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组的稳定和持续调节-微生物群落分布在人体的不同区域。所有生物体都有一个相关的微生物群，它们在生物体的生长、健康、免疫和耐受性中起着密切的作用。目前调节微生物群的方法，包括使用益生元、益生菌和其他活疗法，缺乏诱导微生物群长期变化的证明能力。相比之下，这个项目使用噬菌体——感染细菌但不影响人类宿主的病毒——来长期或永久地改变微生物组中的常驻细菌成员。作为概念验证，最初的开发将集中在改变人类肠道微生物来治疗与谷蛋白相关的疾病，从轻度谷蛋白敏感性到乳糜泻。最终，这项技术可以用于解决大量与微生物组相关的问题，这些问题涉及广泛的临床、商业和社会利益。潜在的健康应用包括微生物组相关的疾病，如肥胖、2型糖尿病、动脉粥样硬化、心血管疾病、炎症性肠病、自身免疫性疾病、牙龈炎和龋齿、多种类型的癌症、皮肤病和复发性细菌感染。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。由于工程噬菌体调节微生物组的尝试尚未超出初步的概念验证实验，因此建立拟议的微生物基因治疗平台将需要研究和开发努力，以克服许多技术挑战。本项目的目标包括：1)构建双歧杆菌靶向型温带噬菌体，使其能够感染长芽胞杆菌，并表达来自荚膜鞘单胞菌的谷蛋白降解酶；2)将表达谷蛋白酶的噬菌体导入长芽胞杆菌体外生物膜模型。该项目将提高对噬菌体-宿主相互作用、遗传电路调制和水平基因转移管理的认识，这将对应用病毒学、基因工程和微生物学领域的创新产生深远的影响。特别是，该项目的最终成功将依赖于合成生物学方法的使用和发展，以增强噬菌体的能力，如灵活地整合和切除宿主基因组中的基因，管理宿主内表达，在不同的生活方式之间转换噬菌体（例如裂解到溶原和返回），扩大噬菌体宿主范围，中和细菌宿主防御系统。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值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.