Local prevention of oxidative stress in the intestinal epithelium of the rat by adhesive liposomes of superoxide dismutase and tempamine.

The purpose of the present study was to investigate whether the local prevention of luminal superoxide-mediated biological damage in the rat jejunal mucosa could be achieved by liposomal superoxide dismutase (SOD) and the SOD mimic tempamine (TMN). Cationic liposomes loaded with either SOD or TMN were perfused in the rat jejunum prior to the induction of oxidative insult. Reactive hydroxyl radicals were generated in situ in a closed circulating intestinal loop of the rat from the reaction between hypoxanthine and xanthine oxidase in the presence of chelated ferrous sulfate. Mucosal activity of lactate dehydrogenase and levels of potassium ions were used to quantify the tissue damage. Intracellular uptake and locality of SOD were examined in HT-29 cells. The intestinal uptake of SOD and TMN was further measured by using rat colon sacs. Entrapment in cationic liposomes was found to significantly enhance the antioxidant effect of SOD and TMN against the induced oxidative damage in the jejunal mucosa, compared with their free forms. The effect was found to be local and was caused by the increased mucosal adhesion of the liposomes. The cationic liposomes also triggered SOD uptake into the HT-29 cell line. It is concluded that the increased residence time of the cationic liposomes of SOD and TMN in the jejunal mucosa resulted in a local effect against oxidative injury. This local protection may be exploited for drug delivery purposes.

Improving cancer therapy through p53 management.

The tumor suppressor p53 normally acts to appropriately coordinate cellular responses to stress stimuli. When p53 activity is disabled, the onset of malignancy is a potential consequence. Engendering wild type p53 activities in cells that lack these functions is an approach that is currently being explored for cancer therapy. Eliciting elevated levels of active p53, imparting p53 activities through gene therapy, compelling mutant p53 to perform normal functions, manipulating p53 regulators, and activating p53 effectors are all approaches that are currently being developed. In this review we will provide a synopsis of the most promising 'p53-based' strategies for fighting cancer, both those under clinical trial and recent innovative concepts.

Manipulation of the tumor suppressor p53 for potentiating cancer therapy.

The tumor suppressor p53 is a linchpin in the regulation of appropriate cellular responses to various stress conditions. Inactivation of the functions of this critical participant can have diabolical consequences, in particular the development of malignant diseases. Elicitation of appropriate p53 functions is an attractive strategy for combating cancer. Triggering p53 responses, reconstituting p53 activities through gene therapy, coercing mutant p53 to perform normal functions, manipulating p53 regulators, and activating p53 effectors are all approaches that are currently being developed. Here, we will overview 'p53-based' strategies for fighting cancer, both those under clinical trial and recent innovative concepts.

Apoptosis - the p53 network.

Exposure to cellular stress can trigger the p53 tumor suppressor, a sequence-specific transcription factor, to induce cell growth arrest or apoptosis. The choice between these cellular responses is influenced by many factors, including the type of cell and stress, and the action of p53 co-activators. p53 stimulates a wide network of signals that act through two major apoptotic pathways. The extrinsic, death receptor pathway triggers the activation of a caspase cascade, and the intrinsic, mitochondrial pathway shifts the balance in the Bcl-2 family towards the pro-apoptotic members, promoting the formation of the apoptosome, and consequently caspase-mediated apoptosis. The impact of these two apoptotic pathways may be enhanced when they converge through Bid, which is a p53 target. The majority of these apoptotic effects are mediated through the induction of specific apoptotic target genes. However, p53 can also promote apoptosis by a transcription-independent mechanism under certain conditions. Thus, a multitude of mechanisms are employed by p53 to ensure efficient induction of apoptosis in a stage-, tissue- and stress-signal-specific manner. Manipulation of the apoptotic functions of p53 constitutes an attractive target for cancer therapy.

P53 licensed to kill? Operating the assassin.

The p53 protein is a key player in the cellular response to stress. Proper regulation of p53 is imperative for the suppression of tumor development. This regulation is largely governed by its master inhibitor, Mdm2, which both blocks p53 activities and promotes its destabilization. This tight regulation of p53 by Mdm2 must be interrupted under stress conditions in order for p53 to be stabilized in an active form. A combined action of partner proteins and modifying enzymes is essential for the relief of p53 from Mdm2. The recent revelation of p53 association with the PML-nuclear bodies provides one explanation of how this regulatory network is coordinated within the nucleus in response to certain stress conditions. Thus, it is not only the nature of the p53 regulatory complex but also the spatial and temporal context of this association that governs the output inhibitory signals mediated by p53.

Regional peptide uptake study in the rat intestinal mucosa: glatiramer acetate as a model drug.

PURPOSE: To identify regions of the rat intestine that are able to internalize from the lumen oligopeptides, using the model drug glatiramer acetate (GA). METHODS: GA was introduced into rat intestinal sacs and the integrity of GA during uptake was monitored using antibody detection. Sodium docecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting of intestinal homogenates that had been exposed to GA were performed to identify GA presence. An enzyme-linked immunosorbent assay (ELISA) protocol was adapted for GA quantification. Immunohistochemistry was undertaken to examine the rat colonic wall for GA uptake, and confocal microscopy was used to differentiate adsorbed and internalized peptide in cultured colorectal adenocarcinoma cells. RESULTS: The colon and the ileum, respectively, were identified to be the intestinal regions in which GA was maximally preserved during uptake from the lumen. GA was identified to cross the colonic wall from the epithelium to the serosa. Internalization of GA into cultured colonic epithelial cells was demonstrated. CONCLUSIONS: The rat colonic wall was identified to be less proteolytically active toward GA compared to the wall of the more proximal regions of the small intestine. GA has the capacity to penetrate from the lumen into the colonic wall. The maintenance of GA integrity within the wall of the colon offers the potential for local biological activity of the drug.

The colon as a possible target for orally administered peptide and protein drugs.

Oral administration offers a potential portal to the superficial layers of the gastrointestinal (GI) tract (local delivery) and to the blood and lymphatics (systemic delivery). The harsh hydrolytic environment of the GI tract and the epithelial barriers to absorption, however, pose major challenges to the success of this mode of drug delivery for peptide and protein drugs. One approach to minimizing enzymatic degradation in the GI tract is to target drugs to the apparently less proteolytically active colon. In this review, the evidence supporting the colon as an attractive site for peptide and protein drug delivery will be discussed. The discussion will be confined to specific examples of delivery systems bearing both peptides and proteins that have been tested in the colonic context.