Use of Bacteriophages to Prevent, Diagnose, and Treat Diseases

部分A.目前正在进行三方面的调查。1.噬菌体在治疗由细菌感染引起的人类疾病中的应用。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，照顾感染过程中出现的噬菌体耐药性。2.我们已经成功地将克雷伯氏菌工程改造成具有多个尾部以增加噬菌体的宿主范围，从而能够感染和杀死多个宿主物种。3.我们已经成功地构建了一个T7噬菌体衍生物，其中我们克隆了一个肽抗生素基因（AMP），当感染给定的宿主时，该基因也会产生大量的抗生素。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性的突变细菌宿主。然而，AMP的表达一直很差，其不能杀死噬菌体抗性宿主。但是当AMP表达与可溶性蛋白共价连接时，表达是合理的。目前，我们已经建立了一个携带肽抗生素随机突变的噬菌体文库，并筛选出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。我们正在使用我们以前开发的噬菌体展示技术，使用噬菌体λ来制造针对传染病和癌症的疫苗。我们已经构建了展示CLL白血病，疟疾和Covid 19病毒的潜在抗原肽和蛋白的载体。我们投入了大量的时间来工程化，纯化，验证和测试多个噬菌体构建体的真实性，这些构建体可能被开发为COVID-19，CLL和疟疾的噬菌体疫苗。目前，我们正在进行几项实验，并分析在小鼠模型中测试基于噬菌体的疫苗的数据，以产生针对COVID-19和胶质母细胞瘤的免疫力。我们目前正在制定未来的实验计划，以证明基于噬菌体的COVID-19疫苗的有效性。两种情况下的目标是相同的：制备纯化的工程λ用于动物免疫原性研究。在该项目的第一个分支中，我们用胶质母细胞瘤中存在的EGFRvIII抗原设计了一种噬菌体，并附着在λ噬菌体的D蛋白上。该项目的另一个分支涉及相同的过程，但抗原是新冠病毒刺突蛋白的一个片段。这些建筑物是作为先知建造的。然后诱导噬菌体产生前噬菌体，并进行快速纯化和浓缩。目前正在动物中测试各自的免疫原性。

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.