Engineered enteric bacteria to deliver therapeutic genes to the colonic mucosa

Taking advantage of the proximity of bowel mucosa to luminal bacteria, we have attempted to deliver a therapeutic gene to the colonic mucosa by oral administration of an invasive and non-pathogenic Escherichia coli. E. coli diamenopimelate (dap) auxotroph, harboring plasmid pGB2Omegainv-hly, express the inv gene from Yersinia pseudotubercolosis that confers the ability to invade nonprofessional phagocytic cells and the hly gene from Listeria monocytogenes that allows expression of lystreriolysin O, a perforin cytolysin able to perfore phagosomal membranes. This bacterial vector invades and transfers functional DNA to epithelial cells in vitro. We have shown that this strain carrying a therapeutic gene (pC1OmegaTGF-beta1) can significantly reduce the severity of experimental colitis in mice. However, as a consequence of mucosal barrier disruption during colitis, vector-specific mRNA transcripts could be recovered from the colon and also from extra-colonic tissues. We therefore replaced the constitutive CMV promoter in pC1OmegaTGF-beta1 by the inflammation-inducible interleukin-8 promoter generating plasmid pC1OmegaTGF-beta1IND. Plasmid-specific TGF-beta1 mRNA transcripts were detectable in mouse CMT-93 epithelial cells incubated with E. coli BM2710/pGB2Omegainv-hly carrying pC1OmegaTGF-beta1IND following exposure to inflammatory cytokines. Furthermore, the transcripts were detectable only within inflamed tissues and the therapeutic effects were comparable to those in animals treated with E. coli BM2710/pGB2Omegainv-hly+pC1OmegaTGF-beta1. In summary, engineered enteric bacteria can efficiently deliver in vivo therapeutic genes to the intact intestinal mucosa and regulation expression of the therapeutic gene by an inflammation-inducible promoter prevents its dissemination during colitis.