In vivo correction of genetic defects of monocyte/ macrophages using attenuated Salmonella as oral vectors for targeted gene delivery

Paglia, Paola; Terrazzini, Nadia; Schulze, Kai; Guzmán, Carlos A.; Colombo, Mario P.

doi:10.1038/sj.gt.3301290

Cited by 52 publications

(44 citation statements)

References 31 publications

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“…Administration to genetically immunodeficient mice of avirulent Salmonella bearing murine IFN-g gene resulted in transgene expression within macrophages and dendritic cells, correction of the genetic defect, and restoration of the cellular phenotype. 32,33 Invasive bacteria can efficiently transfer genes into nondividing differentiated cells, including normal epithelial cells layering the intestinal mucosa, thus offering an alternative to viral vectors.…”

Section: Discussionmentioning

confidence: 99%

Engineered E. coli delivers therapeutic genes to the colonic mucosa

et al. 2005

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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 pGB2Oinv-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 (pC1OTGF-b1) 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 pC1OTGF-b1 by the inflammation-inducible interleukin-8 promoter generating plasmid pC1OTGF-b1 IND . Plasmid-specific TGF-b1 mRNA transcripts were detectable in mouse CMT-93 epithelial cells incubated with E. coli BM2710/pGB2Oinv-hly carrying pC1OTGF-b1 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/ pGB2Oinv-hly+pC1OTGF-b1. 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.

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Section: Discussionmentioning

confidence: 99%

Engineered E. coli delivers therapeutic genes to the colonic mucosa

et al. 2005

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“…SL101 was derived from auxotrophic strain SL7207 (15) and, in addition, contained a deletion of ssrA/B. SL7207 is attenuated in virulence and pathogenesis in vivo and has been shown to function as a gene delivery carrier for the expression of several transgenes in mammalian cells (16,17,19). SsrA/B regulates the expression of Salmonella Pathogenicity Island-2 genes, which are important for Salmonella intracellular survival in macrophages and virulence in vivo (20).…”

Section: Resultsmentioning

confidence: 99%

“…Thus, Salmonella may represent a promising gene delivery agent for gene therapy. Macrophages represent the major in vivo reservoir for Salmonella following their systemic dissemination and are therefore considered an optimal target for any Salmonellabased gene therapy (13,16). However, it has not been reported whether Salmonella can efficiently deliver ribozymes, such as RNase P ribozymes, for expression in animals.…”

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confidence: 99%

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Oral delivery of RNase P ribozymes by Salmonella inhibits viral infection in mice

Bai¹,

Gong

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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Safe, effective, and tissue-specific delivery is a central issue for the therapeutic application of nucleic-acid-based gene interfering agents, such as ribozymes and siRNAs. In this study, we constructed a functional RNase P-based ribozyme (M1GS RNA) that targets the overlapping mRNA region of M80.5 and protease, two murine cytomegalovirus (MCMV) proteins essential for viral replication. In addition, a novel attenuated strain of Salmonella, which exhibited efficient gene transfer activity and little cytotoxicity and pathogenicity in mice, was constructed and used for delivery of anti-MCMV ribozyme. In MCMV-infected macrophages treated with the constructed attenuated Salmonella strain carrying the functional M1GS RNA construct, we observed an 80-85% reduction in the expression of M80.5/protease and a 2,500-fold reduction in viral growth. Oral inoculation of the attenuated Salmonella strain in mice efficiently delivered antiviral M1GS RNA into spleens and livers, leading to substantial expression of the ribozyme without causing significant adverse effects in the animals. Furthermore, the MCMV-infected mice that were treated orally with Salmonella carrying the functional M1GS sequence displayed reduced viral gene expression, decreased viral titers, and improved survival compared to the untreated mice or mice treated with Salmonella containing control ribozyme sequences. Our results provide direct evidence that oral delivery of M1GS RNA by Salmonella-based vectors effectively inhibits viral gene expression and replication in mice. Moreover, this study demonstrates the utility of Salmonellamediated oral delivery of RNase P ribozyme for gene-targeting applications in vivo.gene therapy | herpesvirus | gene delivery | antisense | animal model

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“…Attenuated Salmonella strains have recently been shown to function as a carrier system for delivery of nucleic acid-based vaccines and antitumor small hairpin RNAs (shRNAs) for cancer therapy (10)(11)(12)(13). In these studies, plasmid constructs, which contained the transgenes under the control of an eukaryotic expression promoter, were introduced to the attenuated bacterial strains.…”

mentioning

confidence: 99%

Salmonella -mediated delivery of RNase P-based ribozymes for inhibition of viral gene expression and replication in human cells

Bai¹,

Vu³

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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A fundamental challenge in gene therapy is to develop approaches for delivering nucleic acid-based gene interfering agents, such as small interfering RNAs and ribozymes, to the appropriate cells in a way that is tissue/cell specific, efficient, and safe. Using human cytomegalovirus (HCMV) infection of differentiated macrophages as the model, we showed that Salmonella can efficiently deliver RNase P-based ribozyme sequence in specific human cells, leading to substantial ribozyme expression and effective inhibition of viral infection. We constructed a functional RNase P ribozyme (M1GS RNA) that targets the overlapping mRNA region of two HCMV capsid proteins, the capsid scaffolding protein (CSP) and assemblin, which are essential for viral capsid formation. Substantial expression of ribozymes was observed in human differentiated macrophages that were treated with attenuated Salmonella strains carrying the ribozyme sequence constructs. A reduction of 87-90% in viral CSP expression and a reduction of about 5,000-fold in viral growth were observed in cells that were treated with Salmonella carrying the sequence of the functional ribozyme but not with those carrying the sequence of a control ribozyme that contained mutations abolishing the catalytic activity. To our knowledge, this study showed for the first time that ribozymes expressed following targeted gene transfer with Salmonella-based vectors are highly active and specific in blocking viral infection. Moreover, these results demonstrate the feasibility to develop Salmonella-mediated gene transfer of RNase P ribozymes as an effective approach for gene-targeting applications.antisense | gene targeting | antiviral | gene delivery | RNA cleavage

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In vivo correction of genetic defects of monocyte/ macrophages using attenuated Salmonella as oral vectors for targeted gene delivery

Cited by 52 publications

References 31 publications

Engineered E. coli delivers therapeutic genes to the colonic mucosa

Engineered E. coli delivers therapeutic genes to the colonic mucosa

Oral delivery of RNase P ribozymes by Salmonella inhibits viral infection in mice

Salmonella -mediated delivery of RNase P-based ribozymes for inhibition of viral gene expression and replication in human cells

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