Multi-antigen immunotherapy approaches against Staphylococcus aureus are expected to have the best chance of clinical success when used in combinatorial therapy, potentially incorporating opsonic killing of bacteria and toxin neutralization. We recently reported the development of a murine monoclonal antibody specific for the immunodominant staphylococcal antigen A (IsaA), which showed highly efficient staphylococcal killing in experimental infection models of S. aureus. If IsaA-specific antibodies are to be used as a component of combination therapy in humans, the binding specificity and biological activity of the humanized variant must be preserved. Here, we describe the functional characterization of a humanized monoclonal IgG1 variant designated, hUK-66. The humanized antibody showed comparable binding kinetics to those of its murine parent, and recognized the target antigen IsaA on the surface of clinically relevant S. aureus lineages. Furthermore, hUK-66 enhances the killing of S. aureus in whole blood (a physiological environment) samples from healthy subjects and patients prone to staphylococcal infections such as diabetes and dialysis patients, and patients with generalized artery occlusive disease indicating no interference with already present natural antibodies. Taken together, these data indicate that hUK-66 mediates bacterial killing even in high risk patients and thus, could play a role for immunotherapy strategies to combat severe S. aureus infections.

ABSTRACT Staphylococcus aureus is the most common cause of nosocomial infections. Multiple antibiotic resistance and severe clinical outcomes provide a strong rationale for development of immunoglobulin-based strategies. Traditionally, novel immunological approaches against bacterial pathogens involve antibodies directed against cell surface-exposed virulence-associated epitopes or toxins. In this study, we generated a monoclonal antibody targeting the housekeeping protein IsaA, a suggested soluble lytic transglycosylase of S. aureus , and tested its therapeutic efficacy in two experimental mouse infection models. A murine anti-IsaA antibody of the IgG1 subclass (UK-66P) showed the highest binding affinity in Biacore analysis. This antibody recognized all S. aureus strains tested, including hospital-acquired and community-acquired methicillin-resistant S. aureus strains. Therapeutic efficacy in vivo in mice was analyzed using a central venous catheter-related infection model and a sepsis survival model. In both models, anti-IsaA IgG1 conferred protection against staphylococcal infection. Ex vivo , UK-66P activates professional phagocytes and induces highly microbicidal reactive oxygen metabolites in a dose-dependent manner, resulting in bacterial killing. The study provides proof of concept that monoclonal IgG1 antibodies with high affinity to the ubiquitously expressed, single-epitope-targeting IsaA are effective in the treatment of staphylococcal infection in different mouse models. Anti-IsaA antibodies might be a useful component in an antibody-based therapeutic for prophylaxis or adjunctive treatment of human cases of S. aureus infections.

The increased rates of resistance associated with antibiotic therapy against Staphylococcus aureus, the dominant cause for nosocomial infections has created renewed interest in using alternative treatment options. An antibody-based therapy approach as an emerging option for the prevention and treatment of serious staphylococcal infections is currently under prae-clinical and clinical investigation. In the presented study, the efficacy of passive immunotherapy was directly tested in vivo in two distinct mouse models of catheter-related sepsis and soft tissue infection. The proposed action of monoclonal antibodies is evoked by antibody-mediated phagocytosis. Overall, the results support the idea that implementation of antibodies to S. aureus can play a role for immunotherapy of staphylococci infections in humans.

Functional antibodies targeting IsaA of staphylococcus aureus augment host immune response and open new perspectives for antibacterial therapy ABSTRACT 51 Staphylococcus aureus is the most common cause of nosocomial infections. Multiple 52 antibiotic resistance and severe clinical outcomes provide a strong rationale for 53 development of immunoglobulin-based strategies. Traditionally, novel immunological 54 approaches against bacterial pathogens involve antibodies directed against cell 55 surface exposed virulence-associated epitopes or toxins. In this study, w e generated 56 a monoclonal antibody targeting the housekeeping protein IsaA, a suggested soluble 57 lytic transglycosylase of S. aureus and tested its therapeutic efficacy in two 58 experimental mouse infection models. A murine anti-IsaA antibody of IgG1 subclass 59 (UK-66P) showed highest binding affinity in Biacore analysis. This antibody 60 recognizes all S. aureus strains tested including hospital-acquired and community-61 acquired methicillin-resistant S. aureus strains. Therapeutic efficacy in vivo was 62 analyzed in mice using a central-venous catheter-related infection model and a 63 sepsis survival model. In both models anti-IsaA IgG1 conferred protection to 64 staphylococcal infection. Ex vivo, UK-66P activates professional phagocytes and 65 induces highly microbicidal reactive oxygen metabolites in a dose-dependent 66 manner, resulting in bacterial killing. The study provides proof of concept that 67 monoclonal IgG1 antibodies with high-affinity to the ubiquitous expressed single 68 epitope IsaA are effective in the treatment of staphylococcal infection in different 69 mouse models. Anti-IsaA antibodies might be a useful component in an antibody-70 based therapeutic for prophylaxis or adjunctive treatment of human cases of S.