Development of Heptylmannoside-Based Glycoconjugate Antiadhesive Compounds against Adherent-Invasive Escherichia coli Bacteria Associated with Crohn's Disease
Authors: Sivignon Adeline , Yan Xibo , Alvarez Dorta Dimitri , Bonnet Richard , Bouckaert Julie , Fleury Etienne , Bernard Julien , Gouin Sébastien G. , Darfeuille-Michaud Arlette , Barnich Nicolas 
Published: November 2015

The ileal lesions of Crohn's disease (CD) patients are colonized by adherent-invasive
Escherichia coli
(AIEC) bacteria. These bacteria adhere to mannose residues expressed by CEACAM6 on host cells in a type 1 pilus-dependent manner. In this study, we investigated different antagonists of FimH, the adhesin of type 1 pili, for their ability to block AIEC adhesion to intestinal epithelial cells (IEC). Monovalent and multivalent derivatives of
n
-heptyl α-
d-
mannoside (HM), a nanomolar antagonist of FimH, were tested
in vitro
in IEC infected with the AIEC LF82 strain and
in vivo
by oral administration to CEACAM6-expressing mice infected with LF82 bacteria.
In vitro
, multivalent derivatives were more potent than the monovalent derivatives, with a gain of efficacy superior to their valencies, probably owing to their ability to form bacterial aggregates. Of note, HM and the multi-HM glycoconjugates exhibited lower efficacy
in vivo
in decreasing LF82 gut colonization. Interestingly, HM analogues functionalized with an isopropylamide (1A-HM) or β-cyclodextrin pharmacophore at the end of the heptyl tail (1CD-HM) exerted beneficial effects
in vivo
. These two compounds strongly decreased the amount of LF82 bacteria in the feces of mice and that of bacteria associated with the gut mucosa when administered orally at a dose of 10 mg/kg of body weight after infection. Importantly, signs of colitis and intestinal inflammation induced by LF82 infection were also prevented. These results highlight the potential of the antiadhesive compounds to treat CD patients abnormally colonized by AIEC bacteria and point to an alternative to the current approach focusing on blocking proinflammatory mediators.

IMPORTANCE
Current treatments for Crohn's disease (CD), including immunosuppressive agents, anti-tumor necrosis factor alpha (anti-TNF-α) and anti-integrin antibodies, focus on the symptoms but not on the cause of the disease. Adherent-invasive
Escherichia coli
(AIEC) bacteria abnormally colonize the ileal mucosa of CD patients via the interaction of the mannose-specific adhesin FimH of type 1 pili with CEACAM6 mannosylated proteins expressed on the epithelial cell surface. Thus, we decided to develop an antiadhesive strategy based on synthetic FimH antagonists specifically targeting AIEC bacteria that would decrease intestinal inflammation. Heptylmannoside (HM)-based glycocompounds strongly inhibit AIEC adhesion to intestinal epithelial cells
in vitro
. The antiadhesive effect of two of these compounds of relatively simple chemical structure was also observed
in vivo
in AIEC-infected CEACAM6-expressing mice and was associated with a reduction in the signs of colitis. These results suggest a new therapeutic approach for CD patients colonized by AIEC bacteria, based on the development of synthetic FimH antagonists.