SBIR Phase I: Bacteriophage-Based Microbial Gene Therapy Platform for In Situ Engineering of Microbiomes

这个小企业创新研究(SBIR)第一阶段项目的更广泛影响将是微生物群落的稳定和持续调节--微生物群落分布在人体不同区域。所有生物体都有一个相关的微生物区系，这些微生物区系在生物体的生长、健康、免疫和抗逆性方面发挥着密切的作用。目前调节微生物组的方法，包括使用益生元、益生菌和其他生命疗法，缺乏证明的诱导微生物区系长期变化的能力。相比之下，该项目使用噬菌体--感染细菌但不影响人类宿主的病毒--对微生物组中驻留的细菌成员进行长期或永久的改变。作为概念的证明，最初的开发将专注于改变人类肠道微生物来治疗与面筋相关的疾病，从轻微的面筋敏感到乳糜泻。最终，这项技术可以被改造来解决大量与微生物组相关的问题，这些问题跨越了广泛的临床、商业和社会利益。潜在的健康应用包括与微生物组相关的疾病，如肥胖症、2型糖尿病、动脉粥样硬化、心血管疾病、炎症性肠道疾病、自身免疫性疾病、牙周炎和龋病、多种癌症、皮肤病和反复发生的细菌感染。其他潜在的应用包括农业和环境领域。拟议的项目将利用噬菌体的效用和力量来转导遗传信息。由于除了初步的概念验证实验之外，还没有尝试通过工程噬菌体来调节微生物组，因此建立拟议的微生物基因治疗平台将需要研究和开发工作，以克服许多技术挑战。本课题的研究目标包括：1)构建针对双歧杆菌的温和型噬菌体，表达囊型鞘氨醇单胞菌的面筋蛋白降解酶；2)将表达谷氨酸酶的噬菌体导入长杆菌体外生物膜模型中。该项目将提高对噬菌体-宿主相互作用、基因回路调控和水平基因转移管理的认识，这将对应用病毒学、遗传工程和微生物学领域的创新产生深远影响。特别是，这个项目的最终成功将依赖于使用和开发合成生物学方法来增强噬菌体的能力，例如从宿主基因组灵活地整合和切除基因，管理宿主内的表达，在生活方式之间转换噬菌体(例如，裂解到溶源和返回)，扩大噬菌体宿主范围，以及中和细菌宿主防御系统。这一奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

The broader impact of this Small Business Innovation Research (SBIR) Phase I project will be the stable and persistent modulation of microbiomes – the community of microorganisms that populate the different regions of a person’s body. All organisms have an associated microbiota that play an intimate role in that organism’s growth, health, immunity, and stress tolerance. Current approaches for modulating the microbiome, including the use of prebiotics, probiotics, and other living therapeutics, lack a demonstrated ability to induce long-term changes in the microbiota. In contrast, this project uses bacteriophages—viruses that infect bacteria but do not affect the human host—for long-term or permanent alteration of resident bacterial members of the microbiome. As a proof of concept, initial development will focus on altering human gut microbes to treat gluten-related disorders, ranging from mild gluten sensitivity to celiac disease. Ultimately, this technology could be adapted to address a vast number of microbiome-related issues that span a wide range of clinical, commercial, and societal interests. Potential health applications include microbiome-related conditions such as obesity, type 2 diabetes, atherosclerosis, cardiovascular disease, inflammatory bowel disease, autoimmune disorders, gingivitis and caries, multiple types of cancer, skin disorders, and recurrent bacterial infections. Other potential applications include agricultural and environmental fields.The proposed project will exploit the utility and power of bacteriophages to transduce genetic information. As engineering bacteriophages to modulate the microbiome has not yet been attempted beyond preliminary proof-of-concept experiments, establishment of the proposed microbial gene therapy platform will require research and development efforts to overcome numerous technical challenges. The objectives of this project include: 1) engineering Bifidobacterium-targeting temperate bacteriophage capable of infecting B. longum to express a gluten-degrading enzyme from Sphingomonas capsulata and 2) introducing the glutenase-expressing phage into a B. longum in vitro biofilm model. This project will result in enhanced knowledge of bacteriophage–host interactions, genetic circuit modulation, and management of horizontal gene transfer, which will have far-reaching implications for innovation in the fields of applied virology, genetic engineering, and microbiomics. In particular, the ultimate success of this project will rely upon the use and development of synthetic biological approaches to enhance bacteriophage capabilities, such as flexibly integrating and excising genes from host genomes, managing intra-host expression, converting phages between lifestyles (e.g. lytic to lysogenic and back), expanding phage host range, and neutralizing bacterial host defense systems.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.