doi: 10.1586/17476348.2014.940323. age, broad-spectrum antibiotic exposure, prolonged ventilation, and immunosuppression are most susceptible to these infections (3). To combat the increase in antibiotic-resistant strains, passive immunization with a monoclonal antibody (MAb) targeting the invading pathogen or its virulence factor(s) is being explored as an alternative strategy for protecting at-risk populations (4,C7). One virulence factor under investigation as a target for new therapeutic options against disease is usually alpha-toxin (AT). AT is usually a secreted protein that binds ADAM10 (a disintegrin and metalloproteinase 10) on cell membranes and oligomerizes to form heptameric transmembrane pores (8, 9). AT can directly lyse cells, and it has been demonstrated to exert other toxic effects at sublytic concentrations. For example, AT pore formation on macrophage membranes activates the NLRP3 inflammasome, which, along with staphylococcal pathogen-associated molecular patterns (PAMPs), induces interleukin 1 (IL-1) secretion and promotes cell death (10, 11). AT also activates ADAM10-mediated proteolysis of E-cadherin in cell-cell adhesive contacts, contributing to epithelial and endothelial damage Mesaconine (9, 12, 13). Therefore, targeted AT inhibition may neutralize multiple pathogenic mechanisms, effectively disarming the and enabling the host to combat the infection. MEDI4893 is an extended-half-life, high-affinity, AT-neutralizing MAb under development for the prevention of nosocomial pneumonia in high-risk patients (in a study registered at www.clinicaltrialsregister.eu under registration no. 2014-001097-34). MEDI4893 was generated by introducing the YTE mutations into the previously reported anti-AT Mesaconine MAb LC10, to extend the antibody half-life (4, 14, 15). LC10, also known as MEDI4893*, is identical to MEDI4893 except for the absence Rabbit Polyclonal to CHST10 of the YTE mutations in the Fc domain name (16). While the YTE mutations increase IgG half-lives in humans, they significantly reduce serum exposure in mice and preclude the use of MEDI4893 in murine models (17, 18). Therefore, preclinical animal testing is conducted with Mesaconine MEDI4983*. Mesaconine MEDI4893* was shown to neutralize AT and to promote survival in an acute pneumonia model when administered prophylactically to mice (4). To date, all reported preclinical testing with MEDI4893* has been conducted in immunocompetent animals. However, it is likely that some of the high-risk patients targeted in the MEDI4893 clinical studies will be immunocompromised. In the present study, we tested MEDI4893* in an immunocompromised murine pneumonia model. Herein, we report that MEDI4893* prophylaxis preserves airway structure and the air-liquid barrier, ultimately leading to increased survival rates in this model. MEDI4893* also provides benefits over linezolid or vancomycin monotherapy and extends the therapeutic treatment windows of both drugs, making it a promising candidate for prophylaxis or adjunctive treatment of pneumonia in immunocompromised patients. MATERIALS AND METHODS Bacterial strains and chemicals. NRS382 (type USA100, clonal complex 5 [CC5]) and NRS261 (CC30) were obtained from the Network on Antimicrobial Resistance in (NARSA). SF8300 (type USA300) was generously provided by Binh An Diep (University of California, San Francisco). All strains produced AT, as measured in overnight culture supernatants (at 0.730, 1.273, and 3.44 g/ml, respectively). NRS261 and SF8300 contain the genes encoding Panton-Valentine leucocidin. Bacteria were produced to an optical density at 600 nm (OD600) of 0.8 in trypticase soy broth (TSB) (VWR International), washed twice in ice-cold phosphate-buffered saline (PBS) (Life Technologies), and frozen as aliquots in TSB with 10% glycerol. Challenge inocula were prepared from one frozen vial for each experiment, diluted in ice-cold PBS, and placed on ice until used for contamination. Vancomycin (Sigma-Aldrich) was prepared in.