral genome, making it a primary target for neutralizing antibodies and vaccine development. However, rapid mutations driven by antigenic drift and shift limit the efficacy of current vaccines against evolving HA antigens. Lipid nanoparticle (LNP)-encapsulated mRNA vaccines encoding specific viral antigens offer a promising solution because they can be rapidly developed and elicit strong humoral and cellular immune responses. This study aimed to generate and evaluate three different HA-based mRNA-LNP vaccine candidates including H1 HA isolated from a Taiwanese pig, H1 HA from a 2025 US swine-to-human isolate, and a consensus H1 HA of twelve Taiwanese swine isolated from 1992 to 2013, to assess their potential to induced broad protective immunity. In vivo analysis of vaccine immunogenicity in a mouse model demonstrated that intramuscular (IM) immunization with HA-mRNA-LNPs elicited robust homologous antigen-specific IgG responses, typically after a single dose. Following two doses, the swine-to-human HA-mRNA-LNP vaccine induced the broadest cross-reactive humoral IgG response against all three HA antigens, whereas the Taiwan porcine H1 HA-mRNA-LNP vaccine showed the most limited cross-reactivity. Intranasal (IN) immunization with the Taiwan porcine HA-mRNA-LNP failed to induce humoral immunity and elicited very low mucosal immunity. Although IM immunization across all candidates failed to elicit a mucosal response, importantly, the serum of mice IM immunized with all three vaccine candidate vaccines successfully produced neutralizing antibodies against the Taiwan porcine H1N2 virus. These findings provide a preliminary foundation for utilizing HA-mRNA-LNP platforms against Taiwanese swine influenza. These three IAV-S-HA-mRNA-LNPs may be candidates for multivalent swine influenza vaccines, however further research incorporating cellular immunity assessments, larger sample sizes, and evaluation in swine models is necessary.