Project description:The status quo for combating uprising antibacterial resistance is to employ synergistic combinations of antibiotics. Nevertheless, the currently available combination therapies are fast becoming untenable. Combining antibiotics with various FDA-approved non-antibiotic drugs has emerged as a novel strategy against otherwise untreatable multi-drug resistant (MDR) pathogens. The apex of this study was to investigate the mechanisms of antibacterial synergy of the combination of polymyxin B with the phenothiazines against the MDR Gram-negative pathogens Acinetobacter baumannii, Klebsiella pneumoniae and Pseudomonas aeruginosa. The synergistic antibacterial effects were tested using checkerboard and static time-kill assays. Electron microscopy (EM) and untargeted metabolomics were used to ascertain the mechanism(s) of the antibacterial synergy. The combination of polymyxin B and the phenothiazines showed synergistic antibacterial activity in checkerboard and static time-kill assays at clinically relevant concentrations against both polymyxin-susceptible and polymyxin-resistant isolates. EM revealed that the polymyxin B-prochlorperazine combination resulted in greater damage to the bacterial cell compared to each drug monotherapy. In metabolomics, at 0.5 h, polymyxin B monotherapy and the combination (to a greatest extent) disorganised the bacterial cell envelope as manifested by a major perturbation in bacterial membrane lipids (glycerophospholipids and fatty acids), peptidoglycan and lipopolysaccharide (LPS) biosynthesis. At the late time exposure (4 h), the aforementioned effects (except LPS biosynthesis) perpetuated mainly with the combination therapy, indicating the disorganising bacterial membrane biogenesis is potentially behind the mechanisms of antibacterial synergy. In conclusion, the study highlights the potential usefulness of the combination of polymyxin B with phenothiazines for the treatment of polymyxin-resistant Gram-negative infections (e.g. CNS infections).

Project description:The inability of the adult human heart to regenerate lost or damaged myocardial tissue has created one of the most pressing public health dilemmas due to the devastating impact of heart failure. Our group and others have outlined several regulators of cardiomyocyte mitosis that may impact the regenerative capacity of the adult myocardium in mammals. Recently, we reported that the transcription factors Meis1 and Hoxb13 regulate postnatal cardiomyocyte cell cycle arrest, where concomitant deletion of both genes induced cardiomyocyte proliferation and myocardial regeneration following ischemic injury. These studies suggest that pharmacological targeting Meis1 and Hoxb13 transcriptional activity could be a viable path towards heart regeneration. Therefore, we performed an in-silico screen to identify FDA-approved drugs that can inhibit Meis1 and Hoxb13 transcriptional activity based on the published crystal structure of Meis1 and Hoxb13 bound to DNA. Our screen yielded several candidates based on binding profiles to either Meis1 DNA binding domain, Hoxb13 DNA binding domain, or the interface between Meis1 and Hoxb13, as well as safety and side effect profiles. Out of the shortlist of top hits, paromomycin and neomycin induced proliferation of neonatal rat ventricular myocytes in vitro and displayed dose-dependent inhibition of Meis1 and Hoxb13 transcriptional activity by luciferase assay, and disruption of DNA binding by EMSA. X-ray crystal structure revealed that both paromomycin and neomycin bind to Meis1 near the Hoxb13 interaction site where they interact with 3 out of the 5 Meis1 amino acids that participate in DNA-binding. The crystal structure also demonstrates that both drugs bind at the interface of Meis1 homodimer, therefore placing one of the Meis1 monomers in a position incompatible with DNA-binding. Importantly, administration of paromomycin-neomycin combination by intraperitoneal injection in mice induced cardiomyocyte proliferation, improved left ventricular (LV) systolic function and decreased scar formation in two models of ischemic reperfusion injury in mice. Finally, dual intravenous administration of the paromomycin-neomycin combination in Yorkshire pigs following cardiac ischemia/reperfusion injury induced cardiomyocyte proliferation, improved LV systolic function, and decreased scar formation. Collectively, we identified paromomycin and neomycin as FDA-approved drugs with therapeutic potential for induction of heart regeneration in humans.

Dataset Information

Evaluation of a Library of FDA-Approved Drugs for Their Ability To Potentiate Antibiotics against Multidrug-Resistant Gram-Negative Pathogens.

Publications

Evaluation of a Library of FDA-Approved Drugs for Their Ability To Potentiate Antibiotics against Multidrug-Resistant Gram-Negative Pathogens.

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