SALMONELLA-HIV CHIMERA AS VACCINE

长期目标是构建以沙门氏菌为基础的抗HIV疫苗 用于在志愿者中试验的菌株，由芳香族依赖性菌株组成 色葡萄伤寒（需要对氨基苯甲酸盐，在宿主组织中不可用， 因此非致病性）携带遗传信息， 给药将引起体液和细胞免疫应答， 细菌载体和由乘客DNA编码的HIV表位。 的 外源DNA将是指定氨基的短的合成寡核苷酸 鉴定为HIV蛋白抗原表位的氨基酸序列， 引起病毒中和抗体或其他保护性抗体的产生 反应 该寡核苷酸将插入克隆的鞭毛蛋白基因的一个位点， 使得它所指定的氨基酸序列暴露在鞭毛上， 表面，可接近抗体并预期具有高度免疫原性。 另一种形式的疫苗将由从鞭毛上脱落的鞭毛组成。 活疫苗细菌，浓缩和纯化后用作产品 疫苗，通过注射。 具体目标1是测试小鼠和豚鼠对以下物质的免疫应答： 沙门氏菌活疫苗的肠胃外或口服给药， 由具有寡核苷酸的克隆基因指定的鞭毛蛋白制成的鞭毛 从插入在所述第一位点的HIV的gp 160确定短氨基酸序列。 EcoRV网站，通过使用已经建成或将成为衍生产品的 将质粒pLS 408置于鞭毛蛋白阴性aroA活疫苗株中， S.都柏林 目的二是建立鞭毛蛋白的定量方法， 氨基酸插入物决定什么特征的研究继续 鞭毛的产生或鞭毛蛋白的积累或分泌 不产生鞭毛;和增加量的遗传方法 菌株产生的鞭毛蛋白的特性以及 具有两个或多个指定相同或不同表位的插入片段， 串联在单个鞭毛蛋白分子中。 目的3是构建一株S. 伤寒已经在志愿者中测试过， aro（芳香族生物合成）途径，而不是通过aroA缺失， purA缺失，以及研究purA缺失的参数变化的影响。 剂量、剂型等对口服活疫苗效力的影响。

The long-term objective is to construct Salmonella-based anti-HIV vaccine strains for trial in volunteers, consisting of an aromatic-dependent strain of S. typhi (needing p-aminobenzoate, not available in host tissues, therefore non-pathogenic) carrying genetic information such that its administration will cause humoral and cellular immune responses to both the bacterial carrier and to the HIV epitope encoded by the passenger DNA. The foreign DNA will be a short synthetic oligonucleotide specifying an amino acid sequence identified as an epitope of an HIV protein antigen able to cause production of virus-neutralizing antibody or other protective response. The oligo will be inserted in a cloned flagellin gene at a point such that the amino acid sequence it specifies is exposed at the flagellar surface, accessible to antibody and expected to be highly immunogenic. Another form of the vaccine will consist of flagella broken off from the live-vaccine bacteria, concentrated and purified for use as a product vaccine, to be given by injection. Specific Aim 1 is to test immune responses of mice and guinea pigs to parenteral or oral administration of a Salmonella live vaccine with flagella made of flagellin specified by a cloned gene with oligos determining short amino acid sequences from gp160 of HIV inserted at the EcoRV site, by use of already constructed or to be made derivatives of plasmid pLS408 placed in a flagellin-negative aroA live-vaccine strain of S. dublin. Aim 2 is development of a method for quantitation of flagellin and the investigation of what features of an amino acid insert determine continued production of flagella or the accumulation or secretion of flagellin without production of flagella; and of genetic methods of increasing amount of flagellin produced by a strain and the effect on immunogenicity, etc, of having two or more inserts, specifying the same or different epitopes, in tandem in a single flagellin molecule. Aim 3 is the construction of a replacement for a live-vaccine strain of S. typhi already tested in volunteers by one attenuated by two deletions in the aro (aromatic biosynthesis) pathway, instead of by an aroA deletion and a purA deletion, and investigation of effect of variation of parameters of dosage, presentation, etc, on efficacy of orally administered live vaccine.