Abstract
BackgroundOral thiopurines are effective and widely used in treatment of inflammatory bowel disease (IBD) in humans, although their use is limited due the development of adverse events. Here, we examine the efficacy and toxicity of oral treatment with 6-tioguanine (6-TG) and azathioprine (AZA) in a murine model of IBD.MethodsWe induced acute or chronic colitis in BALB/c mice by one or four cycles of 3% dextran sulphate sodium (DSS), respectively. Mice were treated by daily gavages of various dosages of 6-tioguanine, azathioprine, or by phosphate buffered saline (PBS) starting the first day of DSS or after two cycles of DSS, respectively. We monitored the efficacy and toxicity by measuring the weight change and serum alanine aminotransferase (ALT) activity and by disease severity and histology, at the end of the experiment. Moreover, we measured cytokine production after colon fragment cultivation by enzyme-linked immunoabsorbent assay and numbers of apoptotic cells in the spleen by flow cytometry.Results6-TG is effective in the treatment of acute DSS-induced colitis in a dose-dependent manner and 40 μg of 6-TG is significantly more effective in the treatment of acute colitis than both AZA and PBS. This effect is accompanied by decrease of IL-6 and IFN-γ production in colon. We did not observe histological abnormalities in liver samples from control (PBS) or 6-TG treated mice. However, liver samples from most mice treated with AZA showed mild, yet distinct signs of hepatotoxicity. In chronic colitis, all thiopurine derivatives improved colitis, 20 μg of 6-TG per dose was superior. High doses of 6-TG led to significant weight loss at the end of the therapy, but none of the thiopurine derivatives increased levels of serum ALT. Both thiopurine derivatives reduced the proportion of apoptotic T helper cells, but a high production of both IL-6 and TGF-β was observed only in colon of AZA-treated mice.ConclusionsUse of 6-TG in the treatment of experimental colitis in mice appears superior to AZA administration and placebo. In contrast to 6-TG, the use of AZA resulted in histological liver abnormalities.
Highlights
Oral thiopurines are effective and widely used in treatment of inflammatory bowel disease (IBD) in humans, their use is limited due the development of adverse events
We have shown that 6-TG is effective in the treatment of colitis in mice, in a dose dependent manner, and that a dosage of 40 μg 6-TG per day is superior to both AZA and placebo in acute experimental colitis therapy
Slight portobiliary mononuclear infiltration with or without pericholangitic fibrosis is an accidental finding in mice of the same breed, but damage to the adjacent reticulin was found only in mice that had AZA treatment. These results are in agreement with those of Petit et al, who showed that AZA and 6-MP decreases the viability of human hepatocytes even in clinically relevant concentrations when human treatment is considered, while 6-TG does not [24]. This difference in toxic effect among thiopurines could be explained by the production of toxic metabolites generated during the conversion of pro-drugs AZA and 6-MP to active metabolites 6-TGN, such as 6-methylmercaptopurines (6-MMP) which have been associated with liver test abnormalities [25,26,27]
Summary
Oral thiopurines are effective and widely used in treatment of inflammatory bowel disease (IBD) in humans, their use is limited due the development of adverse events. The immune-modulating thiopurines, 6-mercaptopurine (6-MP), and its pro-drug azathioprine (AZA), are widely used in inflammatory bowel diseases (IBD) treatment [1,2,3]. Both AZA and 6-MP require extensive metabolisation before the pharmacologicaly active metabolites, 6thioguaninenucleotides (6-TGN), are generated. Their mechanism of action is ascribed to both cytotoxic and apoptototic pathways. Activity of the specific metabolite, 6-thioguanine-triphosphate was recently found to contribute to the overall molecular immunosuppressive effect. This end-metabolite induces apoptosis and decreases the expression of proinflammatory molecules in activated T cells [4,5]
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