Human antibody against shiga toxin 2 administered to piglets after the onset of diarrhea due to Escherichia coli O157:H7 prevents fatal systemic complications

Abstract

Infection of children with Shiga toxin (Stx)-producing Escherichia coli (STEC) can lead to hemolytic-uremic syndrome (HUS) in 5 to 10% of patients. Stx2, one of two toxins liberated by the bacterium, is directly linked with HUS. We have previously shown that Stx-specific human monoclonal antibodies protect STEC-infected animals from fatal systemic complications. The present study defines the protective antibody dose in relation to the time of treatment after the onset of diarrhea in infected gnotobiotic piglets. Using the mouse toxicity model, we selected 5C12, an antibody specific for the A subunit, as the most effective Stx2 antibody for further characterization in the piglet model in which piglets developed diarrhea 16 to 40 h after bacterial challenge, followed by fatal neurological symptoms at 48 to 96 h. Seven groups of piglets received doses of 5C12 ranging from 6.0 mg/kg to 0.05 mg/kg of body weight, administered parenterally 48 h after bacterial challenge. The minimum fully protective antibody dose was 0.4 mg/kg, and the corresponding serum antibody concentration in these piglets was 0.7 mug (+/-0.5)/ml, measured 7 to 14 days after administration. Of 40 infected animals which received Stx2 antibody treatment of > or =0.4 mg/kg, 34 (85%) survived, while only 1 (2.5%) of 39 placebo-treated animals survived. We conclude that the administration of the Stx2-specific antibody was protective against fatal systemic complications even when it was administered well after the onset of diarrhea. These findings suggest that children treated with this antibody, even after the onset of bloody diarrhea, may be equally protected against the risk of developing HUS.

FIG. 1.

Neutralization of Stx2-mediated HeLa cell cytotoxicity by Stx2-specific HuMAbs. All Stx2-specific HuMAbs neutralized Stx2 at the highest concentration of 5 μg/ml and showed dose dependency. The HuMAb 5C12 was the best neutralizing HuMAb as it neutralized cytotoxicity by about 40% at the lowest tested concentration (8 ng/ml).

FIG. 2.

Percent survival of mice given i.p. 50, 35, 20, 10, or 5 μg of HuMAbs 3E9, 2F10, 5C12, 5H8, or 6G3 followed 18 h later with i.p. administration of a 250% lethal dose of Stx2. Mice in PBS and IgG1(κ) control groups died within 3 days of Stx2 injection. All HuMAbs protected mice but showed dose dependency except 5C12, which protected 100% of the mice even at the lowest dose administered (5 μg/mouse).