Triple-edged sword; multiple modes of action of a monoclonal antibody against the multi-drug resistant Escherichia coli clone ST131-H30 [PublishAheadOfPrint]

The multi-drug resistant extra-intestinal pathogenic Escherichia coli clone, ST131-H30 has spread worldwide. This clone expresses a conserved LPS O-antigen, O25b. Previously, we described monoclonal antibodies (mAbs) specific to the O25b antigen and characterized them as diagnostic and therapeutic tools. In this study evidence is provided that besides the previously shown complement mediated bactericidal effect, an O25b-specific humanized mAb, A1124 also enhances opsonophagocytic uptake by the murine macrophage cell line RAW264.1. Both phagocyte-dependent and -independent killing, triggered by A1124, was confirmed in human whole blood. Furthermore, A1124 was shown to neutralize endotoxin activity of purified or naturally shed LPS of clinical isolates. This activity was demonstrated in vitro using both RAW264.7 cells and a human TLR4-reporter cell-line, as well as in a murine model of endotoxemia using purified LPS for challenge. Significant protective efficacy of A1124 at low doses (< 1 mg/kg) was shown in murine and rat models of bacteremia. The contribution of the bactericidal and anti-inflammatory effects was dissected in the mouse bacteremia model through depletion of complement with cobra venom factor (CVF). Protective efficacy was lost in complement-depleted mice, suggesting the essential role of complement-mediated activities for protection in this model. These data suggest that A1124 exhibits different mechanisms of action, namely direct complement-mediated and opsonophagocytic killing as well as endotoxin neutralization in various challenge models. Which of these activities are the most relevant in a clinical setting will need to be addressed by future translational studies.

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