Vaccine Development against Infectious Diseases

 描述(申请人提供)：埃博拉、基孔肯雅、肠道病毒D68和流感等病毒的相伴危险和全球传播，使开发针对新出现疾病的有效疫苗成为当务之急。在这方面，活疫苗一直是预防感染的有效资源。然而，它们对免疫力受损的人是有害的。因此，迫切需要生产安全的无病毒疫苗。为此，我们的目标是S操纵调节免疫反应的机制，以产生可被大多数人广泛使用的安全疫苗接种方法。在这方面，我们与JoséRivera博士(UPR-Río Piedras Campus)合作的初步数据显示，一种新型超分子纳米颗粒(NP)在使用DNA疫苗方法进行体内测试时具有强大的免疫调节特性。此外，我们还表征了云芝多糖(PSP)提取物的免疫原性。我们的体外数据显示，PSP可以刺激淋巴细胞增殖并诱导TH1细胞因子谱。这一强有力的初步数据非常鼓舞人心，并激发了我们研究NP和PSP在DNA疫苗接种方法中的免疫调节作用的兴趣。就我们所知，这些新制剂中的任何一种都从未在DNA疫苗接种制度中进行过免疫调节活性测试。我们将我们的模型抗原改造成以树突状细胞(DC)为靶标，因为这些细胞是强大的抗原提呈细胞(APC)。具体地说，HIV-1外区1(OD1)是一个高度甘露糖化的基序，将HIV传递给抗原提呈细胞(APC)。为此，我们测试了一种由编码OD1的密码子优化的嵌合体组成的质粒DNA疫苗，该嵌合体通过连接序列与痘苗病毒(VACV)A27L抗原蛋白融合，以增强对APC的抗原靶向性。我们的初步数据显示，与对照组相比，用我们的质粒(POD1A27Lopt)免疫的小鼠产生了更高的A27特异性细胞和体液介导的免疫反应。基于这一信息，我们推测，在NP和PSP中形成的pOD1A27Lopt产生的免疫反应将诱导长期免疫保护。我们将通过以下特定目标来解决我们的假设：特定目标1.我们的疫苗配方在细胞介导的免疫反应、淋巴增殖和细胞因子谱中的剂量依赖的免疫调节效应。通过ELISPOT分析比较NP和PSP对小鼠脾细胞CD8T细胞和CD4T细胞释放干扰素-γ的抗原特异性频率和CD4T细胞增殖的影响。此外，我们还将研究免疫小鼠血清中的细胞因子谱。具体目标2. 我们的疫苗制剂在体液免疫反应中的免疫调节作用具有剂量依赖性。我们将比较NP和PSP在抗原特异性产生总免疫球蛋白G、IgG1(Th2型)和IgG2a(Th1型)以及产生中和抗体方面的作用。具体目的3.我们的疫苗配方在病毒复制、记忆、激活和增殖方面的免疫调节作用。我们将比较NP和PSP在增强抗原特异性保护性免疫和免疫后病毒攻击方面的效果。此外，我们还将确定该疫苗接种方案对病毒复制和诱导长期记忆、激活和增殖的影响。我们期待我们的疫苗配方为针对新出现的疾病和生物恐怖因子的合理疫苗设计提供新的方法，推动免疫治疗领域的发展。该项目将为治疗干预提供新的靶点，以诱导安全的免疫保护。

 DESCRIPTION (provided by applicant): The concomitant danger and global spread of viruses like Ebola, Chikungunya, Enterovirus D68, and Influenza, makes a priority to develop effective vaccines against emerging diseases. In this regard, live vaccines have been effective resources against infections. However, they are deleterious for immune compromised individuals. Thus, there is a critical need for the production of safe virus-free vaccines. For that reason, our goal s to manipulate the mechanisms that regulate the immune responses, in order to generate safe vaccination approaches that could be broadly used by most of the population. In this regard, our preliminary data in collaboration with Dr. José Rivera (UPR-Río Piedras Campus), shows that a novel Supramolecular Nanoparticle (NP) has strong immunomodulator properties when tested in vivo using a DNA vaccination approach. Also, we characterized the immunogenic potential of a Coriolus versicolor polysaccharide (PSP) extract. Our in vitro data shows PSP to stimulate lymphocyte proliferation and to induce a TH1 cytokine profile. This strong preliminary data is very encouraging, and drove our interest to study the immunomodulatory effects of NP and PSP in a DNA vaccination approach. To the best of our knowledge, the immunomodulator activity of any of these novel agents has never been tested in a DNA vaccination regime. We engineered our model antigen to target Dendritic Cells (DCs), as these are potent antigen-presenting cells (APCs). Specifically, HIV-1 Outer Domain-1 (OD1) is a highly-mannosylated motif that routs HIV to Antigen Presenting Cells (APCs). For that reason we tested a plasmid DNA vaccine consisting of a codon-optimized chimera coding for OD1 fused by a linker sequence to the Vaccinia Virus (VACV) A27L antigenic protein to enhance antigen targeting to APCs. Our preliminary data shows that mice immunized with our plasmid (pOD1A27Lopt) produce higher A27-specific cell- and humoral-mediated immune responses, compared to controls. Based on this information, we hypothesize that immune responses produced by pOD1A27Lopt formulated in NP and PSP, will induce long-term immune protection. We will address our hypothesis with the following specific aims: Specific Aim 1. Dose-dependent immunomodulatory effect of our vaccine formulation in cell-mediated immune responses, lymphoproliferation, and cytokine profile. We will compare the effect of NP and PSP in the antigen-specific frequency of CD8+ and CD4+ T-cells releasing IFN-γ in spleen cells by ELISPOT analysis, and CD4+ T-cell proliferation. Also, we will study the cytokine profile in serum of immunized mice. Specific Aim 2. Dose-dependent immunomodulatory effect of our vaccine formulation in humoral- mediated immune responses. We will compare the effect of NP and PSP in the antigen-specific production of total IgG, IgG1 (Th2-type), and IgG2a (Th1-type) by ELISA, and the generation of neutralizing antibodies. Specific Aim 3. Immunomodulatory effect of our vaccine formulation in viral replication, memory, activation, and proliferation. We will compare the effect of NP and PSP in the enhancement of antigen-specific protective immunity with a viral challenge after immunization. Also, we will determine the effect of this vaccination protocol on viral replication and induction of long-term memory, activation and proliferation. We expect our vaccine formulation to offer novel approaches for rational-vaccine design against emerging diseases and bioterror agents, advancing the field of immunotherapy. This project will provide new targets for therapeutic interventions to induce safe immune protection.