Development of Vaccine for Preventing HTLV-I and HIV Infection and Disease Development Using Vaccinie Virus Vector

(1)我们检测了健康携带者的CTL活性与ATL或HAM患者的CTL活性是否存在差异。研究发现，HAM患者有抗Tax蛋白CTL，而健康携带者没有。利用合成的多肽进行表位定位，揭示了该蛋白氨基末端的抗原性。这些ctl受到HLAA2的限制。测量的。对T细胞的启动显示，在健康的携带者中，一些人启动了T细胞对pX蛋白（Tax， Rex和p2l^X），而另一些人则没有。这些高应答与HAM患者特有的HLA类型相关。T细胞具有CD8抗原。这些结果提示了抗税CTL与HAM之间的关系。(2)为鉴定htlv - 1的中和表位，制备了具有中和活性的抗env单克隆抗体。利用合成肽进行表位定位，发现gp46氨基酸序列191 ~ 196为单抗的结合位点。用该肽免疫家兔可产生中和抗体，预防htlv - 1感染。(3)为了构建更高效的RW载体，我们将突变引入p7.5启动子区域，并将其置于串联重复位置。进一步将其与ATI启动子结合，得到更强的启动子。当我们通过CAT基因作为报告基因的表达来衡量其功效时，合成了CAT蛋白，其数量约占细胞总蛋白的5%。(4)在ATI杂交启动子的控制下构建了含有SIV环境基因的RW。在用这种RVV免疫同种猴后，用SIV攻击它们以检验其预防感染的能力。所有三只被攻击的猴子都感染了SIV。第2个实验是在RW免疫后注射部分纯化的env蛋白作为加强剂，然后用SIV攻毒。接种疫苗的三只猴子中有一只得到了保护。

(1) We examined whether there is deference between CTL activities of healthy carriers and that of ATL or HAM patients. It has been found that the HAM patients had CTL against Tax protein whereas the healthy carriers did not. Epitope mapping using synthesized peptides revealed the antigenicity of amino terminal of Tax. These CTLs were restricted by HLAA2. Measurement of. priming of T cells revealed that among healthy carriers some had primed T cel-is to pX proteins (Tax, Rex, and p2l^X) and the other did not. These high responses correlated to the HLA types characteristic to HAM patients. The primed T cells had CD8 antigen. These results suggest the relationship between anti Tax CTL and HAM.(2) To identify the neutralization epitope of HTLV-I, anti env MAbs which had the neutralizing activity were produced. Epitope mapping using synthetic peptides revealed the gp46 amino acid sequence 191-196 as the binding site of the MAb. Immunization of the rabbits by this peptide elicited neutralizing antibody and prevented HTLV-I infection.(3) To construct more efficient RW vectors, we introduced mutations into the region of p7.5 promoter and then placed them in tandemly repeated position. Furthermore we combined them with ATI promoter to produce stronger promoter. When we measure its efficacy by expression of CAT gene as a reporter, CAT protein was synthesized whose amount was approximately 5% of that of total cell proteins.(4) We constructed the RW which harbors the SIV env gene under the control of ATI hybrid promoter. After cynomologous monkeys were immunized with this RVV, they were challenged by SIV to examine its capability of preventing infection. All three monkeys challenged were infected by SIV. In the second experiment, partially-purified env protein was injected as booster after the immunization by the RW, and then the monkeys were challenged by SIV. One of three monkeys immunized was protected.