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Since effective treatment of patients with chronic lung disease is often complicated by infections with multidrug resistant pathogens like Pseudomonas aeruginosa, bacteriophage therapy is appealing. In our study, we evaluated a phage cocktail, manufactured according to quality standards for good manufacturing practice, for its potential to treat P. aeruginosa experimental lung infection. Human lung tissue samples were infected ex vivo with clinical strains of P. aeruginosa and phages or control solution were applied. Bacterial loads and phage titers were assessed at different time points. Mice were infected with PBS or P. aeruginosa, and intratracheally treated with phages or control solution 6 hpi. Subsequently, phage titers, pathogen load, and inflammatory response were analyzed, and lungs were examined histopathologically. Replicative phages were recovered from human lung tissue, and phage titers increased over time. Bacterial load was reduced by phage application. In mice, active phages were recovered locally and systemically in both infected and sham infected animals. Phage treatment resulted in reduced bacterial load 12 and 48 hpi in BAL and slightly reduced signs of inflammation in histopathology. Analyses of innate immune cell response of infected and phage treated animals compared to the infected placebo group showed no phage-specific effects. Lower levels of IL-1b were detected in BAL and blood of infected and phage treated mice. In sham-infected animals, phage application resulted in increased levels of few cytokines 12 hpi, with no apparent adverse effects. Our study strengthened the approach to develop a manufactured bacteriophage cocktail for P. aeruginosa lung infections.