Development of a Vaccine Against Clostridium difficile Infection (CDI): Design, Purification, and Biological Activities of the Recombinant Toxin 1 Antigens

摘要

Clostridium difficile is the major cause of antibiotic-associated diarrhea and pseudomembranous colitis, a disease associated with significant morbidity and mortality. The disease is mostly of nosocomial origin, with elder patients undergoing anti-microbial therapy being particularly at risk. A new, hypervirulent strain of C. difficile called NAP1 (027) has been implicated in outbreaks associated with increased morbidity and mortality since the early 2000s. This epidemic strain is responsible for increased incidence of C. difficile-associated diarrhea related not only to antibiotic exposure, but infection is also associated with Gl surgery, prolonged hospitalization, and immune-compromising conditions. C. difficile produces 2 large toxins: Toxin A and Toxin B. These 2 toxins act synergistically to damage and impair the colonic epithelium and are primarily responsible for the pathogenesis associated with C. difficile infection (CDI). Testing the feasibility of toxin-based vaccination against C. difficile is being investigated. A native Toxoid A- and B-based vaccine was reported to be safe and immunogenic in healthy volunteers. We generated a toolbox of E. coli expressed Toxin B fragments covering the entire molecule and systematically explored these fragments as components of an experimental vaccine. We observed a robust immune response in hamsters vaccinated with the recombinant toxin fragments. The antiserum obtained from immunized hamsters was shown to neutralize cytotoxic effects of Toxin B in vitro (in cell-based neutralization assay). Hamsters immunized with the combination of full-length Toxoid A and fragments of Toxin B were protected against lethal challenge with C. difficile spores. We also evaluated that recombinant full-length mTcdA and mTcdB vaccine and demonstrated excellent performance in a hamster challenge model, equivalent to benchmark toxoid vaccine. The use of recombinant C. difficile toxins could afford improved protection, vaccine thermal stability, and facilitate manufacturability of the vaccine. The additional studies, including thermal stability, formulation, safety, and dosing, will be needed to establish the final vaccine image.