Antiviral lectins as microbicides

作为杀微生物剂的抗病毒凝集素

基本信息

批准号：
7795249
负责人：
KENNETH E PALMER
金额：
$ 52.59万
依托单位：
UNIVERSITY OF LOUISVILLE
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-04-15 至 2012-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7795249
关键词：
Agriculture Animals Antibodies Antiviral Agents Attention Binding Binding Sites Biological Biological Assay Cells Chlamydia trachomatis Clinical Collaborations Complex Data Data Set Development Dose Drug Kinetics Epithelial Epithelium Evaluation Exhibits Fermentation Gel Genetic Goals HIV HIV-1 Hemophilus Histology Histopathology Homologous Gene Human Human Herpesvirus 2 Human Papillomavirus Immune Immune response In Vitro Infection Inflammation Inflammatory Response Intravaginal Administration Investigation Lectin Leukocyte Chemotaxis Leukocytes Mannose Measurement Measures Modeling Molecular Mus Mutate Oryctolagus cuniculus Peptides Pharmaceutical Preparations Pharmacology and Toxicology Phenotype Placebo Control Plant Leaves Play Predisposition Prevention Production Proteins Recombinant Proteins Recombinants Recruitment Activity Research Resistance Risk Factors Role Safety Series Sexual Transmission Sexually Transmitted Diseases Simplexvirus Site-Directed Mutagenesis Solutions Structure Sum Surface System Testing Tissues Tobacco Toxic effect Toxicology Vaccines Vagina Vertebrates Virus base chemokine cost cytokine genital herpes hydroxyethylcellulose immunogenic immunogenicity in vivo inhibitor/antagonist insight irritation lymphocyte proliferation microbicide molecular marker mutant pathogen practical application prevent public health relevance rectal microbicide research study resistance mechanism response sugar transmission process vaginal microbicide

项目摘要

DESCRIPTION (provided by applicant): For practical application, microbicides must be available in massive amounts, and must be produced cost- effectively. These requirements might eliminate recombinant proteins from consideration as costs of production in cell-based fermentation systems are prohibitively high. We developed a practical solution by expressing and purifying recombinant forms of the algal lectin Griffithsin (GRFT), the most potent HIV-1 entry inhibitor yet described, from tobacco leaf tissues. GRFT has no known homologs in vertebrates, and hence has the potential to be immunogenic and thus possibly to induce an inflammatory response incompatible with its use as a microbicide. This is a theoretical concern that applies to most, if not all, peptide-based microbicides, and it is of vital importance to the field that this phenomenon be investigated more thoroughly. We will generate a mutant of GRFT that lacks lectin activity to allow us to answer the question whether lectin activity plays a role in induction of mucosal inflammatory responses. We will use a comprehensive mouse vaginal administration model, developed by our consultant and collaborator Dr. Betsy Herold, to evaluate the mucosal toxicity of hydroxyethylcellulose-formulated GRFT. Toxicology endpoints will include histology, measurement of molecular markers of inflammation, and phenotypes of leukocyte infiltrate in vaginal tissues. Since immunogenicity of protein microbicides is a concern, we will also measure immune responses to the vaginal microbicide in animals in which we have intentionally raised an anti-GRFT immune response. We will use Dr. Herold's elegant mouse genital herpes simplex virus (HSV) infectivity assay that functions as a surrogate measurement of microbicide safety. Susceptibility to challenge with HSV-2 in animals treated with GRFT will inform us whether the complex of biological responses to treatment with GRFT can increase susceptibility to infection with HSV-2, and hence to possible enhanced susceptibility to HIV-1 infection in humans. We will also study the mechanism of resistance to GRFT in a mutant HIV-1 Clade C virus that we isolated in a selection experiment. Collectively, the data that we generate in the course of this research will provide a comprehensive assessment of the utility of GRFT as a component of a vaginal microbicide to prevent HIV-1 transmission. PUBLIC HEALTH RELEVANCE: In the absence of an effective vaccine against HIV, there is an urgent need for alternative strategies to prevent sexual transmission of HIV, such as vaginally-applied microbicidal gels. We aim to answer the question whether proteins called lectins, which bind the sugar structures found on the surface of the AIDS virus, generate an inflammatory response when administered in the vagina of mice. These studies will provide important information that will help determine whether lectins have acceptable toxicity profiles to justify further development as microbicides.

描述（由申请人提供）：对于实际申请，必须有大量的生体可用，并且必须有效地产生成本。由于基于细胞的发酵系统的生产成本非常高，这些要求可能会从考虑中消除重组蛋白。我们通过表达和纯化的藻类凝集素griffithsin（GRFT）的重组形式开发了一种实用的解决方案，这是尚未从烟叶组织中描述的最有效的HIV-1进入抑制剂。 GRFT在脊椎动物中没有已知的同源物，因此具有免疫原性的潜力，因此可能诱导炎症反应与其用作菌心的不相容性。这是一个理论上的关注点，适用于大多数（如果不是全部）基于肽的杀菌剂，对领域至关重要，即对这种现象进行更彻底的研究至关重要。我们将产生一个缺乏凝集素活性的GRFT突变体，以使我们能够回答凝集素活性是否在诱导粘膜炎症反应中起作用。我们将使用由顾问和合作者Betsy Herold博士开发的全面小鼠阴道管理模型来评估羟基乙基纤维素形成的GRFT的粘膜毒性。毒理学终点将包括组织学，炎症分子标记的测量以及阴道组织中白细胞浸润的表型。由于蛋白质杀菌剂的免疫原性是一个问题，因此我们还将测量动物中对阴道菌心的免疫反应，在这种动物中，我们有意提高了抗Grft免疫反应。我们将使用Herold博士的优雅小鼠生殖器疱疹病毒（HSV）感染性测定，该测定功能是对杀菌剂安全的替代测量。在用GRFT治疗的动物中挑战HSV-2挑战的敏感性将告知我们是否会增加对GRFT治疗的复合物，是否可以增加对HSV-2感染的敏感性，从而增加对人类HIV-1感染的易感性。我们还将研究我们在选择实验中分离出的突变体HIV-1进化枝C病毒中对GRFT的抗性机制。总的来说，我们在这项研究过程中生成的数据将对GRFT作为阴道杀菌剂的组成部分进行全面评估，以防止HIV-1传播。公共卫生相关性：在没有针对艾滋病毒的有效疫苗的情况下，迫切需要采取替代策略来防止艾滋病毒的性传播，例如阴道施加的微生物凝胶。我们的目的是回答一个问题，蛋白质称为凝集素（结合在艾滋病病毒表面上发现的糖结构）是否会在小鼠的阴道中产生炎症反应。这些研究将提供重要的信息，这些信息将有助于确定凝集素是否具有可接受的毒性谱，以证明作为生液剂的进一步发展合理。