A novel fusion protein containing the receptor binding domains of C. difficile toxin A and toxin B elicits protective immunity against lethal toxin and spore challenge in preclinical efficacy models

Tian, Jing-Hui; Fuhrmann, Steven R.; Kluepfel-Stahl, Stefanie; Carman, Robert J.; Ellingsworth, Larry; Flyer, David C.

doi:10.1016/j.vaccine.2012.04.045

Cited by 59 publications

(54 citation statements)

References 38 publications

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“…Mutations that abolished the ADP-ribosylation activity of DT resulted in a toxin that was incapable of inducing caspase activity (28). Similarly, mutations that led to the loss of the glucosyltransferase activities of TcdA and TcdB resulted in toxins that failed to induce caspase activity (24,34,(37)(38)(39). The data strongly suggest that the biological and enzymatic activities of the toxins are primarily responsible for caspase induction that results in the loss of viability and in cell death.…”

Section: Discussionmentioning

confidence: 91%

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Caspase Activation as a Versatile Assay Platform for Detection of Cytotoxic Bacterial Toxins

et al. 2013

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Pathogenic bacteria produce several virulence factors that help them establish infection in permissive hosts. Bacterial toxins are a major class of virulence factors and hence are attractive therapeutic targets for vaccine development. Here, we describe the development of a rapid, sensitive, and high-throughput assay that can be used as a versatile platform to measure the activities of bacterial toxins. We have exploited the ability of these toxins to cause cell death via apoptosis of sensitive cultured cell lines as a readout for measuring toxin activity. Caspases (cysteine-aspartic proteases) are induced early in the apoptotic pathway, and so we used their induction to measure the activities of Clostridium difficile toxins A (TcdA) and B (TcdB) and binary toxin (CDTaCDTb), Corynebacterium diphtheriae toxin (DT), and Pseudomonas aeruginosa exotoxin A (PEA). Caspase induction in the cell lines, upon exposure to toxins, was optimized by toxin concentration and intoxication time, and the specificity of caspase activity was established using a genetically mutated toxin and a pan-caspase inhibitor. In addition, we demonstrate the utility of the caspase assay for measuring toxin potency, as well as neutralizing antibody (NAb) activity against C. difficile toxins. Furthermore, the caspase assay showed excellent correlation with the filamentous actin (F-actin) polymerization assay for measuring TcdA and TcdB neutralization titers upon vaccination of hamsters. These results demonstrate that the detection of caspase induction due to toxin exposure using a chemiluminescence readout can support potency and clinical immunogenicity testing for bacterial toxin vaccine candidates in development.

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Section: Discussionmentioning

confidence: 91%

“…Vero cells have been shown to be very sensitive to C. difficile toxins and have been used in a published F-actin polymerization assay (19,29,33,34). Since we compared the caspase and F-actin polymerization assay, Vero cells were selected for our caspase studies for TcdA and TcdB toxins.…”

Section: Discussionmentioning

confidence: 99%

Caspase Activation as a Versatile Assay Platform for Detection of Cytotoxic Bacterial Toxins

et al. 2013

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“…A recombinant fusion protein comprising the receptor binding domains of both TcdA and TcdB has been shown to reduce disease severity and confer antibody protection in a hamster model of CDI [Tian et al 2012].…”

Section: Parenteral Passive Immunotherapymentioning

confidence: 99%

The host immune response to Clostridium difficile infection

Solomon

2013

Therapeutic Advances in Infection

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Clostridium difficile infection (CDI) is the most common infectious cause of healthcare-acquired diarrhoea. Outcomes of C. difficile colonization are varied, from asymptomatic carriage to fulminant colitis and death, due in part to the interplay between the pathogenic virulence factors of the bacterium and the counteractive immune responses of the host.Secreted toxins A and B are the major virulence factors of C. difficile and induce a profound inflammatory response by intoxicating intestinal epithelial cells causing proinflammatory cytokine release. Host cell necrosis, vascular permeability and neutrophil infiltration lead to an elevated white cell count, profuse diarrhoea and in severe cases, dehydration, hypoalbuminaemia and toxic megacolon. Other bacterial virulence factors, including surface layer proteins and flagella proteins, are detected by host cell surface signal molecules that trigger downstream cell-mediated immune pathways.Human studies have identified a role for serum and faecal immunoglobulin levels in protection from disease, but the recent development of a mouse model of CDI has enabled studies into the precise molecular interactions that trigger the immune response during infection. Key effector molecules have been identified that can drive towards a protective anti-inflammatory response or a damaging proinflammatory response.The limitations of current antimicrobial therapies for CDI have led to the development of both active and passive immunotherapies, none of which have, as yet been formally approved for CDI. However, recent advances in our understanding of the molecular basis of host immune protection against CDI may provide an exciting opportunity for novel therapeutic developments in the future.

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“…In particular, RBDs of TcdA and TcdB have been evaluated for their ability to induce protective immunity. 25,[32][33][34] Recent studies have indicated that the N-terminal GT domain of TcdB can serve as an excellent immunogen. 35,36 This notion was initially supported by our recent construction of a chimeric recombinant vaccine against TcdA and TcdB, i.e., cTxAB, in which the original RBD of a full-length TcdB was replaced with the corresponding portion of TcdA.…”

Section: Introductionmentioning

confidence: 99%

“…32 Moreover, the N terminus of TcdB is more conserved than its RBD. 38,39 In contrast, the RBD of TcdA has potent adjuvant activity, e.g., it has been reported that a TcdA receptor peptide fragment enhanced mucosal antibody responses in mice when coadministered with heterologous protective antigens 40 .…”

Section: Introductionmentioning

confidence: 99%

A chimeric protein comprising the glucosyltransferase and cysteine proteinase domains of toxin B and the receptor binding domain of toxin A induces protective immunity againstClostridium difficileinfection in mice and hamsters

Wang

Yan

Kim

et al. 2015

Human Vaccines & Immunotherapeutics

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A novel fusion protein containing the receptor binding domains of C. difficile toxin A and toxin B elicits protective immunity against lethal toxin and spore challenge in preclinical efficacy models

Cited by 59 publications

References 38 publications

Caspase Activation as a Versatile Assay Platform for Detection of Cytotoxic Bacterial Toxins

Caspase Activation as a Versatile Assay Platform for Detection of Cytotoxic Bacterial Toxins

The host immune response to Clostridium difficile infection

A chimeric protein comprising the glucosyltransferase and cysteine proteinase domains of toxin B and the receptor binding domain of toxin A induces protective immunity againstClostridium difficileinfection in mice and hamsters

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