Use of Bacteriophages to Prevent, Diagnose, and Treat Diseases

A部分目前正在进行三个方面的调查。1.使用噬菌体治疗因细菌感染而引起的人类疾病。我们目前正在处理由克雷伯氏菌引起的感染。我们已经成功地定义了噬菌体鸡尾酒，这种鸡尾酒可以治疗感染过程中产生的噬菌体耐药性。2.我们成功地设计了克雷伯氏菌噬菌体的多个尾巴，以增加一个噬菌体的寄主范围，使其能够感染和杀死多个寄主物种。3.我们成功地构建了T7噬菌体衍生物，在其中克隆了一个多肽抗生素基因(AMP)，当感染给定的宿主时，它也产生大量的抗生素。这使得噬菌体能够通过AMP的作用杀死任何对噬菌体产生抗药性的突变细菌宿主。然而，AMP的表达一直很差，不能杀死抗噬菌体的宿主。但当AMP的表达与可溶性蛋白共价连接时，其表达是合理的。目前，我们已经建立了一个携带抗菌肽随机突变的噬菌体文库，并对其进行筛选，以寻找具有更有效抗菌活性的噬菌体。这些发现鼓励我们进一步推进这一进程。B部分我们正在使用我们之前开发的噬菌体展示技术，使用噬菌体Lambda来制造针对传染病和癌症的疫苗。我们已经构建了CLL白血病、疟疾和Covid19病毒的潜在抗原肽和蛋白的展示。我们投入了大量时间来设计、纯化、验证和测试多个噬菌体构建体的真实性，这些噬菌体构建体有可能被开发为基于新冠肺炎、CLL和疟疾的噬菌体疫苗。目前，我们正在进行几项实验并分析数据，测试基于噬菌体的疫苗在小鼠模型中产生对新冠肺炎和胶质母细胞瘤的免疫力。我们目前正在进行未来的实验计划，以展示基于噬菌体的新冠肺炎疫苗的有效性。在这两种情况下，目标是相同的：制备纯化的、用于动物免疫原性研究的工程Lambda。在该项目的第一个分支中，我们利用胶质母细胞瘤中存在的EGFRvIII抗原设计了一种噬菌体，并将其连接到Lambda噬菌体的D蛋白上。该项目的另一个分支参与了同样的过程，但抗原是新冠肺炎刺突蛋白的一段。这些建筑是作为先知而建造的。然后诱导噬菌体生产，并纯化和浓缩噬菌体。现在正在动物身上测试各自的噬菌体的免疫原性。

Part A. Three lines of investigations are being pursued currently. 1. Use of bacteriophages in treating human diseases because of bacterial infections. We are currently dealing with infections caused by Klebsiella. We have succeeded in defining bacteriophage cocktails that take care of bacteriophage resistance that arises during infection processes. 2. We have succeeded in engineering Klebsiella phages to have multiple tails to increase the host range of a phage to be capable of infecting and killing multiple hosts species. 3. We have succeeded to construct a T7 bacteriophage derivative in which we cloned a peptide antibiotic gene (AMP), which when infects a given host also produces vast amount of the antibiotic. This enables the phage to kill by the action of AMP any mutant bacterial host which becomes resistant to phage. However, the expression of AMP has been poor, which does not kill phage resistant hosts. But when the AMP expression is covalently linked to a soluble protein, the expression was reasonable. Currently, we have made a library of phage carrying random mutations in the peptide anti biotic and screening them for one with more effective anti-bacterial activity. These findings encourage us to pursue the process further. Part B. We are using our previously developed phage display technology using bacteriophage Lambda to make vaccines against infectious diseases as well as against cancer. We already constructed displaying potential antigenic peptides and proteins of CLL leukemia, malaria and Covid 19 virus. We invested a lot of time in engineering, purifying, validating, and testing the authenticity of multiple phage constructs that could potentially be developed as a phage-based vaccine for COVID-19, CLL, and malaria. Currently, we are performing several experiments and analyzing data on testing phage-based vaccines in a mouse model to generate immunity against COVID-19 and glioblastoma. We are currently working in the future experimental plan to demonstrate the efficacy of phage-based COVID-19 vaccine. The goal is the same in both cases: to prepare purified, engineered lambda for animal immunogenecity study. In the first branch of the project, we engineered a phage with the EGFRvIII antigen present in glioblastoma and attached to the D protein of the lambda phage. The other branch of the project involved the same process, but the antigen was a fragment of the covid-19 spike protein. These constructions were made as prophages. The prophages were then induced phage production and phages purified and concentrated. The respective phages are now being tested for immunogenecity in animals.