Supplementary information files for article Targeted delivery of narrow-spectrum protein antibiotics to the lower gastrointestinal tract in a murine model of Escherichia coli colonization

Supplementary information files for Targeted delivery of narrow-spectrum protein antibiotics to the lower gastrointestinal tract in a murine model of Escherichia coli colonization <br> Bacteriocins are narrow-spectrum protein antibiotics that could potentially be used to engineer the human gut microbiota. However, technologies for targeted delivery of proteins to the lower gastrointestinal (GI) tract in preclinical animal models are currently lacking. In this work, we have developed methods for the microencapsulation of Escherichia coli targeting bacteriocins, colicin E9 and Ia, in a pH responsive formulation to allow their targeted delivery and controlled release in an in vivo murine model of E. coli colonization. Membrane emulsification was used to produce a water-in-oil emulsion with the water-soluble polymer subsequently cross-linked to produce hydrogel microcapsules. The microcapsule fabrication process allowed control of the size of the drug delivery system and a near 100% yield of the encapsulated therapeutic cargo. pH-triggered release of the encapsulated colicins was achieved using a widely available pH-responsive anionic copolymer in combination with alginate biopolymers. In vivo experiments using a murine E. coli intestinal colonization model demonstrated that oral delivery of the encapsulated colicins resulted in a significant decrease in intestinal colonization and reduction in E. coli shedding in the feces of the animals. Employing controlled release drug delivery systems such as that described here is essential to enable delivery of new protein therapeutics or other biological interventions for testing within small animal models of infection. Such approaches may have considerable value for the future development of strategies to engineer the human gut microbiota, which is central to health and disease.

大肠杆菌定植小鼠模型中靶向递送窄谱蛋白抗生素至下胃肠道的补充信息文件

细菌素（Bacteriocins）是一类窄谱蛋白抗生素，具备改造人类肠道微生物组的应用潜力。然而当前尚缺乏可在临床前动物模型中实现蛋白质靶向递送至下胃肠道（GI）的技术方案。本研究开发了一种pH响应型制剂，用于靶向大肠杆菌（Escherichia coli）的细菌素——大肠杆菌素E9与Ia的微囊化制备，使其可在大肠杆菌定植的小鼠体内模型中实现靶向递送与可控释放。实验采用膜乳化法制备油包水乳液，随后对水溶性聚合物进行交联以制备水凝胶微胶囊。该微胶囊制备工艺可精准调控药物递送系统的粒径，且包埋治疗性载荷的收率接近100%。通过将市售pH响应型阴离子共聚物与海藻酸盐生物聚合物复配，成功实现了包埋的大肠杆菌素的pH触发释放。采用小鼠大肠杆菌肠道定植模型开展的体内实验结果显示，口服递送包埋的大肠杆菌素可显著降低动物肠道内的定植负荷，并减少粪便中大肠杆菌的排出量。采用本文所述的控释药物递送系统，是在小型动物感染模型中测试新型蛋白疗法或其他生物干预手段的必要前提。此类方法未来在开发改造人类肠道微生物组的策略中具备重要应用价值，而肠道微生物组与机体健康和疾病进程密切相关。