Preventing norovirus and Clostridium difficile gastroenteritis by engineered probiotic yeast Saccharomyces boulardii secreting multi-specific single-domain antibodies

通过分泌多特异性单域抗体的工程益生菌布拉酵母菌预防诺如病毒和艰难梭菌胃肠炎

• 批准号: 10540345
• 负责人: Hanping Feng
• 金额: $ 61.13万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-01-22 至 2024-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10540345
• 关键词: Acute; Animal Disease Models; Animal Model; Antibiotic Resistance; Antibiotic Therapy; Antibiotics; Antibodies; Binding; Body Weight decreased; Carbohydrates; Characteristics; Chromosomal Insertion; Clinic; Clinical; Clostridium difficile; Communicable Diseases; DNA cassette; Data; Development; Diarrhea; Disease; Dose; Dried Yeast; Drug or chemical Tissue Distribution; Dryness; Encapsulated; Engineered Probiotics; Engineering; Enteral; Erythrocytes; Excipients; Family suidae; Formulation; Future; Gastroenteritis; Genes; Genome; Gnotobiotic; Goals; Hand; Health care facility; Hemagglutination; Hospitalization; Human; In Vitro; Individual; Infection; Infection prevention; Intestinal Secretions; Intestines; Lead; Long-Term Care; Marketing; Mediating; Methods; Monitor; Morbidity - disease rate; Mus; Norovirus; Oral; Oral Administration; Patients; Prevention strategy; Preventive; Probiotics; Process; Production; Prophylactic treatment; Saccharomyces; Site; Symptoms; Technology; Testing; Therapeutic; Toxin; Traveler' s diarrhea; Vaccines; Variant; Viral; Viral Load result; Virulence Factors; Virus-like particle; Yeasts; aged; antibiotic-associated diarrhea; antitoxin; capsule; commercialization; enteric pathogen; gut microbiota; high risk population; human model; in vivo; microbiota; microbiota transplantation; mortality; nanobodies; novel; passive antibodies; pathogen; pathogenic bacteria; prevent; scale up; technology platform; tool; transmission process; vaccine access; vaccine development

Abstract Human norovirus (hNoV) and Clostridium difficile (CD) represent two leading causes of acute gastroenteritis worldwide with significant morbidity and mortality. The two infections (one viral and one bacterial) share many of the same characteristics of transmission; and concurrent infections are particularly prevalent in the US in high- risk populations, such as aged patients undergoing antibiotic treatments, hospitalized patients, or patients staying in long-term care facilities. Despite great efforts in the development of vaccines against both infections, to date there is still no single vaccine available on the market. We have developed a novel platform technology against enteric bacterial pathogens by engineering a probiotic yeast, Saccharomyces boulardii, to secrete multi- specific single-domain (VHH) antibodies, directly targeting bacterial virulence factors at the intestinal site of infection. We have successfully applied this technology to target Clostridium difficile. The overall objective of this project is to generate Sb strains secreting mSdAbs against both CD toxins and hNoV, generate proof-of-principle efficacy data in relevant animal disease models, and develop clinic-compatible formulations for drying and encapsulating the Sb-mSdAb strains. We hypothesize that oral administration of Sb-mSdAb strains secreting mSdAb against hNoV and CD toxins will prevent their individual or concurrent infections. To test this hypothesis, we propose to accomplish the following 3 specific aims: 1) Engineer Sb-mSdAb strains (Sb-aNoVCd) secreting VHH fusions against both CD toxins and hNoVs. 2) Determine preventive efficacy of Sb-aNoVCd strains against infections with hNoV and CD in gnotobiotic pigs. 3) Develop a formulation supporting yeast spray drying and encapsulation. With the completion of proposed translational activities, we will have generated lead Sb-aNoVCd strains and evaluated in vivo characteristics in mice and in gnotobiotic pigs which is the most appropriate animal model of hNoV infection and disease. Should the key proof-of-principle efficacy data generated, future scale-up efforts will be justified to include additional VHHs for generating broadly efficacious lead immunoprophylactic strains and eventual commercialization of the yeast products against CD and hNoV gastroenteritis for which we currently have no prophylaxis or vaccines.

抽象的 人类诺如病毒（HNOV）和艰难梭菌（CD）是急性胃炎的两个主要原因 在全球范围内具有明显的发病率和死亡率。两种感染（一种病毒和一种细菌）共享许多 传播的相同特征；在美国，同时感染尤其普遍 风险种群，例如接受抗生素治疗的老年患者，住院患者或患者 留在长期护理设施中。尽管在针对两种感染的疫苗开发中做出了巨大努力，但 迄今为止，市场上仍然没有单一的疫苗。我们已经开发了一种新颖的平台技术 针对肠细菌病原体，通过设计益生菌酵母，saccharomyces boulardii，分泌多种 特定的单域（VHH）抗体，直接靶向细菌毒力因子 感染。我们已经成功地应用了这项技术来针对艰难梭菌。总体目标 项目将产生SB菌株，将MSDAB分泌针对CD毒素和HNOV，生成原理证明 相关动物疾病模型中的功效数据，并开发与诊所兼容的制剂，以进行干燥和 封装SB-MSDAB菌株。我们假设口服SB-MSDAB菌株的口服分泌 针对HNOV和CD毒素的MSDAB将阻止其个体或并发感染。为了检验这一假设， 我们建议完成以下3个具体目标：1）工程师SB-MSDAB菌株（SB-Anovcd）分泌 VHH融合了CD毒素和HNOVS。 2）确定SB-ANOVCD菌株的预防功效 在gnotobiotic猪中用HNOV和CD感染。 3）开发一种配方，以支持酵母喷雾干燥和 封装。随着拟议的翻译活动的完成，我们将产生铅SB-AnoVCD 菌株并评估小鼠和gnotobiotic猪的体内特征，这是最合适的动物 HNOV感染和疾病的模型。应该生成的关键原理证明数据，未来的扩展 努力将是合理的，以包括额外的VHHS来产生广泛有效的铅免疫预防性 酵母产品针对CD和HNOV胃肠炎的菌株和最终商业化 目前没有预防或疫苗。