Bispecific antibody targets multiple Pseudomonas aeruginosa evasion mechanisms in the lung vasculature
Ajitha Thanabalasuriar, … , Antonio DiGiandomenico, Paul Kubes
Ajitha Thanabalasuriar, … , Antonio DiGiandomenico, Paul Kubes
Published May 2, 2017
Citation Information: J Clin Invest. 2017;127(6):2249-2261. https://doi.org/10.1172/JCI89652.
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Research Article Immunology Infectious disease

Bispecific antibody targets multiple Pseudomonas aeruginosa evasion mechanisms in the lung vasculature

Abstract

Pseudomonas aeruginosa is a major cause of severe infections that lead to bacteremia and high patient mortality. P. aeruginosa has evolved numerous evasion and subversion mechanisms that work in concert to overcome immune recognition and effector functions in hospitalized and immunosuppressed individuals. Here, we have used multilaser spinning-disk intravital microscopy to monitor the blood-borne stage in a murine bacteremic model of P. aeruginosa infection. P. aeruginosa adhered avidly to lung vasculature, where patrolling neutrophils and other immune cells were virtually blind to the pathogen’s presence. This cloaking phenomenon was attributed to expression of Psl exopolysaccharide. Although an anti-Psl mAb activated complement and enhanced neutrophil recognition of P. aeruginosa, neutrophil-mediated clearance of the pathogen was suboptimal owing to a second subversion mechanism, namely the type 3 secretion (T3S) injectisome. Indeed, T3S prevented phagosome acidification and resisted killing inside these compartments. Antibody-mediated inhibition of the T3S protein PcrV did not enhance bacterial phagocytosis but did enhance killing of the few bacteria ingested by neutrophils. A bispecific mAb targeting both Psl and PcrV enhanced neutrophil uptake of P. aeruginosa and also greatly increased inhibition of T3S function, allowing for phagosome acidification and bacterial killing. These data highlight the need to block multiple evasion and subversion mechanisms in tandem to kill P. aeruginosa.

Authors

Ajitha Thanabalasuriar, Bas G.J. Surewaard, Michelle E. Willson, Arpan S. Neupane, Charles K. Stover, Paul Warrener, George Wilson, Ashley E. Keller, Bret R. Sellman, Antonio DiGiandomenico, Paul Kubes

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Figure 7

Schematic of pulmonary neutrophil phagocytosis of P. aeruginosa in the presence of MEDI3902.

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Schematic of pulmonary neutrophil phagocytosis of P. aeruginosa in the p...
WT P. aeruginosa avoids detection/phagocytosis by neutrophils in the pulmonary capillaries. P. aeruginosa surface Psl exopolysaccharide inhibits deposition of complement C3b on its surface, cloaking itself from neutrophil recognition. Moreover, phagocytized P. aeruginosa secrete effector molecules via the T3S injectisome to hinder acidification of the neutrophil phagolysosome. Deletion of Psl expression (ΔpslA) or targeting of Psl with an anti-Psl mAb increases bacterial recognition and phagocytosis by neutrophils in the lung. However, phagocytized bacteria interfere with acidification of the phagolysosomal compartment. Inhibition of the T3S injectisome, either by infection of mice with T3S injectisome–deficient P. aeruginosa (ΔpcrV) or by treatment with an anti-PcrV mAb alone, does not increase recognition or phagocytosis by neutrophils, but does lead to greater acidification of phagolysosomes in the few neutrophils that engulf bacteria. In contrast, administration of bispecific antibody MEDI3902, which simultaneously targets Psl and PcrV, leads to increased recognition, phagocytosis, and killing of bacteria by neutrophils. This enhanced activity is not observed with a mixture of parental anti-Psl and anti-PcrV mAbs, indicating the necessity of including both specificities on the same mAb molecule. Therefore, the anti-Psl targeting mediated by MEDI3902 facilitates transfer of anti–T3S injectisome activity (via the anti-PcrV arm) within the phagolysosome after phagocytosis, resulting in enhanced compartment acidification and killing of P. aeruginosa.