Pathway to Clostridium difficile Treatment Stalled by New Discovery

Yongrong Zhang, PhD

Yongrong Zhang, PhD

One of the 2 triggering toxins in Clostridium difficile (C. difficile) Toxin B (TcdB) regulates both the cytotoxicity and proinflammatory activity, according to a new study.

Researchers from the University of Maryland in Baltimore studied a mouse model, human tissues and immune cells in order to understand the inflammatory response to various C. difficile toxins. Toxin A (TcdA) and TcdB are the primary drivers of the C. difficile bacterium and previously, not much was understood about how they regulate their biological functions, the researchers told MD Magazine. Generally, TcdA is more inflammatory while TcdB is more cytotoxic.

The investigators first studied TcdB in the animal model, where it reduced cytotoxicity but enhanced the proinflammatory activities. Then, using the human gut cells and immune cells, the researchers observed a mutant TcdB more potent than a wild-type. They saw the mutant TcdB compared to the wild-type TcdB significantly up-regulated the production of several pro-inflammatory cytokines in the gut tissue. They believe this indicates that the toxins could also regulate the inflammatory response during C. difficile infection manifestation in humans.

"We were surprised to identify an unexpected function of the cysteine protease-mediated autoprocessing in that it regulates the toxin-mediated acute inflammatory response in both human and animal models," study author Yongrong Zhang, PhD told MD Magazine "Blocking the autoprocessing of TcdB by chemical inhibitor or point mutations, while reducing the toxin cytotoxicity, significantly enhanced its proinflammatory activities in animal intestines, human intestinal tissues and human immune cells. Our study demonstrates a novel molecular mechanism of toxin regulation, in that the cysteine protease domains act as an internal regulator to fine tune the different activities of C. difficile toxins: it positively regulates toxins’ cytotoxicity, but negatively regulates their proinflammatory activity."

This new knowledge of the disease's molecular pathogenesis means that researchers can develop new therapeutics against the disease, the researchers continued.

"For example, developing small inhibitors against cysteine protease domain of the toxins as therapeutic is more complicated than previously thought and treatments against C. difficile infection by targeting the cysteine protease domain of C. difficile toxins may not be an optimal choice," Zhang continued.

In a related editorial, Xinhua Chen, PhD and Ciaran P. Kelly, MD from Harvard Medical School wrote that the study may have uncovered a roadblock to traditional targeted therapies.

"This study suggests a dual role of cysteine enzyme domain (CPD)-mediated autoprocessing, which regulates the relative outcomes of cytotoxicity and proinflammatory induction," they said. "There have been suggestions that CPD inhibition could trap TcdB in the endosome, thereby mitigating cytotoxic effects. However, this study suggests that there may be an unintended consequence to CPD inhibition in the form of increased inflammatory pathway activation. The question remains, however, which of the 2 components, cytotoxicity or proinflammatory responses, contribute more to disease pathogenesis."

Both the study authors and the authors of the editorial agree that more research is needed to confirm these findings and their implications.

The research "exposes a new and fascinating aspect to our understanding of the virulence of this difficult bacterium," Chen and Kelly concluded.

The study, titled "Cysteine Protease-Mediated Autocleavage of Clostridium difficile Toxins Regulates Their Proinflammatory Activity," was published in the journal Cellular and Molecular Gastroenterology and Hepatology.

The related editorial is here.