Toxoplasma in the GI tract: Protective role of a parasite protease inhibitor

胃肠道中的弓形虫：寄生虫蛋白酶抑制剂的保护作用

• 批准号: 10197034
• 负责人: Isabelle Coppens
• 金额: $ 20.47万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-17 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10197034
• 关键词: AIDS/HIV problem; Abbreviations; Adverse effects; Affinity; Affinity Chromatography; Animals; Antibody Affinity; Basement membrane; Binding; Biochemical; Biological Assay; Blood; Blood Circulation; Blood Vessels; Brain; Cathepsin G; Cell Death; Cells; Cessation of life; Chorioretinitis; Code; Complement; Cyst; Cytoplasmic Granules; DNA; Data; Development; Disease; Encephalitis; Enterocytes; Environment; Enzymes; Epithelial; Epithelial Cells; Event; Family; Gastrointestinal tract structure; Genes; Genetic; Goals; Histones; Human; Immune; Immune mediated destruction; Immune response; Immunocompetent; Immunocompromised Host; In Vitro; Individual; Infection; Ingestion; Internet; Intestines; Invaded; Lactoferrin; Lamina Propria; Leukocyte Elastase; Leukocyte L1 Antigen Complex; Location; Longevity; Loose connective tissue; Lymphatic; Mass Spectrum Analysis; Membrane; Monitor; Morphology; Muramidase; Mus; Muscle; Myocarditis; Myocardium; NADPH Oxidase; Names; Natural Immunity; Oocysts; Oral; Oral Administration; Oral Ingestion; Organ; Pancreatic Elastase; Parasites; Patients; Peptide Hydrolases; Phagocytosis; Pharmaceutical Preparations; Pharmacology; Pharmacotherapy; Phase; Play; Predisposition; Prevalence; Protease Inhibitor; Proteins; Recombinants; Resistance; Respiratory Burst; Risk; Role; Running; Rupture; Serine Protease; Serine Proteinase Inhibitors; Site; Skeletal Muscle; Small Intestines; Sporozoites; Stomach; Structure; Therapeutic; Tissues; Toxoplasma; Toxoplasma gondii; Toxoplasmosis; Travel; Trypsin; Vaccines; Vacuole; Virulence Factors; alpha-Defensins; antimicrobial; antimicrobial peptide; assault; cell type; chemokine; chymotrypsin; extracellular; in vitro Assay; intraperitoneal; member; monocyte; muscle form; neutrophil; new therapeutic target; novel therapeutics; pathogen; response; scaffold

SUMMARY Toxoplasma gondii is an opportunistic protozoan parasite that poses a significant risk to AIDS/HIV patients. The currently available drugs for toxoplasmosis have significant adverse effects and are ineffective to eradicate long- lived tissue cysts located in the brain and muscles. Identifying new drug targets is imperative to control Toxoplasma infections with new drugs. Natural infection by T. gondii occurs via oral ingestion of tissue cysts (containing bradyzoites) or environmental oocysts (containing sporozoites). After ingestion of tissue cysts, excysted bradyzoites invade enterocytes where they transform into fast-replicating tachyzoite forms. After several cycles of replication in the small intestine and gut organs, tachyzoites migrate to the lamina propria where they enter the bloodstream to reach the brain and muscles to form tissue cysts. How Toxoplasma cysts are able to survive in the harsh environment of the stomach and gut tissues, and escape destruction by immune cells in the lamina propria remains largely unknown. Coccidian parasites (e.g., Toxoplasma), which need to be ingested orally by a host to initiate an infection, have in common the presence of genes coding for serine protease inhibitors that may target host serine proteases. Our proposal focuses on the contribution of a highly abundant serine protease inhibitor of T. gondii, named TgPI-1 to the protection of the parasite against host serine proteases present in the gut lumen (pancreatic elastase, trypsin, chymotrypsin), and/or secreted by immune cells in the lamina propria (neutrophil elastase, cathepsin G). We showed that TgPI-1 is secreted by tachyzoites and bradyzoites from dense granules, and inhibits trypsin, chymotrypsin and neutrophil elastase in vitro. Compared to wild-type parasites, TgPI-1- deficient parasites (cystogenic, type II strain) are more vulnerable to serine proteases added to the medium and have reduced dissemination in the gastro-intestinal tract of mice after oral administration of tissue cysts. Our hypothesis is that TgPI-1 contributes to the protection of T. gondii at the onset of infection in the gut. Specific Aim 1 will identify the TgPI-1 targets and sites where ΔTgPI-1 parasites are killed in the gastro-intestinal tract to reveal the sites where TgPI-1 is secreted to protect Toxoplasma in the gut. Specific Aim 2 will focus on the contribution of TgPI-1 to protect the parasite in the lamina propria by inhibiting neutrophil elastase secreted by activated neutrophils or released by dying neutrophils during extrusion of Neutrophil Extracellular Traps (NET) wherein neutrophil elastase is highly abundant. From a therapeutic point-of-view, TgPI-1 could be a newly identified virulence factor, and its pharmacological inhibition in combination with current drug treatments, may increase the efficacy of anti-Toxoplasma treatments.

