Photo-inactivation of Leishmania for safe and effective delivery of surrogate vac

光灭活利什曼原虫以安全有效地输送替代疫苗

• 批准号: 8386400
• 负责人: Kwang Poo Chang
• 金额: $ 23.18万
• 依托单位: ROSALIND FRANKLIN UNIV OF MEDICINE & SCI
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-25 至 2014-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8386400
• 关键词: Acquired Immunodeficiency Syndrome; Adjuvanticity; Animals; Antigen-Presenting Cells; Antigens; Automation; Biological Assay; Biotechnology; CD8B1 gene; Cell Count; Cell Survival; Cells; Cellular Immunity; Communicable Diseases; Complementary DNA; Cutaneous Leishmaniasis; Cytolysis; Data; Dendritic Cells; Disease; Effectiveness; Endotoxins; Engineering; Enzymes; Epitopes; Eukaryotic Cell; Evaluation; Exposure to; Figs - dietary; Growth; Homing; Human; Immune; Immune response; Immunity; In Situ; In Vitro; Infection; Injection of therapeutic agent; Investigation; Leishmania; Leishmaniasis; Lesion; Life; Light; Lighting; Luciferases; Lytic; Malaria; Measurement; Mediating; Methods; Microscopic; Monoclonal Antibodies; Mus; Organ; Outcome; Ovalbumin; Parasites; Particulate; Phagolysosome; Pharmaceutical Preparations; Phase; Photosensitization; Photosensitizing Agents; Post-Translational Protein Processing; Process; Production; Property; Prophylactic treatment; Proteins; Protocols documentation; Protozoa; Reagent; Resistance; Safety; Singlet Oxygen; Site; Skin; Specificity; Staging; T cell response; T-Cell Activation; T-Lymphocyte; Therapeutic; Time; Transfection; Transgenes; Transgenic Organisms; Uroporphyrins; Vaccination; Vaccines; Viral; Work; chemical genetics; cost; in vivo; luciferin; luminescence; macromolecule; macrophage; novel vaccines; photolysis; phthalocyanine; prophylactic; recombinant peptide; residence; scale up; tumor; vaccine delivery

DESCRIPTION (provided by applicant): Leishmania spp. is innately endowed with exceptionally favorable attributes for exploitation as a potential universal vaccine carrier. Transgenic biotechnology is well-developed for these eukaryotic protozoa to express foreign proteins for vaccination. In addition, adjuvanticity of Leishmania on vaccination is evident from the observation that life-long lasting immunity ensue invariably after spontaneous cure of human simple cutaneous leishmaniasis. Moreover, in natural infection, not only do Leishmania resist humoral lytic factors and parasitize exclusively the antigen-presenting cells (APC), i. e. macrophages/dendritic cells, but also live in their phagolysosomes - a desirable site for vaccine delivery. It has been our long-term objective to safely harness the attributes of Leishmania for homing vaccines to APC to enhance their immune efficacy. Toward that end, we have genetically and chemically modified Leishmania to facilitate their loading with photosensitizers (PS), i. e. uroporphyrin (URO) and phthalocyanines (Pc), thereby rendering them photosensitive to produce ROS for cytolysis to release vaccines in the ROS-resistant phagolysosomes of APC. As shown by our recent study, Leishmania can be PS-loaded so effectively that they are susceptible to photolysis on exposure to very dim light, e. g. luciferase/luciferin-mediated luminescence (semi-auto-light). Leishmania stage-specific URO accumulation and luminescence emission are currently under study by transgenic approach for automation of Leishmania photolysis intraphagolysosomally in APCs (auto-light). Significantly, vaccination of animals with uroporphyric Leishmania followed by illumination in situ (post-light) provided initial evidence for its immuno-prophylactic activity against experimental leishmaniasis. Subsequently, photo-inactivation of Pc-loaded Leishmania (Pre-light) was shown to successfully deliver a surrogate vaccine, i. e. ovalbumin (OVA) to APC, which presented this antigen effectively to activate OVA-specific T cells in vitro and in vivo. Most recently, we obtained preliminary evidence, showing that Leishmania were photolysed to completion far more consistently when sensitized with both URO and Pc than with each individually. We now propose 2 specific aims to study this double photosensitization (DP) of Leishmania in detail under pre-, post- and auto-light conditions as follows: [1] To evaluate the safety of DP-photo-inactivated Leishmania by assessing their survival in vitro in APCs and in vivo in mice; [2] To evaluate the efficacy of DP-photo-inactivated Leishmania to deliver OVA as a surrogate vaccine to APCs for presentation to elicit T cell immune responses in vitro and in vivo. Complete photolysis of Leishmania after DP is expected not only to ascertain their elimination for safe use as a vaccine carrier but also to deliver and release vaccines effectively to increase their immune efficacy. Completion of the proposed studies is thus expected to produce a final version of these carriers optimal for photodynamic immuno-therapy & -prophylaxis not only against leishmaniasis but also other infectious/non-infectious diseases. PUBLIC HEALTH RELEVANCE: The work proposed in this application is to evaluate the safety and efficacy of a combined genetic and chemical approach to photo-inactivation of parasites. The outcome of the proposed studies will help us develop prophylactic and therapeutic photomedicines against infectious and non-infectious diseases.

