The antiarrhythmic drugs amiodarone and dronedarone inhibit intoxication of cells with pertussis toxin.

Pertussis toxin (PT) is a virulent factor produced by Bordetella pertussis, the causative agent of whooping cough. PT exerts its pathogenic effects by ADP-ribosylating heterotrimeric G proteins, disrupting cellular signaling pathways. Here, we investigate the potential of two antiarrhythmic drugs, amiodarone and dronedarone, in mitigating PT-induced cellular intoxication. After binding to cells, PT is endocytosed, transported from the Golgi to the endoplasmic reticulum where the enzyme subunit PTS1 is released from the transport subunit of PT. PTS1 is translocated into the cytosol where it ADP-ribosylates inhibitory α-subunit of G-protein coupled receptors (Gαi). We showed that amiodarone and dronedarone protected CHO cells and human A549 cells from PT-intoxication by analyzing the ADP-ribosylation status of Gαi. Amiodarone had no effect on PT binding to cells or in vitro enzyme activity of PTS1 but reduced the signal of PTS1 in the cell suggesting that amiodarone interferes with intracellular transport of PTS1. Moreover, dronedarone mitigated the PT-mediated effect on cAMP signaling in a cell-based bioassay. Taken together, our findings underscore the inhibitory effects of amiodarone and dronedarone on PT-induced cellular intoxication, providing valuable insights into drug repurposing for infectious disease management.

Alpha-1 antitrypsin inhibits Clostridium botulinum C2 toxin, Corynebacterium diphtheriae diphtheria toxin and B. anthracis fusion toxin.

The bacterium Clostridium botulinum, well-known for producing botulinum neurotoxins, which cause the severe paralytic illness known as botulism, produces C2 toxin, a binary AB-toxin with ADP-ribosyltranferase activity. C2 toxin possesses two separate protein components, an enzymatically active A-component C2I and the binding and translocation B-component C2II. After proteolytic activation of C2II to C2IIa, the heptameric structure binds C2I and is taken up via receptor-mediated endocytosis into the target cells. Due to acidification of endosomes, the C2IIa/C2I complex undergoes conformational changes and consequently C2IIa forms a pore into the endosomal membrane and C2I can translocate into the cytoplasm, where it ADP-ribosylates G-actin, a key component of the cytoskeleton. This modification disrupts the actin cytoskeleton, resulting in the collapse of cytoskeleton and ultimately cell death. Here, we show that the serine-protease inhibitor α1-antitrypsin (α1AT) which we identified previously from a hemofiltrate library screen for PT from Bordetella pertussis is a multitoxin inhibitor. α1AT inhibits intoxication of cells with C2 toxin via inhibition of binding to cells and inhibition of enzyme activity of C2I. Moreover, diphtheria toxin and an anthrax fusion toxin are inhibited by α1AT. Since α1AT is commercially available as a drug for treatment of the α1AT deficiency, it could be repurposed for treatment of toxin-mediated diseases.

Domperidone Inhibits Clostridium botulinum C2 Toxin and Bordetella pertussis Toxin.

Bordetella pertussis toxin (PT) and Clostridium botulinum C2 toxin are ADP-ribosylating toxins causing severe diseases in humans and animals. They share a common translocation mechanism requiring the cellular chaperones Hsp90 and Hsp70, cyclophilins, and FK506-binding proteins to transport the toxins' enzyme subunits into the cytosol. Inhibitors of chaperone activities have been shown to reduce the amount of transported enzyme subunits into the cytosol of cells, thus protecting cells from intoxication by these toxins. Recently, domperidone, an approved dopamine receptor antagonist drug, was found to inhibit Hsp70 activity. Since Hsp70 is required for cellular toxin uptake, we hypothesized that domperidone also protects cells from intoxication with PT and C2. The inhibition of intoxication by domperidone was demonstrated by analyzing the ADP-ribosylation status of the toxins' specific substrates. Domperidone had no inhibitory effect on the receptor-binding or enzyme activity of the toxins, but it inhibited the pH-driven membrane translocation of the enzyme subunit of the C2 toxin and reduced the amount of PTS1 in cells. Taken together, our results indicate that domperidone is a potent inhibitor of PT and C2 toxins in cells and therefore might have therapeutic potential by repurposing domperidone to treat diseases caused by bacterial toxins that require Hsp70 for their cellular uptake.