Preclinical Studies of Vaccines for Pandemic H1N1 Influenza

有必要对流感进行替代或辅助治疗，因为对目前使用的抗病毒药物的耐药性可能很快出现。我们与CEL-SCI公司合作，在小鼠模型中测试了一种配体表位抗原呈递系统（LEAPS）技术作为一种新的基于免疫的流感病毒感染治疗方法。flu -J-LEAPS多肽是通过将源自人MHC I类分子（J-LEAPS）的2 -微球蛋白链的结合配体与源自流感病毒NP、M或HA蛋白的15至30个氨基酸长的肽偶联而合成的。用流感- j - leaps多肽（flu - j - leaps）刺激树突状细胞，并静脉注射到感染小鼠体内。抗原特异性LEAPS刺激的dc可有效减少流感病毒在肺部的复制并提高感染动物的存活率。此外，它们增强了肺部的流感特异性T细胞反应，并通过限制过度的细胞因子反应降低了疾病的严重程度，而过度的细胞因子反应已知会导致流感病毒感染后的发病率和死亡率。我们的数据表明，流感- j - leaps脉冲dc减少了病毒在肺部的复制，提高了存活率，并调节了消除病毒的保护性免疫反应，同时防止了可能伤害宿主的过多细胞因子。这种方法有望作为流感病毒感染抗病毒治疗的辅助手段。

There is a need for alternative or adjunct therapies for influenza, because resistance to currently used antiviral drugs can emerge rapidly. In collaboration with CEL-SCI Corporation, we tested a ligand epitope antigen presentation system (LEAPS) technology as a new immune-based treatment for influenza virus infection in a mouse model. Influenza-J-LEAPS peptides were synthesized by conjugating the binding ligand derived from the β2 -microglobulin chain of the human MHC class I molecule (J-LEAPS) with 15 to 30 amino acidlong peptides derived from influenza virus NP, M, or HA proteins. DCs were stimulated with influenza-J-LEAPS peptides (influenza-J-LEAPS) and injected intravenously into infected mice. Antigen-specific LEAPS stimulated DCs were effective in reducing influenza virus replication in the lungs and enhancing survival of infected animals. Additionally, they augmented influenza-specific T cell responses in the lungs and reduced the severity of disease by limiting excessive cytokine responses, which are known to contribute to morbidity and mortality following influenza virus infection. Our data demonstrated that influenza-J-LEAPS pulsed DCs reduce virus replication in the lungs, enhance survival, and modulate the protective immune responses that eliminate the virus while preventing excessive cytokines that could injure the host. This approach shows promise as an adjunct to antiviral treatment of influenza virus infections. Rapid antigenic variation of HA, the major virion surface protein of influenza A virus, remains the principal challenge to the development of broader and more effective vaccines. Some regions of HA, such as the stem region proximal to the viral membrane, are nevertheless highly conserved across strains and among most subtypes. A fundamental question in vaccine design is the extent to which HA stem regions on the surface of the virus are accessible to broadly neutralizing antibodies. In collaboration with Sriram Subramaniams lab from NCI, we reported 3D structures derived from cryoelectron tomography of HA on intact 2009 pandemic H1N1 virions in the presence and absence of the antibody C179, which neutralizes viruses expressing a broad range of HA subtypes, including H1, H2, H5, H6, and H9. By fitting previously derived crystallographic structures of trimeric HA into the density maps, we deduced the locations of the molecular surfaces of HA involved in interaction with C179. Using computational methods to distinguish individual unliganded HA trimers from those that have bound C179 antibody, we demonstrated that ∼75% of HA trimers on the surface of the virus have C179 bound to the stem domain. Thus, despite their close packing on the viral membrane, the majority of HA trimers on intact virions are available to bind anti-stem antibodies that target conserved HA epitopes, establishing the feasibility of universal influenza vaccines that elicit such antibodies.