Isofagomine Inhibits Multiple TcdB Variants and Protects Mice from Clostridioides difficile-Induced Mortality.

Clostridioides difficile causes life-threatening diarrhea and is one of the leading causes of nosocomial infections. During infection, C. difficile releases two gut-damaging toxins, TcdA and TcdB, which are the primary determinants of disease pathogenesis and are important therapeutic targets. Once in the cytosol of mammalian cells, TcdA and TcdB use UDP-glucose to glucosylate host Rho GTPases, which leads to cytoskeletal changes that result in a loss of intestinal integrity. Isofagomine inhibits TcdA and TcdB as a mimic of the glucocation transition state of the glucosyltransferase reaction. However, sequence variants of TcdA and TcdB across the clades of infective C. difficile continue to be identified, and therefore, evaluation of isofagomine inhibition against multiple toxin variants is required. Here, we show that isofagomine inhibits the glucosyltransferase domain of multiple TcdB variants and protects TcdB-induced cell rounding of the most common full-length toxin variants. Furthermore, we demonstrate that isofagomine protects against C. difficile-induced mortality in two murine models of C. difficile infection. Isofagomine treatment of mouse C. difficile infection also permitted the recovery of the gastrointestinal microbiota, an important barrier to preventing recurring C. difficile infection. The broad specificity of isofagomine supports its potential as a prophylactic to protect against C. difficile-induced morbidity and mortality.

Isofagomine inhibits multiple TcdB variants and protects mice from Clostridioides difficile induced mortality.

Clostridioides difficile causes life-threatening diarrhea and is the leading cause of healthcare associated bacterial infections in the United States. During infection, C. difficile releases the gut-damaging toxins, TcdA and TcdB, the primary determinants of disease pathogenesis and are therefore therapeutic targets. TcdA and TcdB contain a glycosyltransferase domain that uses UDP-glucose to glycosylate host Rho GTPases, causing cytoskeletal changes that result in a loss of intestinal integrity. Isofagomine inhibits TcdA and TcdB as a mimic of the oxocarbenium ion transition state of the glycosyltransferase reaction. However, sequence variants of TcdA and TcdB across the clades of infective C. difficile continue to be identified and therefore, evaluation of isofagomine inhibition against multiple toxin variants are required. Here we show that Isofagomine inhibits the glycosyltransferase activity of multiple TcdB variants and also protects TcdB toxin-induced cell rounding of the most common full-length toxin variants. Further, isofagomine protects against C. difficile induced mortality in two murine models of C. difficile infection. Isofagomine treatment of mouse C. difficile infection permitted recovery of the gastrointestinal microbiota, an important barrier to prevent recurring C. difficile infection. The broad specificity of isofagomine supports its potential as a prophylactic to protect against C. difficile induced morbidity and mortality.

Reducing the Toxicity of Designer Aminoglycosides as Nonsense Mutation Readthrough Agents for Therapeutic Targets.

A significant proportion of genetic disease cases arise from truncation of proteins caused by premature termination codons. In eukaryotic cells some aminoglycosides cause readthrough of premature termination codons during protein translation. Inducing readthrough of these codons can potentially be of therapeutic value in the treatment of numerous genetic diseases. A significant drawback to the repeated use of aminoglycosides as treatments is the lack of balance between their readthrough efficacy and toxicity. The synthesis and biological testing of designer aminoglycoside compounds is documented herein. We disclose the implementation of a strategy to reduce cellular toxicity and maintain readthrough activity of a library of compounds by modification of the overall cationic charge of the aminoglycoside scaffold through ring I modifications.

Stereoselective Synthesis of the I-L Fragment of the Pacific Ciguatoxins.

The I-L ring system found in all the Pacific ciguatoxins has been prepared from a tricyclic precursor in a highly stereoselective manner. Subtle differences in the reactivity of the enones present in the seven- and eight-membered rings of the tricyclic ether starting material have been exploited to allow selective protection of the enone in the eight-membered ring. Subsequent distereoselective allylation of the seven-membered ring has been accomplished by a palladium-mediated Tsuji-Trost reaction. The K-ring methyl and hydroxyl groups have been installed in a highly stereoselective manner by sequential conjugate reduction and enolate oxidation reactions. Ring L has been constructed by a use of a novel relay ring-closing metathesis reaction to complete the tetracyclic framework, which possesses the functionality necessary for elaboration of rings I and L and the introduction of ring M.

Bidirectional Synthesis of the IJK Fragment of Ciguatoxin CTX3C by Sequential Double Ring-Closing Metathesis and Tsuji-Trost Allylation.

A novel four-step bidirectional strategy has been used to synthesize the IJK fragment of the marine polyether natural product CTX3C from a simple monocyclic precursor in a concise and efficient manner. The four-step bidirectional sequence involves ring-closing metathesis, alcohol oxidation, enol carbonate formation, and palladium-mediated Tsuji-Trost allylation.