Development of a mucosal vaccine to prevent Clostridium difficile infection using papilloma pseudovirus as a vector.

摘要 艰难梭菌是卫生保健机构腹泻的主要原因，有50万例感染病例， 每年。艰难梭菌感染和疾病（CDI）通常发生在抗生素治疗后， 消除了大部分肠道微生物菌群，并提供了C.很难有机会殖民 内脏如果殖民C.艰难梭菌产生毒素，许多患者发展CDI并患有症状 从腹泻到危及生命的伪膜性结肠炎CDI的发病率呈上升趋势， 很大程度上是因为抗生素的使用增加。此外，CDI在约20%的治疗患者中复发，使得这一点成为可能。 这是一种严重而昂贵的疾病，目前尚无治愈方法，也没有有效的批准疫苗。这个目标 一项建议是开发一种保护性疫苗来预防CDI及其在医疗保健中的传播 环境而少数C.艰难梭菌疫苗处于II/III期临床试验中，它们针对毒素，但将 不能阻止C.艰难梭菌在肠粘膜，也不是它的传播。我们认为最佳的 保护性疫苗将是一种对毒素产生强烈粘膜伊加抗体反应的疫苗， 一个可以防止C.很难乳头瘤病毒是小的DNA病毒，其中主要的 衣壳蛋白L1与次要衣壳蛋白L2组装成病毒样颗粒（VLP）。这些vlp 感染粘膜表面不具有感染性，但具有强免疫原性，使它们成为理想的载体， 粘膜疫苗我们已经从衣壳蛋白L1和L2开发了乳头状瘤假病毒（PsV）， 含有表达C的受体结合域（RBD）的质粒。艰难梭菌毒素A和B 初步数据显示，这些疫苗可以诱导中和粘膜伊加， C的挑战。很难在目的1中，我们将开发表达毒素A、B（来自不同菌株）的RBD的PsV。 和二元毒素我们将在小鼠和仓鼠中对它们进行免疫。我们将确定一种佐剂， 在小鼠和仓鼠的粘膜Ab应答和产生长期免疫记忆中， CDI动物模型的标准。在目的2中，我们将产生表达表面分子FliD或高表达的PsV。 分子量SLP，影响定植的分子，并确定这些疫苗是否可以诱导 抑制C.很难在目标3中，我们将给小鼠和仓鼠接种 所有PsV的混合物和用致毒C.艰难梭菌孢子从不同的菌株，以确定是否CDI 和C.困难是预防。如果成功，粘膜疫苗将产生一个强大的 中和针对毒素和表面抗原FliD和SLP的粘膜伊加应答。这种疫苗 将为改善卫生保健机构中数千名患者的卫生保健做出重大贡献， 并节省数十亿美元的医疗费用。

Abstract Clostridium difficile is a major cause of diarrhea in health care settings, with one-half million cases of infection, annually. Clostridium difficile infection and disease (CDI) usually occurs after antibiotic treatment, which eliminates much of the intestinal commensal microbiota and provides C. difficile the opportunity to colonize the gut. If the colonizing C. difficile produces toxins, many patients develop CDI and suffer from symptoms ranging from diarrhea to life-threatening pseudomembranous colitis. The incidence of CDI is on the rise, in large part because of increased use of antibiotics. Further, CDI recurs in ~20% of treated patients, making this a serious and expensive disease, for which there is no cure and no effective approved vaccine. The goal of this proposal is to develop a protective vaccine to prevent CDI and also, its transmission in the health care environment. While a few C. difficile vaccines are in Phase II/III clinical trials, they target the toxins, but will not prevent colonization of C. difficile in the intestinal mucosa, nor its transmission. We think that the optimal protective vaccine would be one that generates a strong mucosal IgA antibody response against toxins, and also one that prevents colonization of C. difficile. Papillomaviruses are small DNA viruses in which the major capsid protein, L1, assembles with the minor capsid protein, L2 into virus-like particles (VLP). These VLPs infect mucosal surfaces, are not infectious but are strongly immunogenic, making them ideal vectors for mucosal vaccines. We have developed papilloma pseudoviruses (PsV) from capsid proteins L1 and L2, that contain plasmids expressing the receptor binding domain (RBD) of the C. difficile toxins A and B, and we have preliminary data showing that these vaccines can induce neutralizing mucosal IgA and protect mice from challenge by C. difficile. In Aim 1, we will develop PsV expressing RBD of toxins A, B (from different strains) and binary toxin. We will immunize them in mice and hamsters. We will identify an adjuvant that will increase the mucosal Ab response and generate long-term immunologic memory in mice and hamsters, the gold standard for CDI animal models. In Aim 2, we will generate PsV expressing surface molecules FliD or high molecular weight SLP, molecules that will affect colonization, and determine if these vaccines can induce antibodies that inhibit colonization of C. difficile. In Aim 3, we will vaccinate mice and hamsters with a mixture of all the PsV and challenge with toxogenic C. difficile spores from different strains to determine if CDI and colonization of C. difficile are prevented. If successful, the mucosal vaccine will generate a robust neutralizing mucosal IgA response against both toxins and the surface antigens FliD and SLP. Such a vaccine would make a major contribution to improving the health care of thousands of patients in health care settings, and save billions of dollars in health care costs.