Engineered E coli delivers therapeutic genes to the colonic mucosa

Taking advantage of the proximity of bowel mucosa to luminal bacteria, we have attempetd 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 pGB2 Omega inv-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 (pC1 Omega TGF-beta 1) 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 pC1 Omega TGF-beta 1 by the inflammation-inducible interleukin-8 promoter generating plasmid pC1 Omega TGF-beta 1(IND). Plasmid-specific TGF-beta 1 mRNA transcripts were detectable in mouse CMT-93 epithelial cells incubated with E. coli BM2710/pGB2 Omega inv-hly carrying pC1 Omega TGF-beta 1(IND) 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/ pGB2 Omega inv-hly+pC1 Omega TGF-beta 1. 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.