MALARIA VACCINE DEVELOPMENT BASED ON THE RECOMBINANT SERA

本研究纯化的SE36蛋白是基于恶性疟原虫血清N-末端结构域的重组蛋白(丝氨酸重复抗原，120kd)。在本研究过程中，在GMP条件下纯化的无内毒素SE36蛋白由大阪大学微生物病研究基金会Kannonji研究所提供。针对SE36疟疾疫苗的临床开发，我们进行了以下实验，并取得了良好的效果，为开展SE36疟疾疫苗的临床试验奠定了基础。(1)检测了SE36蛋白在三只黑猩猩体内的免疫原性。SE36以氢氧化铝为佐剂。在所有动物中，SE36都能诱导出效价较高的特异性抗体。诱导的免疫球蛋白中包含了所有的免疫球蛋白亚类。(2)抗体应答持续1年以上。初次免疫2年后，单剂加强免疫可增强抗体的诱生，最高…为L两周内会有更多的报道。(3)为了解人体内与SE36交叉反应的天然获得性抗体在抗疟疾免疫中的作用，我们在恶性疟疾全流行的所罗门群岛进行了血清流行病学研究。我们检测了针对SE36蛋白和MSP-1(MSP-1)BLOCK 1-6的抗体效价以及血液寄生虫血症和疟疾症状。SE36和MSP-1、1-6、Gt；的血清阳性率分别为48%和78%。最显著的是，抗SE36抗体效价与血液寄生虫血症及症状呈显著负相关。(4)混合高滴度血清，检测血清对体外培养的寄生虫细胞增殖是否有抑制作用。高滴度混合血清对细胞增殖的抑制作用是低滴度混合血清的两倍。当高滴度血清与SE36蛋白预先孵育以中和特异性抗体时，对细胞增殖的抑制被取消50%以上。对三个独立的寄生虫菌株也观察到了类似的结果，这三个菌株拥有三种具有代表性的血清等位基因类型。这些结果表明，血清N-端区是疟疾保护性免疫的主要靶抗原之一。

SE36 protein that we have purified before this study is a recombinant protein based on the N-terminal domain of P.falciparum SERA (Serine Repeat Antigen, 120kd). In the course of this study, the endotoxin-free SE36 protein purified under GMP condition has been supplied by Kannonji Institute of Research Foundation of Microbial Diseases of Osaka University. For clinical development of SE36 malaria vaccine, we have conducted the following experiments and obtained the promising results for pursuing the clinical trials of SE36 malaria vaccine. (1) Immunogenicity of SE36 protein was examined in three chimpanzees. SE36 was administered with aluminum hydroxide as adjuvant. In all animals, SE36 induced the specific antibodies with superior titers. All of the IgG subclasses were included in the induced IgG. (2) The duration of the antibody response was more than 1 year. After 2 years from the primary immunization, single booster administration of SE36 enhanced antibody induction at the highest l … More evel within two week. (3) To see the contribution of naturally acquired antibodies cross-reactive to SE36 in human to their anti-malarial immunity, we have conducted sero-epidemiological study in Solomon Islands where is falciparum malaria holo-endemic. We have measured antibody titers against SE36 protein and block 1-6 of MSP-1 (MSP-1_<1-6>) protein as well as blood parasitemia and malaria symptoms. Sero-positive rate for SE36 and MSP-1_<1-6> were 48% and 78% respectively. Most significantly, a strong negative correlation was found between antibody titer against SE36 and blood parasitemia as well as the symptoms. (4) The high titer serum were pooled and examined whether the sera inhibits the parasite cell proliferation or not in the culture. The inhibition of the cell proliferation of the high titer pooled serum was twice as much as that observed with low titer pooled serum. When high titer serum was pre-incubated with SE36 protein to neutralize the specific antibodies, the inhibition of the cell proliferation was cancelled more than 50%. The similar results were observed against three independent parasite strains that have three representative allelic types of SERA. These results demonstrate that N-terminal domain of SERA is one of the major target antigens for the protective malaria immunity Less