描述（由申请人提供）：利什曼尼亚属。天生就具有将剥削作为潜在通用疫苗载体的异常有利的属性。这些真核原生动物的转基因生物技术已经发达，以表达异物进行疫苗接种。此外，利什曼尼亚对疫苗接种的辅助性可以从以下观察到自发治疗人类简单皮肤利什曼病后总是会持续的持久免疫力。此外，在自然感染中，利什曼原虫不仅可以抵抗体液裂解因子，并仅寄生于抗原呈递细胞（APC），i。 e。巨噬细胞/树突状细胞，但也生活在其吞噬体中 - 疫苗输送的理想部位。我们的长期目标是安全利用利什曼尼亚的属性来归纳疫苗以增强其免疫功效。为此，我们具有遗传和化学修饰的利什曼原虫，以促进其使用光敏剂（PS）的负载，即。 e。泌尿原磷脂（URO）和邻苯二甲胺（PC），从而使它们具有光敏性，以产生用于细胞溶解的ROS以在APC的ROS耐药吞噬体中释放疫苗。如我们最近的研究所示，可以有效地将利什曼原虫（Leishmania）加载，以至于它们容易受到光解的影响。 g。荧光素酶/荧光素蛋白介导的发光（半自动）。 Leishmania阶段特异性的URO积累和发光发射目前正在通过转基因方法进行研究，以自动化Leishmania光溶解在APC中（自动光）中吞噬性肌液内体的自动化。值得注意的是，用尿形利什曼原虫的动物接种疫苗，然后在现场照明（灯后）提供了初始的疫苗接种 其针对实验利什曼病的免疫 - 肺活性活性的证据。随后，显示了PC负载的利什曼原虫（PRIGHT）的照片灭活已显示成功提供替代疫苗，即。 e。卵蛋白（OVA）到APC，该抗原有效地呈现了体外和体内的OVA特异性T细胞。最近，我们获得了初步证据，表明利什曼原虫（Leishmania）在用URO和PC敏感时，将Leishmania的光化量要比单独敏感得多。现在，我们提出了2个具体目标，以在前，后和自动灯条件下详细研究利什曼尼亚的双重光敏化（DP），如下所示：[1]通过评估dp-Photo灭活的利什曼尼亚的安全性，通过评估其在APC中的体外APC和Vivo中评估其生存率； [2]评估DP-Photo灭活利什曼原虫将OVA作为替代疫苗传递给APC的疗效，以表现出来，以在体外和体内引起T细胞免疫反应。 DP之后的利什曼原虫的完全光解E不仅可以确定其消除作为疫苗载体的安全使用，而且还可以有效地输送和释放疫苗以提高其免疫功效。因此，预计拟议的研究的完成将产生这些载体的最终版本，用于光动力免疫疗法和 - prophaxis，不仅针对利什曼病，而且还针对其他传染病/非传染性疾病。 公共卫生相关性：本申请中提出的工作是评估寄生寄生虫的遗传和化学方法的合并方法的安全性和功效。拟议的研究的结果将有助于我们开发针对感染性和非感染性疾病的预防性和治疗性摄影药。