The pathogens resistance to numerous antibiotics, including new drugs, leaves few therapeutic options available for infected patients, who often are too sick to fight the infection themselves

The pathogens resistance to numerous antibiotics, including new drugs, leaves few therapeutic options available for infected patients, who often are too sick to fight the infection themselves. inoculated with a mixture of CR CPS conjugated to anthrax protective antigen. Two of these MAbs, 17H12 and 8F12, bind whole and oligosaccharide epitopes of the CPS of clade 2 ST258 CR infections in the United States. These antibodies were shown to agglutinate all clade 2 strains and were also shown to promote extracellular processes killing these bacteria, including biofilm inhibition, complement deposition, and deployment of neutrophil extracellular traps. Additionally, they promoted opsonophagocytosis and OTS964 intracellular killing of CR by human-derived neutrophils and cultured murine macrophages. Finally, when mice were intratracheally infected with preopsonized clade 2 CR are possible. Such MAbs and vaccines would benefit those susceptible populations at risk of infection with this group of multidrug-resistant bacteria. KEYWORDS: adjuvant therapy, antibiotic resistance, carbapenem resistance, infection protection, is an enteric bacterium that has been responsible for an increasing number of deadly outbreaks and hospital-acquired infections. The pathogens resistance to numerous antibiotics, including new drugs, leaves few therapeutic options available for infected patients, who often are too sick to OTS964 fight the infection themselves. Immunotherapy utilizing monoclonal antibodies has been successful in other medical fields, and antibodies targeting the outer polysaccharide capsule of these bacteria could be a valuable treatment alternative. This study presents two anticapsular antibodies, 17H12 and 8F12, that were found to be protective against the most virulent carbapenem-resistant clinical strains. These antibodies are shown to promote the killing of these strains through several extracellular and intracellular processes and prevent the spread of infection in mice from the lungs to distal organs. Thus, they could ultimately treat or protect patients infected or at risk of infection by this multidrug-resistant bacterium. INTRODUCTION The Gram-negative bacterium presents a dire health care problem. Already a common nosocomial pathogen that causes chronic urinary tract infections, pneumonias, and sepsis, the pathogen has become even more successful in the last decade because of emerging multidrug resistance to antibiotics. Infections with these strains are associated with mortality rates of >50%, particularly in hospitalized patients with comorbidities (1,C3). Additionally, recent reports of carbapenem-resistant (CR) strains that have acquired a hypervirulent phenotype indicate that these strains OTS964 could soon also cause disease in healthy people in the community (4). With reports of already emerging resistance to ceftazidime-avibactam (5) and limited development of other Gram-negative antibiotics (6), novel strategies to combat CR are urgently needed. Though still an emerging field, treatment of infections with antibodies has shown some success, and to date, four such monoclonal antibodies (MAbs) are FDA licensed (7). Numerous laboratories, including ours, have demonstrated protective efficacy of anti-infective antibodies in murine models (8,C12). As MAbs target specific pathogens, they are less likely to disturb the microbiome. Yet the specificity of antibodies also drives antigenic variability of pathogens and can make finding appropriate targets the most difficult hurdle in MAb development. While carbohydrate antigens, such as the antiphagocytic capsular polysaccharide (CPS), are the most accessible targets on the surface of and other OTS964 members of the family strains and has been subcategorized into two evolutionary groups, termed clades 1 and 2 (13). Infection with clade 2 strains, which encompasses about 50% of the United States ST258 strains (1) and up to 88% of the ST258 strains in other countries, has been associated with a higher mortality rate than infection with clade 1 strains (1, 14,C16). Notably, the 2009 2009 outbreak at the National Institutes of Health Clinical Center in Bethesda, which resulted in nine deaths, was also caused by a clade 2 strain (17). Genetic differentiation of these two clades results from DNA recombination events in an ~215-kb genomic region that includes the gene cluster involved in CPS biosynthesis (13). Molecular typing of the gene alleles, which has replaced serological typing, indicates higher conservation of CPS in clade 2 strains than in clade 1 strains, as nearly all clade 2 strains possess the allele (18). We previously developed an IgM antibody, 1C9, that was shown to agglutinate 16/16 clade 2 strains tested and 1/7 clade 1 strains. However, as an IgM isotype, its affinity was too low and attempts to switch the isotype of this antibody failed (18). In this study, we developed novel IgG antibodies by immunizing mice with protein-conjugated CPS, which improved the probability of generating IgG-producing hybridomas. We now present data showing that IgG3s 17H12 and 8F12, which bind to clade 2 CPS, exhibit broad opsonophagocytic killing efficacy against clade 2 strains and demonstrate protective efficacy in a murine intratracheal (i.t.) infection model. Interestingly, both MAbs bind to the same glycan epitope as 1C9, but both MAbs can simultaneously bind to the clade 2 CPS. In IGKC summary, we propose that 17H12.