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Early Auto-processing of Bacterial Toxins as a Therapeutic Approach

Applicant Leroux Jean-Christophe
Number 141263
Funding scheme Project funding (Div. I-III)
Research institution Institut für Pharmazeutische Wissenschaften ETH Zürich
Institution of higher education ETH Zurich - ETHZ
Main discipline Organic Chemistry
Start/End 01.07.2012 - 31.10.2014
Approved amount 205'516.00
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Keywords (4)

Medicinal chemistry; Drug discovery; Bacterial toxins; Clostridium difficile

Lay Summary (English)

Lead
Lay summary

Bacteria rely on a host of different virulence factors to colonize the host and create pathogenesis. For some infections, therapeutic strategies targeting virulence factors such as toxins or adhesins have the advantage of not putting selective pressure on the organism, and thereby avoiding resistance problems. In recent years, the precise mechanism of pathogenesis and the crucial virulence factor for colonization of several important infections has been uncovered. This understanding provides fertile grounds for chemists to devise innovative strategies to prevent pathogenesis.

Clostridium difficile is a so-called “superbug” that is the cause of serious infections, especially in hospitalized patients. When the healthy gut flora of a patient is depleted, C. difficile can colonize the colon and cause a series of life-threatening diseases. Recently, emerging hypervirulent strains of C. difficile are causing more frequent and severe diseases. The current treatment for severe C. difficile infection (CDI) is vancomycin, but is plagued by high relapse rates and the threat of resistance against this last-resort antibiotic. A newly approved antibiotic for CDI, fidaxomicin, does not improve relapse rates in common hypervirulent strains. Therefore, there is a need for new therapy that would ideally not put selective pressure on the bacteria. Two toxins (TcdA, TcdB) secreted by the bacteria in the colon are responsible for the pathogenesis. Thus, there is an opportunity to manage the disease by targeting the toxins in the colon rather than killing the bacteria.

TcdA and TcdB are closely related large multi-domain proteins. The “warhead” of the toxin is a glucosyltransferase that inactivates important proteins in the cell cytosol. The remainder of the toxin is responsible for cell-binding, translocation and auto-processing, and is essential for the glucosyltransferase to reach the cytosol.

In this project we aim at interfering with the uptake mechanism of the "warhead" by using small molecules. This research could provide an important therapeutic alternative to antibiotics that would not be susceptible to resistance problems.

Direct link to Lay Summary Last update: 21.02.2013

Responsible applicant and co-applicants

Employees

Publications

Publication
A Chiral Phosphoramidite Reagent for the Synthesis of Inositol Phosphates.
Durantie Estelle, Huwiler Samuel, Leroux Jean-Christophe, Castagner Bastien (2016), A Chiral Phosphoramidite Reagent for the Synthesis of Inositol Phosphates., in Organic letters, 18(13), 3162-5.
New paradigms for the chiral synthesis of inositol phosphates.
Durantie Estelle, Leroux Jean-Christophe, Castagner Bastien (2015), New paradigms for the chiral synthesis of inositol phosphates., in Chembiochem : a European journal of chemical biology, 16(7), 1030-2.
Investigational New Treatments for Clostridium difficile Infection
Ivarsson Mattias, Leroux Jean-Christophe, Castagner Bastien, Investigational New Treatments for Clostridium difficile Infection, in Drug Discovery Today.

Collaboration

Group / person Country
Types of collaboration
Dr. Premysl Bercik, McMaster University Canada (North America)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Research Infrastructure

Scientific events

Active participation

Title Type of contribution Title of article or contribution Date Place Persons involved
Meakins-Christie Laboratory Individual talk Inhibition of a Clostridium difficile Toxin by Auto-Processing Triggers 10.11.2014 Montreal, Canada Castagner Bastien;
Young Faculty Meeting – Swiss Academy of Sciences Talk given at a conference Inhibition of a Clostridium difficile Toxin by Allosteric Triggers of Auto-Processing 05.06.2014 Bern, Switzerland Castagner Bastien;
Université Laval Individual talk Inhibition de Toxines Bactériennes - Comment déjouer une toxine 02.12.2013 Québec, Canada Castagner Bastien;
5th EMBO Meeting Talk given at a conference Small-molecule activators of Clostridium difficile toxin B auto-proteolysis 21.09.2013 Amsterdam, Netherlands Ivarsson Mattias;
McGill University Individual talk Inhibition of Bacterial Toxins - How to trick a toxin 19.09.2013 Montreal, Canada Castagner Bastien;
Université du Québec à Montréal Individual talk Inhibition of Bacterial Toxins - How to trick a toxin 11.09.2013 Montréal, Canada Castagner Bastien;
Queens University Individual talk Inhibition of Bacterial Toxins - How to trick a toxin 03.09.2013 Kingston, Canada Castagner Bastien;
The European Workshop on Bacterial Protein Toxins (ETOX) Poster Soluble Allosteric Triggers of Clostridium difficile Toxin B auto-processing 22.06.2013 Freiburg, Germany Castagner Bastien;
Max Planck Institute of Colloids and Interfaces Individual talk Confronting superbugs, or how to trick a toxin 27.02.2013 Golm, Germany Castagner Bastien;
Globalization of Pharmaceutics Education Network Meeting Talk given at a conference Toxin-targeted polymeric binders for the treatment of Clostridium difficile infection 28.11.2012 Melbourne, Australia Ivarsson Mattias; Castagner Bastien;
4th International Clostridium difficile Symposium Poster Alternatives to antibiotics for the treatment of Clostridium difficile infection 20.09.2012 Bled, Slovenia Castagner Bastien; Ivarsson Mattias;


