Prophylactic and therapeutic HPV16 vaccine using yeast expressing chimeric L1 and E7 protein.

宫颈癌是世界范围内妇女常见的继发性恶性肿瘤，高危型HPV（如HPV 16型）是宫颈癌最重要的致病因素。因此，开发HPV疫苗是降低宫颈癌发病率的最佳途径之一，尤其是在许多发展中国家，Koutsky等人首次证明，由酵母产生的HPV 16-L1病毒样颗粒（VLP）可以保护接受VLP胃肠外疫苗接种的女性的HPV 16感染。这表明VLP疫苗对HPV 16感染是高度有效的。然而，胃肠外VLP疫苗在许多发展中国家可能没有用，因为它非常昂贵，并且在医疗材料有限的这些国家可能难以进行注射程序。我们已经发现，含有HPV 16-L1蛋白的可食用冻干酵母疫苗，通过鼻内给予次优剂量的HPV 16-VLP增强，能够在小鼠中诱导HPV 16特异性抗体应答， ...更多信息 del系统本研究尝试构建表达L1-E7嵌合蛋白的重组酵母，以合成含E7蛋白的VLP。我们制备了三个不同大小的含有E7全长和截短的L1基因的DNA片段，并将它们转染到酵母中，得到了三株分别表达57 kD、63 KD和67 KD蛋白的酵母。由于我们成功地从产生63 KD蛋白的酵母中纯化了VLP，因此我们将该菌株用于小鼠模型系统中的可食用酵母疫苗。如果嵌合的L1-E7 VLP能诱导产生抗HPV 16的抗体，并能诱导细胞毒性T淋巴细胞杀伤HPV 16感染的细胞，则该疫苗有可能作为预防和治疗HPV 16感染的疫苗。初步实验中发现该疫苗免疫小鼠后可诱导产生HPV 16 L1抗体。现在，我们将进一步研究这一点，并在小鼠中测试细胞毒性T淋巴细胞对HPV 16-E7蛋白的反应。

High-risk HPV, such as HPV type 16 was the most important etiologic factor for cervical cancer, which is the secondary many cancer in women worldwide. Therefore, HPV vaccine development is one of the best ways to reduce the incidence of cervical cancer, especially in manu developing countries Koutsky et al. first demonstrate that HPV16-L1 virus-like particles (VLP), which was produced from yeast can protect HPV16-infection in women who undertook parenteral vaccination of the VLP. This suggests that VLP vaccine is highly effective against HPV16 infection. However, the parenteral VLP vaccine may be not useful in many developing countries, since it is very expensive and the injection procedure may be difficult to perform in such countries where medical stuffs are limited. We have found that an edible freeze-dried yeast vaccine containing HPV16-L1 protein enhanced with intranasal administration of a suboptimal dose of HPV16-VLP is able to induce HPV16-specific antibody responses in mice mo … More del system. In the present study, we tried to make recombinant yeast expressing L1-E7 chimeric protein in order to synthesize VLP containing oncoprotein E7. We made three different-sized L1-E7 DNA constracts which contained full-length of E7 and truncated L1 genes, and transfected them into yeast, and had got three strains of yeast expressing 57kD, 63KD, and 67KD proteins, respectively. Since we succeeded to purify VLP from the yeast producing 63KD protein, we used this strain for an edible yeast vaccine in the mice model system. If the chimeric L1-E7 VLP induced antibody against HP16, and incorporated E7 protein induced cytotoxic T lymphocytes killing HPV16-infected cell, this vaccine might be useful as prophylactic and therapeutic vaccine against HPV16 infection. In the preliminary experiment, we found that this vaccine induces HPV16 L1 antibody in the immunized mice. Now we are going to investigate this further, and to test the cytotoxic T lymphocyte response to HPV16-E7 protein in the mice Less