Mechanisms of Protection against Clostridium difficile Infection by the Monoclonal Antitoxin Antibodies Actoxumab and Bezlotoxumab

Abstract

dClostridium difficile infection (CDI) represents the most prevalent cause of antibiotic-associated gastrointestinal infections in health care facilities in the developed world. Disease symptoms are caused by the two homologous exotoxins, TcdA and TcdB. Standard therapy for CDI involves administration of antibiotics that are associated with a high rate of disease recurrence, highlighting the need for novel treatment paradigms that target the toxins rather than the organism itself. A combination of human monoclonal antibodies, actoxumab and bezlotoxumab, directed against TcdA and TcdB, respectively, has been shown to decrease the rate of recurrence in patients treated with standard-of-care antibiotics. However, the exact mechanism of antibodymediated protection is poorly understood. In this study, we show that the antitoxin antibodies are protective in multiple murine models of CDI, including systemic and local (gut) toxin challenge models, as well as primary and recurrent models of infection in mice. Systemically administered actoxumab-bezlotoxumab prevents both the damage to the gut wall and the inflammatory response, which are associated with C. difficile in these models, including in mice challenged with a strain of the hypervirulent ribotype 027. Furthermore, mutant antibodies (N297Q) that do not bind to Fc␥ receptors provide a level of protection similar to that of wild-type antibodies, demonstrating that the mechanism of protection is through direct neutralization of the toxins and does not involve host effector functions. These data provide a mechanistic basis for the prevention of recurrent disease observed in CDI patients in clinical trials. Clostridium difficile is an anaerobic, spore-forming, Gram-positive bacterium that causes infections in the lumen of the colon and is the most frequent cause of nosocomial diarrhea in the developed world (1, 2). C. difficile infections (CDI) contribute to thousands of deaths and are associated with over $1 billion in health care-related costs in the United States each year (3-5). The symptoms of CDI range from asymptomatic carriage or mild diarrhea to fatal pseudomembranous colitis, colonic rupture, and death (6, 7). The disease occurs mainly in patients undergoing (or who have recently undergone) a course of broad-spectrum antibiotics; in such patients, composition of the gut microbiota is altered, disrupting the body's natural defense against C. difficile infections. Clinical management of CDI consists of discontinuation of the offending antibiotic and treatment with either metronidazole, vancomycin, or the newly approved fidaxomicin (8). A major concern with CDI is that even when treatment of a primary infection is successful, 20 to 30% of patients experience a recurrence of the disease within days or weeks of symptom resolution. Disease recurrence results from continued disruption of the gut microbiota by standard-of-care antibiotics (9) combined with persistence of resistant C. difficile spores (relapse) or reacquisition of new spores from the environment (rein...

Broad Coverage of Genetically Diverse Strains of Clostridium difficile by Actoxumab and Bezlotoxumab Predicted by In Vitro Neutralization and Epitope Modeling

“…To better understand how in vitro potency can predict in vivo clinical efficacy, it is relevant to correlate in vitro neutralization of specific strains with in vivo clinical efficacy against these same strains. In this regard, actoxumab-bezlotoxumab was efficacious against ribotype 027 strains in a gnotobiotic piglet infection model (22) and in mouse models of primary and recurrent CDIs (23). Furthermore, the antibodies reduced the rate of recurrent CDIs in patients infected with ribotype 027 strains to the same extent as in the overall patient population (although the reduction did not reach statistical significance; P ϭ 0.06) (7).…”

Adaptive immune response to Clostridium difficile infection: A perspective for prevention and therapy

“…Bezlotoxumab was confirmed to bind to epitopes in the carbohydrate-binding domain of TcdB, possibly explaining its efficacy [40]. In mouse models, bezlotoxumab mutated in the Fcγ receptor binding site was as protective as unmodified Ab, suggesting that direct toxin neutralization is likely the mechanism of protection in CDI [41]. In contrast, the failure of actoxumab in human trials provides additional evidence that TcdB is more damaging in human disease, and also may explain the discrepancies in earlier studies that found conflicting reports of whether naturally occurring anti-TcdA antibodies protect against CDI.…”

The Impact of Therapeutic Antibodies on the Management of Digestive Diseases: History, Current Practice, and Future Directions

“…C. difficile infection can complicate the use of broad-spectrum antibiotics, causing a severe infectious toxigenic colitis rising in incidence and incurring significant morbidity and mortality [101]. Human monoclonal antibodies that neutralize C. difficile toxins A and B, lower TNFa levels and improve survival in preclinical mouse models are currently under investigation [102][103][104]. A phase II study compared treatment with monoclonal antibodies against toxins A and B combined with standard antibiotic treatment to antibiotic treatment alone.…”