• The bacterial cell envelope as delimiter of anti-infective bioavailability - An in vitro permeation model of the Gram-negative bacterial inner membrane.

      Graef, Florian; Vukosavljevic, Branko; Michel, Jean-Philippe; Wirth, Marius; Ries, Oliver; De Rossi, Chiara; Windbergs, Maike; Rosilio, Véronique; Ducho, Christian; Gordon, Sarah; et al. (2016)
      Gram-negative bacteria possess a unique and complex cell envelope, composed of an inner and outer membrane separated by an intermediate cell wall-containing periplasm. This tripartite structure acts intrinsically as a significant biological barrier, often limiting the permeation of anti-infectives, and so preventing such drugs from reaching their target. Furthermore, identification of the specific permeation-limiting envelope component proves difficult in the case of many anti-infectives, due to the challenges associated with isolation of individual cell envelope structures in bacterial culture. The development of an in vitro permeation model of the Gram-negative inner membrane, prepared by repeated coating of physiologically-relevant phospholipids on Transwell®filter inserts, is therefore reported, as a first step in the development of an overall cell envelope model. Characterization and permeability investigations of model compounds as well as anti-infectives confirmed the suitability of the model for quantitative and kinetically-resolved permeability assessment, and additionally confirmed the importance of employing bacteria-specific base materials for more accurate mimicking of the inner membrane lipid composition - both advantages compared to the majority of existing in vitro approaches. Additional incorporation of further elements of the Gram-negative bacterial cell envelope could ultimately facilitate model application as a screening tool in anti-infective drug discovery or formulation development.