Knowledge transfer events

Active participation

Title Type of contribution Date Place Persons involved
Falling Walls Lab Talk 08.11.2014 Berlin, Germany Ivarsson Mattias;
BIO Europe Talk 13.11.2012 Hamburg, Germany Ivarsson Mattias;
ETH Industry Day Poster 07.09.2012 Zurich, Switzerland Ivarsson Mattias;


Communication with the public

Communication Title Media Place Year
Media relations: print media, online media An innovative fight against the bacterium Clostridium difficile ETH Life Western Switzerland 2014
Media relations: radio, television Du nouveau contre le clostridium difficile RTS 1 Western Switzerland 2014
Media relations: print media, online media ETH doctoral student wins Zurich’s Falling Walls Lab ETH Life German-speaking Switzerland 2014
Media relations: print media, online media Von Darm, Charme und einer Mauer Tages Anzeiger German-speaking Switzerland 2014
New media (web, blogs, podcasts, news feeds etc.) Video décrivant le mécanisme des toxines de Clostridium difficile YouTube International 2013
Media relations: print media, online media Toxine statt Bakterien bekämpfen ETH Life Western Switzerland 2012

Awards

Title Year
Jury and audience prize of the Falling Walls Lab Zurich competition 2014
Venture Leaders 2014, VentureLab & Swiss CTI (Business development training course in Boston & New York for top 20 promising entrepreneurs in Switzerland) 2014
EMBO travel grant, The EMBO Meeting, Amsterdam, Netherlands 2013

Abstract

Clostridium difficile is a bacterium that causes serious nosocomial infection in patients under antibiotic treatment, because this so-called “superbug” is resistant to a number of broad-spectrum antibiotics. When the healthy gut flora of a patient is depleted, C. difficile can colonize the colon and cause a series of life-threatening diseases. Recently emerging hypervirulent strains of C. difficile are causing more frequent and severe diseases. The current treatment for severe C. difficile infection (CDI) is vancomycin, but is plagued by high relapse rates and the threat of resistance against this last-resort antibiotic. A newly approved antibiotic for CDI, fidaxomicin, does not improve relapse rates in common hypervirulent strains. Therefore, there is a need for new therapy that would ideally not put selective pressure on the bacteria. Two toxins (TcdA, TcdB) secreted by the bacteria in the colon are responsible for the pathogenesis. Therefore, there is an opportunity to manage the disease by targeting the toxins in the colon rather than killing the bacteria. TcdA and TcdB are closely related large multi-domain proteins. The “warhead” of the toxin is a glucosyltransferase that inactivates important proteins in the cell cytosol. The rest of the toxin is responsible for cell-binding, translocation and auto-processing, and is essential for the glucosyltransferase to reach the cytosol. An auto-processing step triggered by the intracellular co-factor inositol hexakisphosphate (IP6) is required to release the glucosyltransferase in the cytosol. The cysteine protease domain (CPD) responsible for this cleavage binds to IP6, which then stabilizes the active conformation of the enzyme and results in cleavage.Since the glucosyltransferase itself is innocuous to cells because it is incapable of reaching the cytosol, we propose to trigger prematurely the auto-processing of the toxin in the colon lumen, and therefore inactivate the toxin. This constitutes a novel and unexplored strategy for targeting bacterial toxins. IP6 is a strictly intracellular co-factor because of its insolubility in the high concentrations of calcium found outside the cells. Therefore, IP6 itself cannot be used for this purpose. We propose to synthesize analogs of IP6 that are soluble in the presence of calcium and retain the ability to induce the toxin auto-cleavage. The specific aims of this proposal are:Aim 1: The design, synthesis, and characterization of IP6 analogs soluble in high calcium concentrations.Aim 2: The in vitro evaluation of the candidates’ activity against the CPD of the toxin. The results of this aim will inform the design of further analogs in a feedback loop.Aim 3: Further in vitro evaluation of selected candidates will be performed. In addition, the candidates will be evaluated in a cytotoxicity inhibition assay in cell cultures. The compounds’ toxicity to colon cancer cells will also be evaluated.
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