Human & mouse antibodies against influenza virus

描述(由申请人提供)：我们工作的长期目标是开发一种针对流感的疫苗策略，该策略比目前的疫苗病毒更新更有效地对抗抗原漂移变异，目前的疫苗病毒更新通常比正在传播的病毒晚一年。本供资期间拟议工作的总体目标是确定由接种疫苗或感染引起或召回的人血清中抗体所识别的抗原部位的特征，并确定多克隆抗体如何在人群中选择抗原变体。下一个授权期的具体目标是基于一个假设，即表位不具有同等的免疫原性，大部分抗体反应可能是对一小部分抗原位点的反应。在抗原漂移过程中突变的积累可能会导致特定的抗原部位变得或多或少地成为免疫优势。具体目标1是获得和鉴定从2002年起获得的与H3N1流感病毒分离株HA和NA反应的单抗(MAbs)。人类单抗将与生成的鼠单抗面板一起使用，以填补任何空白。具体目标2是绘制HA和NA上的表位图。中和表位是构象的，不被多肽模仿。在少数情况下，构象表位将通过逃逸突变体的选择和表征、竞争分析、定向突变和抗原-抗体复合体的X射线结晶学来确定。具体目的3是利用来自Aim 1的单抗与人血清进行竞争试验，确定单个表位在体内的相对免疫优势，以确定在接种疫苗或感染后，人的某些抗原位点是否占主导地位，以及在老年人或免疫抑制的受试者中是否存在差异。具体目标4是确定在拨款的5年中，随着病毒经历抗原漂移，免疫优势是否会发生变化。每年将从接种疫苗的受试者身上检测人类血清中针对漂移病毒个别抗原部位的抗体。具体目的5是确定是否可以通过制造旨在抑制特定表位的免疫原性的突变病毒或抗原来改变免疫原性。那么，对其他表位的反应会得到加强吗？这些实验的结果将显示某些抗原位置是否存在免疫优势，免疫优势是否在抗原漂移过程中发生变化，以及免疫优势是否可以系统地改变。这一知识最终可能被用来操纵疫苗抗原，以便针对更广泛的中和表位制造抗体，并更有效地对抗抗原漂移。公共卫生相关性：流感疫苗是安全的，但由于人类人群中流行的流感病毒的抗原变异，其效力各不相同。流感疫苗每年都会用新病毒更新，但由于确定新毒株正在传播并使其适应疫苗使用所需的时间，疫苗通常比病毒晚一年。这项研究的目标是确定在人类群体中选择新的流感变种的机制，并利用这一知识修改疫苗，使其对更广泛的变种病毒有效。如果和当新的大流行开始时，所获得的知识将适用于一种新的流感亚型。

DESCRIPTION (provided by applicant): The long term goal of our work is to develop a vaccine strategy for influenza that is more effective against antigenic drift variants than the current updating of vaccine viruses, which is typically a year behind the circulating viruses. The overall goal of the work proposed for this funding period is to characterize the antigenic sites recognized by antibodies in human sera that are induced or recalled by vaccination or infection and to determine how antigenic variants are selected by polyclonal antibodies in the human population. The Specific Aims for the next granting period are based on a hypothesis that epitopes are not equally immunogenic and the bulk of the antibody response may be to a small subset of antigenic sites. Accumulation of mutations during antigenic drift may cause particular antigenic sites to become less or more immunodominant. Specific Aim 1 is to obtain and characterize panels of monoclonal antibodies (mAbs) that react with HA and NA of influenza H3N2 isolates from 2002 through the 5 year grant. Human mAbs will be used with panels of mouse mAbs generated to fill in any gaps. Specific Aim 2 is to map the epitopes on the HA and NA. Neutralizing epitopes are conformational and not mimicked by peptides. Conformational epitopes will be mapped by selection and characterization of escape mutants, competition assays, directed mutagenesis and X-ray crystallography of antigen-antibody complexes in a few cases. Specific Aim 3 is to determine the relative immunodominance in vivo of individual epitopes using mAbs from Aim 1 in competition assays with human sera to determine if certain antigenic sites are dominant in people after vaccination or infection, and if there are differences in elderly or immunosuppressed subjects. Specific Aim 4 is to determine if immunodominance changes as the virus undergoes antigenic drift over the 5 years of the grant. Antibodies in human sera against individual antigenic sites of drifted viruses will be measured from subjects vaccinated each year. Specific Aim 5 is to determine if immunogenicity can be altered, by making mutant viruses or antigens that are designed to suppress immunogenicity of particular epitopes. Will response to other epitopes then be enhanced? The results of these experiments will show if there is immunodominance of certain antigenic sites, if the immunodominance changes during antigenic drift, and if immunodominance can be systematically altered. This knowledge could ultimately be used to manipulate vaccine antigens so that antibodies would be made against a wider spectrum of neutralizing epitopes and be more effective against antigenic drift. PUBLIC HEALTH RELEVANCE: Influenza vaccines are safe but have varying effectiveness due to antigenic variation of circulating influenza viruses in the human population. Influenza vaccine is updated every year with new viruses but due to the time required to determine that a new strain is spreading and to adapt it for vaccine use, the vaccine is usually a year behind the virus. The goal of this research is to determine the mechanisms by which new influenza variants are selected in the human population and to use this knowledge to modify the vaccine so that it is effective against a broader range of variant viruses. The knowledge gained will be applicable to a new subtype of influenza if and when a new pandemic begins.