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- Bacillus cereus (1)
- LasB (1)
- aerodyanmic properties (1)
- antibiotic resistance (1)
- bacillus subtilis (1)
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Scientific Unit
Pseudomonas aeruginosa (PA) infections can be notoriously difficult to treat and are often accompanied by the development of antimicrobial resistance (AMR). Quorum sensing inhibitors (QSI) acting on PqsR (MvfR) – a crucial transcriptional regulator serving major functions in PA virulence – can enhance antibiotic efficacy and eventually prevent the AMR. An integrated drug discovery campaign including design, medicinal chemistry-driven hit-to-lead optimization and in-depth biological profiling of a new QSI generation is reported. The QSI possess excellent activity in inhibiting pyocyanin production and PqsR reporter-gene with IC50 values as low as 200 and 11 × 10−9 m, respectively. Drug metabolism and pharmacokinetics (DMPK) as well as safety pharmacology studies especially highlight the promising translational properties of the lead QSI for pulmonary applications. Moreover, target engagement of the lead QSI is shown in a PA mucoid lung infection mouse model. Beyond that, a significant synergistic effect of a QSI-tobramycin (Tob) combination against PA biofilms using a tailor-made squalene-derived nanoparticle (NP) formulation, which enhance the minimum biofilm eradicating concentration (MBEC) of Tob more than 32-fold is demonstrated. The novel lead QSI and the accompanying NP formulation highlight the potential of adjunctive pathoblocker-mediated therapy against PA infections opening up avenues for preclinical development.
Liposomes have been studied for decades as nanoparticulate drug delivery systems for cytostatics, and more recently, for antibiotics. Such nanoantibiotics show improved antibacterial efficacy compared to the free drug and can be effective despite bacterial recalcitrance. In this work, we present a loading method of bacteriomimetic liposomes for a novel, hydrophobic compound (HIPS5031) inhibiting energy-coupling factor transporters (ECF transporters), an underexplored antimicrobial target. The liposomes were composed of DOPG (18:1 (Δ9-cis) phosphatidylglycerol) and CL (cardiolipin), resembling the cell membrane of Gram-positive Staphylococcus aureus and Streptococcus pneumoniae, and enriched with cholesterol (Chol). The size and polydispersity of the DOPG/CL/± Chol liposomes remained stable over 8 weeks when stored at 4 °C. Loading of the ECF transporter inhibitor was achieved by thin film hydration and led to a high encapsulation efficiency of 33.19% ± 9.5% into the DOPG/CL/Chol liposomes compared to the phosphatidylcholine liposomes (DMPC/DPPC). Bacterial growth inhibition assays on the model organism Bacillus subtilis revealed liposomal HIPS5031 as superior to the free drug, showing a 3.5-fold reduction in CFU/mL at a concentration of 9.64 µM. Liposomal HIPS5031 was also shown to reduce B. subtilis biofilm. Our findings present an explorative basis for bacteriomimetic liposomes as a strategy against drug-resistant pathogens by surpassing the drug-formulation barriers of innovative, yet unfavorably hydrophobic, antibiotics.
Abstract Despite the progress in surgical techniques and antibiotic prophylaxis, opportunistic wound infections with Bacillus cereus remain a public health problem. Secreted toxins are one of the main factors contributing to B. cereus pathogenicity. A promising strategy to treat such infections is to target these toxins and not the bacteria. Although the exoenzymes produced by B. cereus are thoroughly investigated, little is known about the role of B. cereus collagenases in wound infections. In this report, the collagenolytic activity of secreted collagenases (Col) is characterized in the B. cereus culture supernatant (csn) and its isolated recombinantly produced ColQ1 is characterized. The data reveals that ColQ1 causes damage on dermal collagen (COL). This results in gaps in the tissue, which might facilitate the spread of bacteria. The importance of B. cereus collagenases is also demonstrated in disease promotion using two inhibitors. Compound 2 shows high efficacy in peptidolytic, gelatinolytic, and COL degradation assays. It also preserves the fibrillar COLs in skin tissue challenged with ColQ1, as well as the viability of skin cells treated with B. cereus csn. A Galleria mellonella model highlights the significance of collagenase inhibition in vivo.
Pulmonary fibrosis, a disabling lung disease, results from the fibrotic transformation of lung tissue. This fibrotic transformation leads to a deterioration of lung capacity, resulting in significant respiratory distress and a reduction in overall quality of life. Currently, the frontline treatment of pulmonary fibrosis remains limited, focusing primarily on symptom relief and slowing disease progression. Bacterial infections with Pseudomonas aeruginosa are contributing to a severe progression of idiopathic pulmonary fibrosis. Phytic acid, a natural chelator of zinc, which is essential for the activation of metalloproteinase enzymes involved in pulmonary fibrosis, shows potential inhibition of LasB, a virulence factor in P. aeruginosa, and mammalian metalloproteases (MMPs). In addition, phytic acid has anti-inflammatory properties believed to result from its ability to capture free radicals, inhibit certain inflammatory enzymes and proteins, and reduce the production of inflammatory cytokines, key signaling molecules that promote inflammation. To achieve higher local concentrations in the deep lung, phytic acid was spray dried into an inhalable powder. Challenges due to its hygroscopic and low melting (25 °C) nature were mitigated by converting it to sodium phytate to improve crystallinity and powder characteristics. The addition of leucine improved aerodynamic properties and reduced agglomeration, while mannitol served as carrier matrix. Size variation was achieved by modifying process parameters and were evaluated by tools such as the Next Generation Impactor (NGI), light diffraction methods, and scanning electron microscopy (SEM). An inhibition assay for human MMP-1 (collagenase-1) and MMP-2 (gelatinase A) allowed estimation of the biological effect on tissue remodeling enzymes. The activity was also assessed with respect to inhibition of bacterial LasB. The formulated phytic acid demonstrated an IC50 of 109.7 µg/mL for LasB with viabilities > 80 % up to 188 µg/mL on A549 cells. Therefore, inhalation therapy with phytic acid-based powder shows promise as a treatment for early-stage Pseudomonas-induced pulmonary fibrosis.
