IMMUNOGENICITY AND STRUCTURES OF RHINOVIRUS:HIV CHIMERAS

描述：（改编自申请人的摘要） 为竞争性更新提出的工作是产生和识别艾滋病 来自嵌合人鼻病毒的组合文库的疫苗组分 （HRV）展示HIV-1免疫原。我们使用随机系统的技术 诱变以产生展示HIV-1表位的嵌合HRV文库 具有许多不同的长度、序列和构象。然后我们使用 免疫选择以富集具有所需抗原性特征的嵌合体。 这项赠款的前四年使我们能够证明成功， 这个系统已经导致嵌合体有效地展示HIV-1表位。 产生并表征了四个文库，其中两个展示V3环， 来自gp120的序列和两个显示来自gp41的保守ELDKWA表位的序列。 这项工作导致了三个V3环嵌合体的鉴定， 抗HIV-1中和反应是所有艾滋病报告中最有效的反应之一 疫苗系统ELDKWA嵌合体以及一些V3环嵌合体显示出 复合序列的特征在于它们能够中和 不同的HIV-1分离株初步数据显示， 初级分离物可以被抗这些嵌合体的抗血清中和。在 在另一个实验中，四只慢性HIV感染的黑猩猩被免疫 与两个V3环嵌合体中的任一个，导致增强的V3环特异性 免疫反应。在这次更新申请中，两个HRV14：HIV-1 将构建和/或表征复合序列V3环文库。 一个涉及"交叉进化枝" V3环序列，另一个涉及进化枝B 序列的此外，还将建造两个图书馆， 其将展示ELDKWA免疫原（使用不同的接头和插入 网站）。被多种抗HIV-1抗体良好中和的免疫选择嵌合体 将使用抗体对豚鼠进行免疫。诱导的抗血清 将测定HRV14：HIV-1嵌合体中和 HIV-1的实验室适应性和原始分离株。我们亦会探讨 用展示SIV Mamu-A * 01的HRV免疫恒河猴的潜力 MHC I类限制性gag CTL表位，CTPYDINQM。我们将与大卫博士合作 威斯康星州地区灵长类动物研究中心的沃特金斯博士， 在猕猴细胞中复制。如果是这样，一只或两只猕猴将被免疫， 心率变异性14。如果HRV14复制，我们将继续免疫Mamu-A * 01猕猴 与嵌合病毒，并监测SIV导向的CTL反应。嵌合 HRV：HIV系统补充了许多其他正在探索的疫苗系统， 努力研制艾滋病疫苗。HRV：HIV嵌合体有可能成为 作为活病毒疫苗组分，能够刺激体液、粘膜和， 也许是细胞介导的免疫反应。

DESCRIPTION: (Adapted from Applicant's Abstract) The continuing goal of the work proposed for the competitive renewal is to generate and identify AIDS vaccine components from combinatorial libraries of chimeric human rhinoviruses (HRVs) displaying HIV-1 immunogens. We use the technique of random systematic mutagenesis to generate libraries of chimeric HRVs that display HIV-1 epitopes with many varying lengths, sequences, and conformations. We then use immunoselection to enrich for chimeras with desired features of antigenicity. The first four years of this grant allowed us to demonstrate the success with which this system has led to chimeras that effectively display HIV-1 epitopes. Four libraries were generated and characterized, two displaying V3 loop sequences from gp120 and two displaying the conserved ELDKWA epitope from gp41. This work led to the identification of three V3 loop chimeras that elicited anti-HIV-1 neutralizing responses among the most potent reported for any AIDS vaccine system. The ELDKWA chimeras as well as some V3 loop chimeras displaying composite sequences are now being characterized for their ability to neutralize diverse isolates of HIV-1. The preliminary data indicate that a number of primary isolates can be neutralized by antisera against these chimeras. In another experiment, four chronically HIV-infected chimpanzees were immunized with either of two V3 loop chimeras, resulting in boosted V3 loop-specific immune responses. In this renewal application, two HRV14:HIV-1 composite-sequence V3 loop libraries will be constructed and/or characterized. One involves "cross-clade" V3 loop sequences and the other involves clade B sequences. In addition, two libraries will be constructed and characterized that will display the ELDKWA immunogen (using different linkers and insertion sites). Immunoselected chimeras that are neutralized well by diverse anti-HIV-1 antibodies will be used to immunize guinea pigs. Antisera elicited by HRV14:HIV-1 chimeras will be assayed for their ability to neutralize both laboratory-adapted and primary isolates of HIV-1. We will also explore the potential for immunizing rhesus macaques with HRVs displaying the SIV Mamu-A*01 MHC class I restricted gag CTL epitope, CTPYDINQM. We will work with Dr. David Watkins of the Wisconsin Regional Primate Research Center to see if HRV can replicate in macaque cells. If so, one or two macaques will be immunized with HRV14. If HRV14 replicates, we will pursue immunization of Mamu-A*01 macaques with chimeric viruses and monitor the SIV-directed CTL responses. The chimeric HRV:HIV system complements the many other vaccine systems being explored in an effort to develop an AIDS vaccine. HRV:HIV chimeras have the potential to serve as live-virus vaccine components, capable of stimulating humoral, mucosal and, perhaps, cell-mediated immune responses.