Scientific Abstract

Proposal No.  IBD-0027R
Principal Investigator: Eugene B. Chang, M.D.
Applicant Organization: The University of Chicago (Illinois, U.S.A.)
Project Title:  Identification of probiotic-derived cytoprotective and anti-inflammatory proteins:  novel agents for treatment of inflammatory bowel diseases
Period of Award:  November 1, 2002 - October 31, 2003
 
Colonic bacterial flora appear to play a major role in the development of experimental colitis and Crohn’s disease.  The combination of genetic background, exposure, or colonization by certain inciting commensal bacteria could result in the development of inflammatory bowel disease (IBD) in susceptible individuals. Because little can be done presently to correct genetic susceptibility, changing the gut flora of IBD patients with probiotic agents is being intensely studied as a therapeutic strategy.  Probiotics are living organisms in food and dietary supplements that have beneficial health effects beyond their inherent nutritive value.  These agents have been shown to be effective in either preventing or mitigating the intestinal mucosal inflammation associated with experimental colitis.  A number of preliminary clinical trials have also shown that probiotics are effective in the treatment of pouchitis and IBD.

The mechanism(s) of probiotic action, however, remains unclear.  It has been assumed that colonization by probiotics is a necessary first step to alter enteric flora, thereby reducing or blocking their adverse effects.  Arguing against this hypothesis are findings that probiotic agents and their soluble extracts have direct and rapid effects in promoting intestinal epithelial barrier function and viability, as well as in protecting cells against injury induced by a variety of pathogenic conditions and agents

Our studies are focused on the examination of the beneficial effects of the probiotic formulations such as VSL#3 (composed of many strains, including Bifidobacteria, Lactobacillus and Streptococcus) and other native enteric flora (e.g., Bacteroides fragilis).  At least two mechanisms underlying their potential effects have been identified:  (1) induction of two cytoprotective heat shock proteins, hsp25 and hsp72, and (2) inhibition of the pro-inflammatory NF-kappaB pathway through stabilization of phospho-IkappaB.  While these findings are interesting, this proposal will, instead, focus on the identification and isolation of the active factor(s) of VSL#3 and other enteric organisms, as these studies will have immediate relevance to the treatment of IBD. We will first isolate specific bacterial strains of VSL#3 and other reference bacterial strains that have probiotic activity.  Several soluble, heat-labile proteins secreted by these organisms of less than 100kD size have already been partially isolated.  Standard biochemical approaches (sizing, affinity, hydrophobicity, etc.) will be used to identify, purify and concentrate the active factors of the probiotic supernatant.  Two read-outs, hsp induction and inhibition of NF-kappaB, will be used to achieve accuracy and high throughput screening.

Identification of bioactive probiotic proteins will be a major step forward in eventual harvest, concentration and therapeutic application of these agents.  We believe this approach has a significant advantage over current probiotic practice where efficacy may be limited by the need for bacterial colonization, by variations and unregulated composition of formulations and by homeopathic delivery of active agents.  These investigations can therefore lead to the development of new types of therapeutic agents that will benefit patients with IBD.