概括 弓形虫是一种机会性原生动物寄生虫，对艾滋病/艾滋病毒患者构成重大风险。 目前可用的治疗弓形虫病的药物具有显着的副作用，并且不能有效根除长期弓形虫病。 确定新的药物靶点对于控制弓形虫至关重要。 新药感染。弓形虫通过口腔摄入组织包囊（含有包囊）而发生自然感染。 缓殖子）或环境卵囊（含有子孢子）摄入组织包囊后，排出。 缓殖子侵入肠细胞，经过几个循环后，它们转化为快速复制的速殖子形式。 在小肠和肠道器官中复制时，速殖子迁移到固有层，在那里它们进入 血液到达大脑和肌肉形成组织囊肿，弓形虫囊肿如何在体内存活。 胃和肠道组织的恶劣环境，并逃避固有层免疫细胞的破坏 球虫寄生虫（例如弓形虫）仍然很大程度上未知，它们需要被宿主口服摄入。 引发感染，共同点是存在编码丝氨酸蛋白酶抑制剂的基因，这些抑制剂可能靶向 我们的提案重点关注高丰度丝氨酸蛋白酶抑制剂的贡献。 刚地弓形虫，命名为 TgPI-1，用于保护寄生虫免受肠腔中存在的宿主丝氨酸蛋白酶的侵害 （胰腺弹性蛋白酶、胰蛋白酶、胰凝乳蛋白酶）和/或由固有层中的免疫细胞（中性粒细胞）分泌 我们发现，TgPI-1 是由致密颗粒中的速殖子和缓殖子分泌的， 与野生型寄生虫相比，TgPI-1- 在体外可抑制胰蛋白酶、胰凝乳蛋白酶和中性粒细胞弹性蛋白酶。 缺陷寄生虫（囊原性，II 型菌株）更容易受到添加到培养基中的丝氨酸蛋白酶的影响，并且 口服组织囊肿后，小鼠胃肠道的传播减少。 假设 TgPI-1 在肠道感染开始时有助于保护弓形虫。 1 将确定胃肠道中 ΔTgPI-1 寄生虫被杀死的 TgPI-1 目标和位点，以揭示 具体目标 2 将重点关注分泌 TgPI-1 以保护肠道内弓形虫的位点。 TgPI-1 通过抑制活化的中性粒细胞弹性蛋白酶分泌来保护固有层中的寄生虫 中性粒细胞或在中性粒细胞胞外陷阱 (NET) 挤出过程中由垂死的中性粒细胞释放 从治疗的角度来看，TgPI-1 可能是一种新发现的。 毒力因子及其药理学抑制与目前的药物治疗相结合，可能会增加 抗弓形虫治疗的功效。

项目成果

MalDA, Accelerating Malaria Drug Discovery.

• DOI: 10.1016/j.pt.2021.01.009
• 发表时间: 2021-06
• 期刊: Trends in parasitology
• 影响因子: 9.6
• 作者: Yang T;Ottilie S;Istvan ES;Godinez-Macias KP;Lukens AK;Baragaña B;Campo B;Walpole C;Niles JC;Chibale K;Dechering KJ;Llinás M;Lee MCS;Kato N;Wyllie S;McNamara CW;Gamo FJ;Burrows J;Fidock DA;Goldberg DE;Gilbert IH;Wirth DF;Winzeler EA;Malaria Drug Accelerator Consortium
• 通讯作者: Malaria Drug Accelerator Consortium