Behind the Scenes: Azathioprine in IBD

256px-Azathioprine-structureFor this post, I decided to take a little break from the current literature and discuss something that I’ve wanted to look at for quite some time: the immunological mechanisms of azathioprine. Azathioprine is one of the first line drugs for IBD and appears to work via T cells; interfering with proliferation and causing apoptosis during activation.

 Main points:

  • Azathioprine interferes with lymphocyte proliferation by blocking nucleotide synthesis.
  • Azathioprine encourages apoptosis during T cell activation.

Take home message: Azathioprine appears to have its strongest effects on T cells.

Azathioprine, also well known under the trade name, Imuran, is one of the mainstay drugs in IBD treatment. It has a long history, having been developed in 1957 by the Nobel Prize winner, Gertrude B. Elion and George Hitchings.

It is a pro-drug of the molecule, 6-MP, which is easily metabolized into three other molecules: 6-TU, which is inactive; 6-MMP, which is also inactive and 6-TIMP, which is further metabolized to the active molecules, TIMP, MeTIMP and TGTP/TdGTP. MeTIMP is a purine synthesis inhibitor. This means that it can block the enzymes that are needed to produce purine nucleotides (adenine and guanine). Nucleotides are necessary for the production of DNA during cell division. T and B cells are particularly dependent on this process as they lack to the ability to form nucleotides from nucleotide degradation products in a process called “nucleotide salvage”. Thus, azathioprine has the ability to put a damper on the proliferation of T and B cells, the strong men of the adaptive immune response.

For many years, this was considered THE function of azathioprine. But, in the last ten years, more mechanisms surfaced. It is now known that the metabolites TIMP and TdGTP interfere with DNA synthesis and TGTP is actually incorporated into RNA during transcription. However, a more interesting immunological finding is that the metabolite TGTP also interacts with GTP-binding protein, Rac1. Rac1 has a multitude of effects and also works downstream of the CD28 signaling cascade.

This phenomenon was described in the Journal of Clinical Investigation in 2003. The authors initially determined that activation of human T cells in combination with azathioprine or the metabolitie 6-MP led to more apoptotic cells. An examination of apoptotic mononuclear cells in azathioprine-treated IBD patient colon samples showed that the number of apoptotic cells were increased 2-3 fold as compared to untreated patients. It was ultimately found that azathioprine blocked the Rac1/MEC pathway during CD28 stimulation. CD28 stimulation is essential for full T cell activation and the Rac1 pathway helps block apoptosis during this period. By blocking Rac1 function during CD28 activation, azathioprine delivers a deadly punch just when the T cells are getting ready to gear up for battle.

A more recent article in Immunology Letters looking at other co-stimulatory receptors found that the effects of azathioprine might not be limited to CD28 alone. They saw reductions in in vitro T cell proliferation after co-stimulating cells via CD2, LFA-1, ICOS or 4-1BB. It could be that Rac1 or another GTP-ase is involved in all of these co-stimulatory pathways.

After mucking around in the details, it’s nice to take some distance and look at the bigger picture. The main point is that azathioprine appears to have the strongest influence on activated T cells either via the blockage of proliferation or the induction of apoptosis. This also appears to be the case for another popular drug for IBD, anti-TNFα, which has been shown to lead to both apoptosis and a reduction of T cell proliferation. It would really seem that T cells are at the crux of IBD.

In my December 7th blog post, I talked about memory T cells and their interesting relationship to mesenchymal stem cells that produced IL-7, a factor needed for their survival. The articles discussed really sent home the message that memory T cells are extremely important to the development of colitis in mice. I suspect that it is during the restimulation of memory T cells where azathioprine has its strongest effect.

References

Bradford, K. (2011). Optimizing 6-mercaptopurine and azathioprine therapy in the management of inflammatory bowel disease. World Journal of Gastroenterology, 17(37), 4166. doi:10.3748/wjg.v17.i37.4166

Leitner, J., Drobits, K., Pickl, W. F., Majdic, O., Zlabinger, G., & Steinberger, P. (2011). The effects of Cyclosporine A and azathioprine on human T cells activated by different costimulatory signals. Immunology Letters, 140(1-2), 74–80. doi:10.1016/j.imlet.2011.06.010

Tiede, I., Fritz, G., Strand, S., Poppe, D., Dvorsky, R., Strand, D., et al. (2003). CD28-dependent Rac1 activation is the molecular target of azathioprine in primary human CD4+ T lymphocytes. The Journal of clinical investigation, 111(8), 1133–1145. doi:10.1172/JCI200316432

 

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