Antibiotic Adjuvant Activity Revealed in a Photoaffinity Approach to Determine the Molecular Target of Antipyocyanin Compounds.

Infections with Pseudomonas aeruginosa are a looming threat to public health. New treatment strategies are needed to combat this pathogen, for example, by blocking the production of virulence factors like pyocyanin. A photoaffinity analogue of an antipyocyanin compound was developed to interrogate the inhibitor's molecular mechanism of action. While we sought to develop antivirulence inhibitors, the proteomics results suggested that the compounds had antibiotic adjuvant activity. Unexpectedly, we found that these compounds amplify the bactericidal activity of colistin, a well-characterized antibiotic, suggesting they may represent a first-in-class antibiotic adjuvant therapy. Analogues have the potential not only to widen the therapeutic index of cationic antimicrobial peptides like colistin, but also to be effective against colistin-resistant strains, strengthening our arsenal to combat P. aeruginosa infections.

Mesenchymal transformation in epithelial ovarian tumor cells expressing epidermal growth factor receptor variant III.

Overexpression of the epidermal growth factor (EGF) receptor occurs frequently in ovarian cancer and is associated with poor patient prognosis. A constitutively active mutant EGF receptor termed variant III (EGFRvIII) has been detected at a high frequency in many human tumors, including those of the ovary. To identify the consequences of EGFRvIII expression in ovarian tumor cells, we introduced EGFRvIII into the epithelial ovarian cancer cell line (OVCA 433). The EGFRvIII-transfected cells displayed a dissociated, motile phenotype and fibroblastic morphology. The EGFRvIII-dependent phenotype was comparable to that observed in EGF-stimulated parental OVCA 433 cultures and required the catalytic activity of the mutant receptor. Disruption of adherens and desmosomal junctions in EGFRvIII expressing cells was evident by immunofluorescent detection of specific junctional components. In addition, Western blot analysis confirmed decreased levels of cellular plakoglobin and beta-catenin in EGFRvIII-expressing cells, and E-cadherin protein and mRNA were nearly absent. The loss of E-cadherin was accompanied by decreased expression of additional ovarian epithelial markers, including keratins 7, 8, and 18 and mucins 1 and 4. In contrast, the mesenchymal markers N-cadherin and vimentin were elevated in EGFRvIII expressing cells. Overall, the switch in cadherins from E-cadherin to N-cadherin, coupled with gain of vimentin expression and loss of the epithelial keratins and mucins typically expressed in well-differentiated epithelial ovarian carcinomas, are consistent with transition to a mesenchymal phenotype as an outcome of EGFRvIII expression. These findings suggest that EGFRvIII expression may regulate phenotypic plasticity in ovarian cancer and thereby contribute to more aggressive disease.