PuSH - Publication Server of Helmholtz Zentrum München: Cell-free production of personalized therapeutic phages targeting multidrug-resistant bacteria.

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Emslander, Q.* ; Vogele, K.* ; Braun, P.* ; Stender, J.* ; Willy, C.* ; Joppich, M.* ; Hammerl, J.A.* ; Abele, M.* ; Meng, C.* ; Pichlmair, A.* ; Ludwig, C.* ; Bugert, J.J.* ; Simmel, F.C.* ; Westmeyer, G.G.

Cell-free production of personalized therapeutic phages targeting multidrug-resistant bacteria.

Cell Chem. Bio. 29, 1434-1445.e7 (2022)

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Bacteriophages are potent therapeutics against biohazardous bacteria, which rapidly develop multidrug resistance. However, routine administration of phage therapy is hampered by a lack of rapid production, safe bioengineering, and detailed characterization of phages. Thus, we demonstrate a comprehensive cell-free platform for personalized production, transient engineering, and proteomic characterization of a broad spectrum of phages. Using mass spectrometry, we validated hypothetical and non-structural proteins and could also monitor the protein expression during phage assembly. Notably, a few microliters of a one-pot reaction produced effective doses of phages against enteroaggregative Escherichia coli (EAEC), Yersinia pestis, and Klebsiella pneumoniae. By co-expressing suitable host factors, we could extend the range of cell-free production to phages targeting gram-positive bacteria. We further introduce a non-genomic phage engineering method, which adds functionalities for only one replication cycle. In summary, we expect this cell-free methodology to foster reverse and forward phage engineering and customized production of clinical-grade bacteriophages.

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